19th annual upstate new york immunology conference · keynote speakers are dr. wayne m. yokoyama...

19th Annual Upstate New York

Immunology Conference

Major Corporate Sponsors:

BD Biosciences

BioLegend, Inc.

Keynote Speakers:

Wayne M. Yokoyama, M.D.

Thomas J. Braciale, M.D., Ph.D.

Workshops by:

Dr. Yokoyama, Dr. Braciale, and Members of NIH

Panel Discussion:

Careers Outside of Academia

AAI Young Investigator Awards

eBioscience/affymetrix Trainee Travel Awards

Welcome to

The Sagamore Resort

and Conference Center

Bolton Landing, NY

October 24-27, 2016

Supported by NIH/NIAID

Grant Funding

3.

Conference and Venue ............................................................................... 5

Schedule of Events .................................................................................... 6

Memories from 2015 .................................................................................. 15

Platinum Corporate Sponsors

BD Biosciences ..................................................................................... 16

BioLegend, Inc. .................................................................................... 18

Silver Plus Corporate Sponsors

eBioscience/affymetrix .......................................................................... 20

Krackeler Scientific .............................................................................. 22

Silver Corporate Sponsors .......................................................................... 24

Agilent Technologies ............................................................................ 25

EMD Millipore ...................................................................................... 26

Shenandoah ......................................................................................... 27

ThermoFisher ....................................................................................... 28

Bronze Corporate Sponsors ........................................................................ 29

American Association of Immunologists ....................................................... 30

NYIC Scientific Advisory Board .................................................................... 31

Institutional Financial Supporters ................................................................ 32

Grant Support ........................................................................................... 33

Keynote Speaker—Sponsored by BD Biosciences

Wayne M. Yokoyama, M.D. ................................................................... 34

Symposium I: Current Topics in Immunoregulation ...................................... 35

Symposium II: Neonatal Immunity .............................................................. 39

Oral Poster Presentations

Session A: Lymphocyte Immunobiology I ............................................... 42

Session B: Tumor Immunology .............................................................. 48

Table of Contents

4.

Corporate Presentation: BioLegend, Inc. ...................................................... 54


Session C: Lymphocyte Immunobiology II .............................................. 55

Session D: Immunoregulation ................................................................ 61

Workshop I - Dr. Wayne Yokoyama

“Opportunities to Study the Immunology of Human Diseases” ................... 67

Corporate Presentation: BD Biosciences ....................................................... 68

Symposium III: Cancer Immunity ............................................................... 69

Symposium IV: Immune Mediated Disease .................................................. 72

Industry Panel

“Careers Outside of Academia” ............................................................. 75

Workshop II - Dr. Thomas Braciale

“The Business End of Academic Research” .............................................. 76

Workshop III—NIH Members

“Grantsmanship, Funding, and Mock Peer Review” ................................. 77

Symposium V: Immunity in Infectious Disease ............................................. 78

Keynote Speaker—Sponsored by BioLegend

Dr. Thomas J. Braciale .......................................................................... 82

Poster Listing ............................................................................................ 83

Poster Abstracts ........................................................................................ 84

Attendee Contact Information ..................................................................... 133

Author Index ............................................................................................. 139

5.

UPSTATE NEW YORK IMMUNOLOGY CONFERENCE (NYIC)

We’ve come a long way from Garnet Hill! This meeting started in 1997 as a small retreat to facilitate

interactions among young scientists, institutions, and renowned experts in the field of Immunology. In

just a few short years, the number of attendees grew and a larger venue was needed to meet the future

needs of the Conference.

We are happy to announce the American Association of Immu-

nologists (AAI) is once again providing 10 Young Investigator

Awards. eBioscience/affymetrix is also proving 10 Trainee Travel

Awards. All award winners will give Oral Poster Presentations.

There will also be two Workshops. Keynote speakers are Dr. Wayne M. Yokoyama

(Washington University) and Dr. Thomas J. Braciale (University of Virginia). As part of

our leisure activities, there will be a cruise on Lake George, as well as a recreational night including min-

iature golf, Wii, X-box, whiffle ball, and movies! Caldwell’s and Mr. Brown’s Pub will be open Wednes-

day night for informal discussions. Trainees will also have an opportunity to win an iPad during one of

two drawings. You must be present at the drawing to win!

While all these elements lend to the atmosphere, one simple principle goal of this Conference re-

mains. To provide an opportunity for young and senior scientists to gather in a setting that is diverse

enough to meet the needs of all attendees while remaining small enough to allow for personal interac-

tions. While always challenging, it is the goal of the NYIC Scientific Advisory Board and the NYIC Confer-

ence Organizers to give graduate students and postdoctoral fellows the opportunity to present their re-

search and engage in conversations that will stimulate further discussions, collaborations, and interest in

pursuing a new or different way of looking at their research.

We hope you share our enthusiasm and enjoy your time with us!

THE SAGAMORE RESORT

The Sagamore Resort and Conference Cen-

ter celebrated it’s 125 year anniversary in

2008. Since that time, the resort has gone

through some remarkable renovations. If this is

your first visit to the Resort, take some time to

enjoy the beauty that surrounds you. There

are many breath-taking views to be seen.

The staff are friendly, courteous, and hard-working. If you require any information or have a special

need, please see either Dawn Bellville, Administrative Coordinator for NYIC, or any of the Resort person-

nel. Many thanks to Lori Rehm (Director of Sales), Shelly Yeager (National Sales Manager), Derrick Ham-

mond (Conference Services Manager), Don Vilmar (Banquet Manager), Joel Clark (Function Set-up Man-

ager) and his amazing crew, CMI Communications-Audio/Visual Technician, Glen, along with all of the

associates who attend to our many needs. Thank you!

6.

Upstate New York Immunology Conference

Schedule of Events

Monday, October 24th

3:00-5:00 p.m. Hotel Check-in (Main Hotel Lobby) 3:00-5:00 p.m. Conference Registration (Albenia—Conference Center) 5:00-6:00 p.m. Welcome Reception (Conference Center Lobby) 6:00-6:30 p.m. Dinner (Bellvue) 6:30 p.m. Welcome and Introductions

Keynote Presentation

Sponsored by BD Biosciences Introduction: Dr. Dennis W. Metzger

Wayne M. Yokoyama, M.D. Levin Professor of Medicine Investigator, Howard Hughes Medical Institute Washington University

“Tissue-resident Natural Killer Cells” 7:30 p.m. American Association of Immunologists Young Investigator Awards and eBioscience Travel Awards (Award Presentations & Photos) Immediately Following Awards Presentation—Meet and Greet Activities

(Recreation Center)

7.

Tuesday, October 25th 7:00-8:15 a.m. Breakfast at Leisure (Use voucher towards total cost) (La Bella Vita)

8:25-8:30 a.m. Morning Announcements (Nirvana)

8:30-10:00 a.m. Symposium I: Current Topics in Immunoregulation (Nirvana) Chair: Dr. Brent Berwin 8:30-9:00 Michael A. Lynes, Ph.D. (University of Connecticut)

“The Stress Protein Metallothionein: a Small Protein with a Large Immunomodulatory Role”

9:00-9:30 Qi Yang, Ph.D. (Albany Medical College) “Biology of Inflammatory Innate Lymphoid Cells” 9:30-10:00 Karin Schneider, Ph.D. (SUNY Upstate)

“Influence of Colony Stimulating Factors on Virus Infections of Monocyte Lineage Cells”

10:00-10:15 a.m. Break and Display Posters (Conference Center Foyer/Bellvue) 10:15-11:15 p.m. Symposium II: Neonatal Immunity (Nirvana) Chair: Dr. Gary Winslow 10:15-10:45 Brain Rudd, Ph.D. (Cornell University) “The Fate of Neonatal and Adult CD8+ T cells During Infection is Linked to Their Developmental Origin” 10:45-11:15 David A. Lawrence, Ph.D. (Wadsworth/NYSDOH)

“Maternal-placental-fetal Interactions Affecting Offspring Im-munity and Behavior”

11:15-11:30 a.m. Break and Display Posters (Conference Center Foyer/Bellvue) 11:30-12:45 p.m. Oral Poster Presentations (Albenia) Session A—Lymphocyte Immunobiology I

8.

11:30-12:45 p.m. Oral Poster Presentations (Albenia) Session A - Lymphocyte Immunobiology I Chairs: Drs. Margaret Bynoe and Brian Rudd

11:30-11:45 Oyebola Oyesola, M.S. (Roswell Park Cancer Institute)

“The Prostaglandin D2 Receptor CRTH2 Mediates Interleukin-33-elicited Group 2 Innate Lymphoid Cell Accumulation in Tissues” (5)

11:45-12:00 Weishan Huang, Ph.D. (Cornell University)

“ITK Signaling via IRF4 Regulates the Development and Function of Type 1 Regulatory T Cells” (6)

12:00-12:15 Christina M. Post, M.S. (University of Rochester)

“The Ancestral Environment Shapes Antiviral CD8+ T Cell Responses Across Generations” (11)

12:15-12:30 Jennifer Yates, Ph.D. (Wadsworth Center)

“Early IL-10 Signals Favor Regulatory B Cell Over Memory B Cell Development during Cognate iNKT Cell Help” (17)

12:30-12:45 Victoria L. DeVault, B.S. (University of Vermont)

“SLAMf6 Modulates the NKT Cell Death Threshold” (20) 11:30-12:45 p.m. Oral Poster Presentations (Evelley) Session B - Tumor Immunology Chairs: Drs. Michael Robek and Yasmin Thanavala 11:30-11:45 Kelly L. Singel, B.S. (Roswell Park Cancer Institute)

“A Novel Barrier to Endogenous Anti-Tumor Immunity: Ovarian Cancer Ascites-activated Neutrophils Suppress T Cell Proliferation in a Contact-dependent Mechanism” (4)

11:45-12:00 Colin A. Powers, Ph.D. (Roswell Park Cancer Institute) “Tumor-induced Myeloid-derived Suppressor Cells Act via Remote Control to Inhibit L-selectin-dependent Adaptive Im-munity in Lymph Nodes” (24)

9.

12:00-12:15 Adaobi Amobi, B.S. (Roswell Park Cancer Institute)

“Tumor-Derived Indoleamine 2,3- Dioxygenase Regulates Density of Tumor Infiltrating CD8+ T cells and Myeloid Derived Suppressor Cells in a Murine Model of Ovarian Cancer” (26)

12:15-12:30 Amy Ku, B.S. (Roswell Park Cancer Institute)

“Negative Impact of Myeloid-derived Suppressor Cells on CD8 Effector T Cell Trafficking Within the Tumor Microenvironment” (27)

12:30-12:45 Anand Sharda, B.A. (Roswell Park Cancer Institute)

“Pretreatment Peripheral Blood Monocyte Subset Signature is Predictive of Patient Response to Dendritic Cell Vaccination” (48)

1:00-1:30 p.m. Lunch Buffet (Wapanak)

1:30-2:00 p.m. Platinum Corporate Sponsor—BioLegend, Inc. (Wapanak) Patrick Murphy, Ph.D. 2:00-3:15 p.m. Oral Poster Presentations (Albenia) Session C - Lymphocyte Immunobiology II Chairs: Drs. Nicholas Mantis and Gary Winslow

2:00-2:15 Stephanie L. Schell, B.S. (Penn State College)

“Autoantigen Availability Determines the Innate Sensing Requirement during Self-Antigen Driven Germinal Center Responses in Autoimmunity” (13)

2:15-2:30 Kevin Kenderes, B.S. (SUNY Upstate Medical University)

“IgM Memory B Cells Reconstitute Multiple B Cell Lineages and Provide Protection” (22)

2:30-2:45 Adam Utley, B.S. (Roswell Park Cancer Institute) “CD28 Induces Metabolic Fitness in LLPCs through NFκB-Mediated Irf4 Expression and ROS-Dependent Survival” (23)

10.

2:45-3:00 Joel R. Wilmore, Ph.D. (University of Pennsylvania)

“Commensal Microbes Drive the Generation of Systemic IgA Responses” (38)

3:00-3:15 Amber Papillion, B.S. (SUNY Upstate Medical University)

“Regulation of IgM Memory B cell Pool Size by the Inhibitory receptor FcγRIIb” (44)

2:00-3:15 p.m. Oral Poster Presentations (Evelley) Session D - Immunoregulation Chairs: Drs. Eyal Amiel and William O’Connor 2:00-2:15 Scott B. Minchenberg, B.S. (SUNY Upstate Medical University)

“Cytokine-mediated Regulation of Oligodendrocyte Metabolism” (8)

2:15-2:30 Camila Rosat Consiglio, M.S. (Roswell Park Cancer Institute) “Inflammation, Androgens and Macrophages in the Prostate:

Are We Missing the Link?” (18) 2:30-2:45 Phyu Thwe, B.S. (University of Vermont)

“Cell-Intrinsic Glycogen Metabolism Supports Early Activation and Maintains Metabolite Homeostasis in Dendritic Cells” (21)

2:45-3:00 Angelica Costello, B.A. (Albany Medical College)

“Macrophages Negatively Regulate Hematopoietic Stem Cells in Murine Aplastic Anemia” (40)

3:00-3:15 Travis Walrath, B.S. (Albany Medical College)

“Antagonistic Control of Intestinal Wnt Expression by IBD-Associated Cytokines” (49)

AAI Young Investigator Award and Oral Poster Presentation

eBioscience Trainee Travel Award and Oral Poster Presentation

3:15-3:30 p.m. Break 3:30-5:00 p.m. Workshop I - Dr. Wayne Yokoyama (Wapanak) “Opportunities to Study the Immunology of Human Diseases”

11.

5:00-6:30 p.m. Vendor/Poster Mixer (Bellvue) Poster Viewing and Questions (Odd Numbers) 6:30-7:00 p.m. Dinner (Wapanak)

7:00-7:30 p.m. Platinum Corporate Sponsor-BD Biosciences (Wapanak) Arielle Ginsberg 7:30-9:00 p.m. Vendor/Poster Mixer (Bellvue) Poster Viewing and Questions (Even Numbers) iPad drawing during this event. Must be present to win. 9:00-9:15 p.m. Remove Posters (Posters left behind will be discarded) Wednesday, October 26th 7:00-8:15 a.m. Breakfast at Leisure (Use voucher towards total cost) (La Bella Vita) 8:25-8:30 a.m. Morning Announcements (Nirvana)

8:30-9:30 a.m. Symposium III: Cancer Immunity (Nirvana) Chair: Dr. Edith Lord 8:30-9:00 Joseph Barbi, Ph.D. (Roswell Park) “Exploring the Mechanisms of Neurotrophin-mediated Immune Tolerance and Their Implications for Autoimmunity and Cancer” 9:00-9:30 Fiona Ginty, PhD. (GE Global) “In situ Analysis of Multiple Immune Cells in Tumor and Microenvironment” 9:30-10:30 a.m. Symposium IV: Immune Mediated Disease (Nirvana) Chair: Dr. Wayne Yokoyama

12.

9:30-10:00 Iwona Buskiewicz, Ph.D. (University of Vermont) “Reactive Oxygen Species Induce Virus-dependent MAVS-oligomerization in Systemic Lupus Erythematosus” 10:00-10:30 Timothy Chapman, Ph.D. (University of Rochester) “Increasing Regulatory Tone to Suppress Allergic Inflammatory Responses in the Lung” 10:30-10:45 a.m. Beverage Break (Conference Center Foyer)

10:45-11:45 a.m. Industry Panel (Albenia) “Careers Outside of Academia” 11:45-12:45 p.m. Lunch Buffet (Wapanak) 12:45 -2:00 p.m. Workshop II - Dr. Thomas Braciale (Nirvana) “The Business End of Academic Research” 2:15-2:30 p.m. Meet at Dock—Prepare to board The Morgan (Please have your boarding pass.)

2:30-4:00 p.m. Cruise sponsored by Krackeler Scientific, Inc. (Refreshments provided/Cash Bar)

4:00-6:00 p.m. Free Time 4:15-5:15 p.m. Scientific Advisory Board Meeting (Board Members Only) (Empire Room)

6:00-7:00 p.m. Dinner Buffet (Wapanak)

7:00-8:30 p.m. Workshop III—NIH Members “Grantsmanship, Funding, and Mock Peer Review”

8:30-11:30 p.m. Mr. Brown’s Pub and Caldwell’s Informal Discussions

13.

Thursday, October 27th

7:00-8:15 a.m. Breakfast at Leisure (Use voucher towards total cost)

(La Bella Vita)

8:25-8:30 a.m. Morning Announcements (Nirvana)

8:30-10:00 a.m. Symposium V: Immunity in Infectious Disease (Nirvana) Chair: Dr. Thomas Braciale

8:30-9:00 Richard I. Enelow, M.D. (Dartmouth College) “Control of the CD8+ Effector Population in Influenza Infection”

9:00-9:30 Christine King, Ph.D. (SUNY Upstate) “Mast Cells in Kaposi’s Sarcoma – A Model for the Pathogenesis of KSHV-driven Oncogenesis”

9:30-10:00 Ira J. Blader, Ph.D. (University at Buffalo) “Toxoplasma Infections of the Nervous System”

10:00-10:15 a.m. Beverage Break and Check-out (Conference Center Foyer/Hotel Lobby)

10:15-11:15 a.m. Keynote Presentation – Sponsored by BioLegend (Nirvana)

Introduction: Dr. Richard Enelow

Dr. Thomas J. Braciale Director, Emeritus Carter Immunology Center Professor of Pathology and Molecular Medicine University of Virginia

“Dendritic Cells in the Host Response to Stress or Dendritic

Cells Can Do More Than Present Antigens”

11:15-11:30 a.m. Closing Remarks(Nirvana) iPad drawing during this event. Must be present to win.

14.

Departure

Please join us next year for the

20th Annual

Upstate New York Immunology Conference

October 23-26, 2017

(Monday-Thursday)

The Sagamore Resort and Conference Center

Bolton Landing, NY

15.

NYIC 2015

16.

Platinum Corporate Sponsor

BD Biosciences

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18.

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20.

Silver-Plus Corporate Sponsor

22.

Silver Plus Corporate Sponsor

We are proud to sponsor

The 19th Annual Upstate New YorkImmunology Conference 2016

Krackeler Scientific is a leading distributor of quality scientific products. We serve customers nationwide in biotechnology, nanotechnology,

life sciences, pharmaceutical, biomedical, environmental, & industrial sectors.

Our comprehensive product line represents all of the leading manufacturers including Corning, Sigma, Agilent, Eppendorf & Celltreat. Our home is Albany, NY.

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24.

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29.

Bronze Corporate Sponsors

Tonbo Biosciences

30.

In recognition of the significance

of this meeting and work being done by

Graduate Students

and Postdoctoral Fellows,

the

American Association

Of Immunologists

has provided

Ten(10) Young Investigator Awards.

Each will receive a monetary award,

as well as the opportunity

to present their research both in poster format and brief talks.

31.

NYIC Scientific Advisory Board

Institutional Representatives

Albany Medical College

Jim Drake and Kate MacNamara

(NYIC Conference Organizers)

Cornell University

Margaret Bynoe

Dartmouth College

Brent Berwin

Roswell Park Cancer Institute

Yasmin Thanavala

SUNY Upstate Medical University

Gary Winslow

University at Buffalo

Beth Wohlfert

University of Connecticut

Steven Szcezpanek

University of Rochester Medical Center

Edith Lord

University of Vermont

Eyal Amiel

Wadsworth Center/SUNY Albany

Nicholas Mantis

32.

Institutional Financial Supporters


Alumni Association

Cornell University

Microbiology & Immunology

Dartmouth College

Department of Microbiology & Immunology


Department of Immunology


Microbiology & Immunology Program


Buffalo School of Medicine



Department of Molecular & Cellular Biology

Center of Excellence for Vaccine Research

University of Rochester Medical Center



Vermont Center for Immunology & Infectious Diseases

Wadsworth Center

33.

Grant support provided to

Graduate Students

and

Postdoctoral Fellows

by the

National Institutes of Health

National Institute of

Allergy and Infectious Diseases

R13AI051522

“Thank You”

34.

Keynote Speaker

Wayne M. Yokoyama, M.D. Levin Professor of Medicine

Investigator, Howard Hughes Medical Institute

Rheumatology Division, Dept. of Medicine

Washington University School of Medicine

St. Louis, Missouri

“Tissue-Resident Natural Killer Cells”

Most of what we know about natural killer (NK) cells comes from study of NK cells from the

mouse spleen and human peripheral blood. However, solid organs, such as the mouse liver, con-

tains two populations of NK cells, circulating conventional NK cells that resemble splenic NK cells,

and tissue-resident NK cells that do not recirculate. These NK cell populations can be distin-

guished by cell surface markers and transcription factor-dependence, indicating that they form

distinct lineages. Humans may also have tissue-resident NK cells but lineage-specific markers in

mice may not be easily translated to humans. Here I will describe our work on mouse tissue-

resident NK cells and progress we have made in characterizing human liver tissue-resident NK

cells.

Dr. Yokoyama’s research includes the role of NK cells in anti-pathogen defense led to studies of

innate immune responses to large DNA viruses, MCMV and CPXV. His lab is especially interested

in how these viruses interact with the host. They are also studying the interplay between innate

and adaptive immunity. They are also embarking on human studies aimed at understanding the

etiology and pathogenesis of autoimmune disease, especially rheumatic disorders (i.e., rheuma-

toid arthritis and Kawasaki disease.)

35.

Symposium I

Current Topics in Immunoregulation

Chair : Dr. Brent Berwin

36.

The Stress Protein Metallothionein: A Small Protein with a Large

Immunomodulatory Role

Michael A. Lynes

University of Connecticut, Storrs CT

Metallothionein (MT) is a protein that makes many interwoven contributions to cellular homeosta-

sis and to the management of cellular function under stressful conditions. These contributions include man-

aging essential metals such as copper and zinc, and regulating the local redox microenvironment, which

have been linked to MT’s influence on intracellular signaling cascades and many transcription factors (e.g.

Sp1 and NF-kB). MT can serve in these roles as a consequence of its thiol-rich nature: fully 33 mol% of

the protein is cysteine. MT has a complex upstream regulatory region, which is responsible for its expres-

sion as a consequence of cellular exposure to divalent heavy metal cations, reactive oxygen species, endo-

toxin, acute phase cytokines (IL-1, IL-6, and TNF-a), glucocorticoids, IFN-g, and aromatic hydrocarbons.

These inducers and regulatory roles suggest that MT plays an important role in the progression of an im-

mune response. We have shown that MT can influence elements of both the innate and adaptive immune

response. At least some of this effect is a consequence of its effect on intracellular NF-kB, while other

changes appear to rely on the MT-mediated management of intracellular Zn and the role that ion plays in

phosphatase activity. Under stressful circumstances, MT can exit the cell, despite the lack of a signal pep-

tide, and appears in extracellular fluids and spaces such as serum, urine, milk, bronchoalveolar lavage and

pancreatic acini, bronchoalveolar spaces, and liver sinusoids. We have shown that this extracellular pool

can serve as an activator of chemotactic responses, and that a targeted disruption of the Mt1 and Mt2 genes

or monoclonal antibody blockade of the extracellular pool of MT can significantly diminish the severity of

DSS-induced colitis in mice. Taken together, these observations suggest MT as an interesting and novel

target for therapeutic manipulation of the immune response.

37.

Biology of Inflammatory Innate Lymphoid Cells

Qi Yang and Kangning Zhang

Department of Immunology and Microbial Disease, Albany Medical College,

Albany, NY 12208

Mechanisms that control lymphocyte lineage stability and plasticity remain elusive. Recent work

indicates that innate lymphoid cells (ILC) possess substantial functional plasticity. Whereas natural type-2

innate lymphoid cells (nILC2) respond to IL-33 and produce type-2 cytokines, inflammatory ILC2 (iILC2)

respond to IL-25 and can co-produce both type-2 cytokines and also the ILC3-characteristic cytokine IL-

17. The transcriptional regulatory mechanisms that drive this plasticity, and the importance in health and

disease, remain unknown. Here we show that iILC2 are potent inducers of airway inflammation in re-

sponse to acute house-dust mite challenge. We find that Notch signaling drives the emergence of iILC2.

Acute blockade of Notch signaling abolished functional iILC2, but not nILC2, in vivo. Exposure of isolat-

ed nILC2 to Notch ligands in vitro altered their cytokine responsiveness and elicited dual IL-13/IL-17 pro-

duction, thus converting nILC2 into iILC2. In mature ILC2, Notch transcriptional complex bound to the

Rorc gene locus and promoted its expression, but did not affect the expression of Gata3, therefore confer-

ring ILC3-like capability without compromising primary ILC2 function. Together these results reveal a

novel role for Notch signaling in eliciting the plasticity of ILC2 and driving the emergence of highly pro-

inflammatory innate lymphocytes.

38.

Influence of Colony Stimulating Factors on Virus infections of Monocyte

Lineage Cells

Karin Schneider


Monocyte lineage cells are common targets for infection and innate immune activation by many

pathogenic viruses. As both infection and innate activation are key determinants of viral pathogenesis espe-

cially in the central nervous system, an analysis of colony stimulating factors on these events was per-

formed in cultures infected with Theiler’s Murine Encephalomyelitis Virus (TMEV) or Zika virus (ZIKV).

Our results suggest that the two single-stranded, positive sense viruses can uniquely interact to different

monocyte lineage cells.

39.

Symposium II

Neonatal Immunity

Chair : Dr. Gary Winslow

40.

The Fate of Neonatal and Adult CD8+ T cells During Infection is Linked to Their

Developmental Origin

Jocelyn Wang1, Erin Wissink2, Neva Watson1, Norah L. Smith1, Arnold Reynaldi3,

Andrew Grimson2, Miles P. Davenport3, and Brian D. Rudd1

Department of Microbiology and Immunology1, Cornell University, Ithaca, NY 14853

Department of Molecular Biology and Genetics2, Cornell University, Ithaca, NY 14853

Kirby Institute for Infection and Immunity3, University of New South Wales,

Sydney, Australia

We recently showed that neonatal CD8+ T cells fail to become memory cells because of an inher-

ent propensity to rapidly proliferate and become terminally differentiated. However, the underlying basis

for these differences remained unclear. To determine whether neonatal CD8+ T cells behave differently

than adults because of age-related differences in T cell homeostasis or thymopoiesis, we directly compared

the behavior and transcriptomes of neonatal and adult CD8+ T cells that had undergone similar amounts of

homeostatic proliferation in the periphery or were at the same stage of thymic development. We also com-

pared T cell maturation by fetal and adult precursors in the adult thymus and examined whether fetal-

derived CD8+ T cells behave differently than their adult counterparts. Our data demonstrates that neonatal

and adult CD8+ T cells adopt different fates after infection because they are derived from distinct HSCs,

which express different amounts of Lin28b. We also developed a system to ‘timestamp’ CD8+ T cells in

situ at various stages of development (1d, 1wk, and 4wks) and examined their behavior at 8 wks of age.

These data indicates that the spectrum of CD8+ T cell differentiation observed after infection is influenced

by when the responding cells were initially made.

41.

Maternal-placental-fetal Interactions Affecting Offspring Immunity and Behavior

David A. Lawrence1,2, Tapan Mondal1, Alvaro Mendoza1, Jane Lubliner2, and

Lori Kearsing3 1 Wadsworth Center/New York State Department of Health,

2University at Albany School of Public Health, and 3Center for Disability Services

Luminex and grating coupled surface plasmon coupled fluorescence (GCSPCF) technologies were

used to screen samples from humans and mice. The human samples were obtained from newborn dried

blood spots (NDBSs) collected from babies born in New York and later diagnosed with autism spectrum

disorder (ASD) or typical development. The proteins eluted from the NDBSs were compared to plasma

protein differences from the BTBRT+ltpr3tf/J (BTBR) mouse strain with ASD-like behavior and the

C57BL/6J (B6) strain, which has normal behavior. BTBR mice have reduced social interactions, impaired

play, low exploratory behavior, unusual vocalizations and high anxiety; they also have substantial deple-

tion of the corpus callosum and severely reduced hippocampal commissure. Antibody specificities and iso-

types from the NDBSs of babies that developed ASD and those from BTBR mice were assessed by

GCSPCF and Luminex analysis and shown to differ from the antibody specificities and isotypes from hu-

mans and mice with typical development and usually normal behaviors. The human and mouse compari-

sons demonstrate that humans and mice with different behaviors have different antibody and cytokine pro-

files from the control populations. The human and mouse comparisons demonstrate that humans and mice

with different behaviors have different antibody and cytokine profiles from the control populations. Mater-

nal-placental-fetal interactions leading to heightened antenatal inflammation and oxidative stress are sug-

gested to affect the aberrant immunity and behavior of offspring.

42.


Session A:

Lymphocyte Immunobiology I

Chairs: Drs. Margaret Bynoe

and Brian Rudd

43.

The Prostaglandin D2 Receptor CRTH2 Mediates Interleukin-33-elicited Group 2

Innate Lymphoid Cell Accumulation in Tissues

Oyebola Oyesola, Lauren M. Webb, Rebecca Cubitt, Elia Tait Wojno

Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca NY 14853, United

States

Group 2 innate lymphoid cells (ILC2s) are rare innate immune cells that contribute to the development of

type 2 inflammation. These cells and associated type 2 inflammation promote allergic lung inflammation, which

affects nearly 8% of the general population in the United States. Previous studies have shown that the accumulation

of ILC2s at inflammatory sites is mediated by epithelial cell-derived cytokines and alarmins, particularly interleukin-

33 (IL-33). In addition, prostaglandin D2 (PGD2), a bioactive lipid produced by mast cells, induces accumulation of

ILC2s at inflammatory sites by binding to its receptor chemoattractant receptor-homologous molecule expressed on

Th2 cells (CRTH2), which is expressed by ILC2s. However, whether the IL-33-IL-33 receptor (IL-33R) and PGD2-

CRTH2 pathways coordinate or intersect to regulate ILC2 responses during type 2 lung inflammation has not fully

been explored in vivo. In this study, we investigated the role of the PGD2-CRTH2 pathway during IL-33-elicited

ILC2 accumulation and type 2 inflammation in the lung. When wild-type mice were treated either systemically by

intraperitoneal injection or locally by intranasal injection with recombinant murine IL-33 (rmIL-33) to induce type 2

inflammation, ILC2 frequency and number increased in the lung parenchyma. In contrast, ILC2s accumulated in the

lung of mice deficient in CRTH2 (CRTH2KO) following intranasal but not intraperitoneal treatment with rmIL-33.

The defect in ILC2 accumulation in the CRTH2KO lung following systemic rmIL-33 treatment was not due to dif-

ferences in expression of the IL-33R, cell death, apoptosis, or cell proliferation. Therefore, our data suggest that the

PGD2-CRTH2 pathway acts downstream of IL-33 to partially mediate ILC2 accumulation in the lung during type 2

inflammation that originates in the periphery, acting via pathways other than those controlling cell responsiveness to

IL-33, death, or proliferation. Future studies will focus on determining if ILC2 accumulation in the lung in response

to IL-33 is mediated via CRTH2-dependent migration to or retention in the lung. A better understanding of how cy-

tokines and bioactive lipid mediators interact to regulate ILC2 responses will be important in informing the use and

development of drugs that target these pathways to treat ILC2-associated type 2 airway inflammation.

Poster 5

44.

ITK Signaling via IRF4 Regulates the Development and Function of Type 1

Regulatory T cells

Weishan Huang*, Sabrina Solouki, Nicholas Koylass, and Avery August*

Department of Microbiology and Immunology, Cornell University, Ithaca, NY 14853.

Type 1 regulatory T (Tr1) cells lack the expression of Foxp3 but have significant regulatory function in sup-

pressing inflammation and promoting tolerance, in part via their expression of the immunosuppressive cytokine IL-

10. Tr1 cells differentiate in response to signals engaging T cell receptor (TCR) and/or the regulatory cytokine mi-

lieu. The non-receptor tyrosine kinase ITK is a key modulator downstream of TCR, playing critical roles in T cell

development and function. Using mouse models carrying Foxp3RFP and IL-10GFP dual reporters, we found that, in

the absence of ITK, TCR activation-driven development of Foxp3- IL-10+ Tr1 cells is severely impaired in various

organs (spleen, blood, lung, gut, and fat). Itk-/- mice were also deficient in mounting a mucosal Tr1 cell response

during parasitic (Nippostrongylus Brasiliensis) and viral (Influenza A) infections.

Naïve Itk-/- thymic and splenic Foxp3- CD4+ T cells also exhibited severe deficiency in Tr1 differentiation

under Tr1 polarizing condition. Although Itk-/- CD4+ T cells proliferated under Tr1 differentiating conditions, they

failed to up-regulate IL-10, and Tr1 cell markers LAG3, CD49b, ICOS, PD-1, c-Maf, AHR, and IRF4 to levels ob-

served in WT cells, suggesting that ITK is critical for Tr1 cell fate programming. Utilizing a transgenic mouse model

carrying an allele sensitive mutant of ITK (ITKas) that allows ITK kinase specific blockade by a small molecule

3MB-PP1, we determined that the expression of the aforementioned markers, as well as the balance between IL-10

and IFN-g (gamma) production during Tr1 differentiation, are dependent on ITK kinase activity. Furthermore, using

cells from an ITKas-Foxp3RFP/IL-17AGFP dual reporter mouse model, we find that ITK kinase activity is required

for optimal Th17 trans-differentiation to Tr1 cells. We also find that inhibiting ITK kinase activity diminished Tr1

differentiation by human CD4+ T cells. The requirement for ITK function during Tr1 cell development can be re-

stored by the expression of the transcription factor IRF4. Finally, specifically targeting ITK kinase activity in already

differentiated Tr1 cells diminished their suppressive function.

We conclude that the TCR/ITK/IRF4 pathway is required for the development and function of Tr1 cells,

which may be targeted to modulate regulatory immunity for clinical benefit.

*Correspondence: Weishan Huang ([email protected]) and Avery August ([email protected]).

Poster 6

45.

The Ancestral Environment Shapes Antiviral CD8+ T cell Responses

Across Generations

Christina M. Post, Jason Myers, and B. Paige Lawrence

Department of Environmental Medicine, University of Rochester School of Medicine & Dentistry,

Rochester NY

Recent studies have linked health fates of great-grandchildren to environmental exposures of their great

grandparents. However, few studies have considered whether ancestral exposures influence the immune system

across generations. Here we report novel findings regarding transgenerational transmission of altered T cell respons-

es resulting from maternal (F0) exposure to an environmentally relevant aryl hydrocarbon receptor (AHR) ligand.

The AHR is a transcriptional regulator that plays diverse roles in cellular function, including modulating immune

responses. AHR ligands comprise several classes of pollutants, such as dioxins and PCBs, as well as molecules from

foods and other sources. AHR-binding pollutants cross the placenta and are excreted in breast milk. In animal mod-

els and human populations, early life exposure to dioxins and PCBs is associated with persistent defects in the off-

spring’s immune function. Using a mouse model, maternal exposure to the AHR ligand and pollutant 2,3,7,8-

tetrachlorodibenzo-p-dioxin (TCDD) results in a significantly reduced CD8+ T cell response to influenza A virus

(IAV) in the adult offspring (F1), compared to the response of infected offspring of control-treated dams. Specifical-

ly, there are significantly fewer cytotoxic T lymphocytes (CTL; CD44hiCD62Llo), virus-specific CD8+ T cells, and

CD8+ T cells that produce interferon gamma (IFNγ). Transcriptomic analyses using sorted CD8+ T cells from F1

offspring of TCDD and control dams support new evidence that triggering AHR during development changes pro-

gramming of senescence or exhaustion regulatory pathways. Follow up studies show increased expression of pro-

teins associated with hindered T cell responses, such as CTLA-4 and KLRG1 on CD8+ T cells. We next asked

whether the diminished CD8+ T cell response in the F1 generation was observed in the F3 generation. We detected

fewer CTL and virus-specific CD8+ T cells in the TCDD F3 lineage, as well as increased expression of CTLA-4 and

KLRG1 compared to control F3 lineage following IAV infection. These data indicate that F0 maternal exposure to

AHR ligands is capable of disrupting immune function not only via direct activation of the AHR in the F1 genera-

tion, but also by reprogramming immune responses in subsequent generations. This has broad implications for un-

derstanding how the environment of prior generations shapes susceptibility to pathogens and antiviral immunity in

later generations.

Poster 11

46.

Early IL-10 Signals Favor Regulatory B cell Over Memory B cell Development during Cognate

iNKT Cell Help

Jennifer Yates1,4, Emilie Vomhof-Dekrey1, Paula Lanthier1, Katja Mohrs1, Thomas Hägglöf 2,

Natacha Veerapen3, Gurdyal Besra3, Mikael Karlsson2, and Elizabeth Leadbetter1,5 1Trudeau Institute, Saranac Lake, NY 12983; 2Karolinska Institutet, Stockholm, Sweden;

3School of Biosciences University of Birmingham, Birmingham, UK; 4Wadsworth Center, NYSDOH, Albany, NY; 5University of Texas Health Science Center at San Antonio,

San Antonio, TX

Effective generation of humoral B cell memory is dependent upon help from CD4+ T cells. We and

others have found that invariant natural killer T (iNKT) cells can provide both cognate and non-cognate

helper signals to enhance B cell responses. While both cognate and non-cognate iNKT cell help induce

class-switched, antigen-specific humoral immune responses - only non-cognate iNKT cell help drives the

formation of humoral memory. Rather, cognate iNKT cell help drives an early, un-sustained expansion of

germinal center B cells and antigen-specific antibody production. Therefore, we posit that cognate help

provided to B cells by iNKT cells is fundamentally different from the help provided by conventional CD4+

T cells. We now find that glycolipid immunization drives considerable IL-10 transcription by many differ-

ent spleen cell populations including dendritic cells, plasmablasts, iNKT cells, and B regulatory cells. Cog-

nate iNKT cell help expands antigen-specific IL-10 producing B regulatory cells upon primary immuniza-

tion, and IL-10 producing iNKT10 cells following secondary antigen challenge. Early, but not late, block-

ade of the IL-10 receptor resulted in a significant, and sustained increase in antigen-specific antibody titers

during cognate iNKT cell help, but had no effect when traditional CD4+ T cell help was present. We con-

clude that the early composite cytokine environment is critical for dictating the long-term course of the B

cell response. Based on these data, we suggest that B cell antigens which recruit only cognate help from

iNKT cells experience a regulatory rather than inflammatory environment.

Poster 17

47.

SLAMf6 Modulates the NKT Cell Death Threshold

Victoria L. DeVault1, Oliver Dienz1, Graham W.J. Lilley1, Patrick Benoit1, Pamela L. Schwartzberg2,

Jonathan E. Boyson1 1Department of Surgery, University of Vermont College of Medicine, Burlington, VT 2National Human Genome Research Institute (NHGRI), National Institutes of Health,

Bethesda, MD

Signaling lymphocyte activation marker family member 6 (SLAMf6) is a cell surface signaling re-

ceptor that plays a critical role in NKT cell development. Surprisingly, the exact mechanisms through

which SLAMf6 regulates NKT cell development and function remain unclear. To investigate the function

of SLAMf6 on peripheral NKT cells, we challenged C57BL/6 (B6) or B6.Slamf6-/- mice with the NKT cell

agonist, aGalCer. While we detected no difference between B6 and B6.Slamf6-/- mice in NKT cell IFN-g,

IL-4, and TNFa production, we did find a significant difference in NKT cell numbers in the periphery.

Three days after challenge, we observed a 50-fold increase in NKT cells in B6 mice over vehicle-treated

controls. In contrast, we found a 220-fold increase in NKT cells in B6.Slamf6-/- mice versus controls. A

comparison between B6 and B6.Slamf6-/- mice of in vivo BrdU uptake by NKT cells revealed no signifi-

cant differences in proliferation. We then compared NKT cell apoptosis using Annexin V staining and

live/dead discriminator 3 h after aGalCer administration. This analysis revealed a significantly lower per-

centage of apoptotic NKT cells in B6.Slamf6-/- versus B6 mice, suggesting that SLAMf6 expression on

NKT cells was associated with activation-induced cell death (AICD). Consistent with this observation, we

found significantly diminished expansion of sorted B6 NKT cells when they were stimulated by SLAMf6+

antigen-presenting cells in an in vitro cell culture system. We conclude that in the presence of a strong,

high-affinity agonist, SLAMf6 contributes to significantly increased NKT cell AICD and concomitant di-

minished NKT cell expansion. Interestingly, when we conducted similar comparisons in naïve mice under

homeostatic conditions, we observed an increased percentage of apoptotic NKT cells in B6.Slamf6-/- mice

versus their B6 counterparts, which was associated with lower NKT cell numbers in B6.Slamf6-/- mice.

Taken together, these data support a model in which SLAMf6 regulates NKT cell activation and death

thresholds depending on the strength of activation. These data also suggest that SLAMf6 blockade could

be a useful tool to manipulate NKT cell populations in vivo.

Poster 20

48.


Session B

Tumor Immunology

Chairs: Drs. Michael Robek and

Yasmin Thanavala

49.

A Novel Barrier to Endogenous Anti-Tumor Immunity: Ovarian cancer

ascites-activated neutrophils suppress T cell proliferation in a contact-dependent mechanism

Kelly L. Singel1, ANM Nazmul H. Khan2, Kirsten B. Moysich3, Kunle Odunsi4,5, Brahm H. Segal1,2,6

Departments of 1Immunology, 2Medicine, 3Cancer Prevention and Control, 4Gynecologic Oncology, and

the 5Center for Immunotherapy, Roswell Park Cancer Institute, Buffalo, NY 6Department of Medicine, University at Buffalo Jacobs School of Medicine and Biomedical Sciences,

Buffalo, NY

Neutrophils are the first responders to infection and injury, and critical for antimicrobial host de-

fense. Through the generation of reactive oxidants, activation of granular constituents, and neutrophil ex-

tracellular traps (NETs), neutrophils target microbes and prevent their dissemination. While these path-

ways are beneficial in the context of trauma and infection, their roles in the context of tumor are less un-

derstood. Ovarian cancer (OC) is often diagnosed at advanced stages and presents with ascites. Necrosis is

a hallmark of advanced cancer and releases DAMPs that activate innate immune responses. Cytotoxic T

lymphocyte (CTL) immunity is critical in OC, and barriers to durable anti-tumor immunity include TAMs,

MDSCs, and Tregs. While activated neutrophils can kill tumor cells, knowledge is limited on the role of

activated neutrophils in the tumor microenvironment.

Our prior studies showed that granulocytic cells from ascites of patients with newly diagnosed OC

variably suppressed stimulated normal donor T cell proliferation ex vivo. In ascites, ~90% of cells were

inflammatory (CD45+) with varying proportions of neutrophils, monocytes, and lymphocytes. Neutrophils

comprised ~15% of CD45+ cells and the neutrophil:CD8+ T cell ratio was 1.5:1. In ex vivo studies, cell-

free ascites (CFA) attracted normal donor neutrophils (NDN) and induced NETs. We next evaluated the

effects of CFA-treated NDN on T cell proliferation. In co-culture studies, a subset of CFA (5/10 tested)

activated NDN to completely suppress CD3/CD28-stimulated T cell proliferation. Neither CFA nor NDN

alone impaired T cell proliferation as measured by [3H] thymidine at a 1:1 target:effector cell ratio. Sup-

pression did not affect T cell viability or induce apoptosis, but required T cell contact with NDN in the

presence of CFA. In addition, we found that PD-1 expression on the suppressed T cells phenocopied the

unstimulated controls. When T cells were CD3/CD28-stimulated overnight before exposure to CFA and

NDN, T cell proliferation was not inhibited by the addition of CFA and NDN, suggesting that CFA-treated

NDN interrupt T cell proliferation at an early stage of activation.

These results support a model in which neutrophils in the ascites of a subset of OC patients sup-

press T cell responses, which may be an important barrier to endogenous anti-tumor immunity and immu-

notherapy. Further studies will identify ascites constituents that activate neutrophils and mechanisms for

neutrophil-mediated T cell suppression. This work may lead to novel prognostic biomarkers regarding in-

flammatory responses in the OC tumor microenvironment and therapeutic approaches that target neutro-

phils.

Supported by the NIH (R01, CA188900; T32, CA085183), the RPCI-UPCI Ovarian Cancer SPORE (P50

CA159981), and the Cancer Center Support Grant to RPCI (CA016056).

Poster 4

50.

Tumor-induced Myeloid-derived Suppressor Cells Act via Remote Control to Inhibit L-selectin-

dependent Adaptive Immunity in Lymph Nodes

Amy W. Ku1, Jason B. Muhitch1, Colin A. Powers1, Michael Diehl1, Anand P. Sharda1,

Kieran O'Loughlin1, Hans Minderman1, Joseph J. Skitzki1, Suzanne Ostrand-Rosenberg2,

Scott I. Abrams1, and Sharon S. Evans1 1Roswell Park Cancer Institute, Buffalo, NY 14263, USA; 2University of Maryland Baltimore County,

Baltimore, MD 21250, USA

Myeloid derived suppressor cells (MDSC) are potent immunomodulatory cells that play an exten-

sive role in cancer progression and immune evasion. These immature myeloid cells are known to accumu-

late within the spleen and tumor, and their ability to suppress effector T cell functions within these tissues

is well described. In contrast, the impact of MDSC on naive T cells within lymph nodes (LN) has been

largely overlooked as MDSC are rare within these critical sites of immune priming. Previous reports have

shown that peripheral MDSC from tumor-bearing mice downregulate the LN homing receptor L-selectin

on naive CD4 and CD8 T lymphocytes, but the molecular mechanisms and target cell-specificity has been

unclear. Furthermore, the biological relevance of moderate fluctuations of L-selectin is questionable as the

high density of L-selectin molecules normally present on T cells could theoretically buffer against the ef-

fects of such loss during trafficking. Using stringent murine mammary tumor models of high and low

MDSC burden (4T1 and AT-3, respectively), we demonstrate that MDSC downregulate L-selectin on na-

ive T and B cells post-transcriptionally via a contact-dependent mechanism. MDSC-driven loss of L-

selectin occurs within 24 hours both in vitro and in vivo, and does not appear to be species-restricted as L-

selectin on human lymphocytes can also be targeted by MDSC. By employing real-time intravital micros-

copy and immunofluorescence histology to visualize and assess naive CD8 T cell trafficking within vascu-

lar gateways for lymphocyte trafficking known as high endothelial venules (HEV), we found that even

moderate losses of L-selectin mediated by MDSC causes a profound reduction in the quality of lympho-

cyte-HEV interactions. Ultimately, this results in significantly fewer T cells trafficking and infiltrating

into the LN parenchyma. In an in vivo vaccination model, MDSC-mediated loss of L-selectin on naive

CD8 T cell and subsequent reduction in lymphocyte trafficking severely diminishes antigen-driven T cell

expansion within draining LN. These data reveal a novel mechanism by which tumor-induced MDSC lo-

calized outside of the LN shape the magnitude of T cell responses within the intranodal compartment,

which has unanticipated implications for systemic immunity in cancer. Supported by the NIH (CA79765,

AI082039, T32CA085183, 5T32CA108456, 5P30 CA016056), the Breast Cancer Coalition of Rochester,

the Mark Diamond Research Fund and the Jennifer Linscott Tietgen Family Foundation.

Poster 24

51.

Tumor-Derived Indoleamine 2,3- Dioxygenase Regulates Density of Tumor

Infiltrating CD8+ T cells and Myeloid Derived Suppressor Cells in a Murine Model of Ovarian

Cancer

Adaobi Amobi1,3, and Kunle Odunsi1,2,3

Departments of Immunology1, Gynecologic Oncology2 and Center for Immunotherapy3

Roswell Park Cancer Institute, Buffalo, NY

Amino-acid withdrawal is an important molecular mechanism regulating anti-tumor immune re-

sponses. The catabolism of the essential amino-acid tryptophan (TRP) by indoleamine 2,3-dioxygenase

(IDO1) is a central pathway that contributes to the immunosuppressive microenvironment in many types

of cancer. IDO1 enzymatic activity results in depletion of TRP and the generation of immunosuppressive

metabolites, such as kynurenine. Our lab has previously shown that IDO1 expression in human ovarian

tumor correlates with poor prognosis and poor tumor infiltration by CD8+ T cells. Moreover, our lab

demonstrated that increased infiltration of CD8+ T cells into the tumor is associated with improved sur-

vival. Thus, IDO1 inhibition represents an attractive target for cancer immunotherapy.

To establish the mechanism by which IDO1 inhibition augments immune responses in a murine

model of metastatic ovarian cancer, we utilized a murine ovarian surface epithelial cancer cell line, IE9-

mp1. We generated a stable IDO1-overexpressing cell line (IE9mp1-mIDO1) by transfecting murine IDO

cDNA into parental IE9-mp1 cells and confirmed functional IDO1 enzyme activity. C57BL/6 mice were

challenged intraperitoneally with either parental IE9mp1-Empty Vector (IE9mp1-EV) or IE9mp1-

mIDO1 tumor cells. Syngeneic immunocompetent mice inoculated with IE9mp1-mIDO1 cells displayed

earlier onset of tumor burden and decreased overall survival compared with IE9mp1-EV challenged

mice.

To delineate the role of host- and tumor-derived IDO1 on immune cell infiltration to the tumor

site, we utilized the IDO1 genetic knockout (IDO1KO) mouse model. IDO1KO and C57BL/6 mice were

challenged intraperitoneally with either the IE9mp1-EV or IE9mp1-mIDO1 tumor cells. C57BL/6 and

IDO1KO mice challenged with IE9mp1-mIDO1 demonstrate reduced CD8+ T cell infiltration within the

tumor. Interestingly, IDO1KO mice challenged with IE9mp1-EV tumor cells demonstrate increased tu-

mor infiltration by CD8+ T cells compared to C57BL/6 mice. Moreover, tumor-derived IDO1 mediates

increased frequency in the CD11b+Gr1+ myeloid derived suppressor cell (MDSC) population in ascites

fluid early-on along tumor burden in C57BL/6 and IDO1KO tumor-bearing animals.

From these data, we conclude that regulation of IDO1 will promote anti-tumor immune responses,

by permitting increased frequency of effector T cells in tumor tissues. Moreover, tumor-derived IDO1

inhibition may decrease the frequency of CD11b+Gr1+ MDSCs in ascites fluid. Future studies are ongo-

ing to further delineate the specific contribution of IDO1 by tumor cells, host cells, or both mutually to

the regulation of immunosuppressive MDSCs in ovarian cancer. Experiments are ongoing to characterize

the mechanism by which IDO1 inhibition may augment vaccine-induced immune responses in a murine

model of ovarian cancer.

Supported by: NCI SPORE P50 CA159981

Poster 26

52.

Negative Impact of Myeloid-derived Suppressor Cells on CD8 Effector T cell

Trafficking Within the Tumor Microenvironment

Amy Ku1, Michelle Appenheimer1, Jason Muhitch2, Scott I. Abrams1, and Sharon S. Evans1

Departments of Immunology1 and Urology2, Roswell Park Cancer Institute, Buffalo, NY

The success of T cell-based immunotherapy and, unexpectedly, thermal therapy, standard chemo-

therapy and radiation hinges on cytotoxic T cells gaining access to tumor targets. These observations have

prompted interest in strategies to improve T cell trafficking to tumors although the mechanisms that posi-

tively or negatively regulate extravasation at tumor vascular checkpoints are poorly understood. Here, we

report that the ability of tumor vessels to respond to IL-6-dependent preconditioning regimens that boost

CD8 effector T cell homing is temporally and inversely related to the accumulation of myeloid-derived

suppressor cells (MDSC) within the tumor microenvironment. Using real-time intravital imaging and im-

munofluorescence histology, IL-6 therapies were shown to convert vessels from T cell-low to -high recruit-

ment sites in murine tumors with minimal MDSC infiltration (i.e., CT26 colorectal, B16 melanoma, EMT6

mammary tumors). This conversion requires induction of the ICAM-1 trafficking molecule on tumor ves-

sels. Conversely, mammary (4T1, AT-3 and PyMT-MMTV) and pancreatic (Pan02) tumors with high

MDSC burdens were refractory to IL-6 therapies, but became responsive after acute MDSC depletion. To

further investigate contributions of MDSC to poor trafficking, IL-6-responsive tumors were admixed with

syngeneic CD11b+Gr-1+ MDSC isolated from spleens of tumor-bearing mice at a ratio of 2:1, thus mim-

icking the high MDSC burden detected in IL-6-refractive tumors. Sustained intratumoral elevation of

MDSC in admixed tumors resulted in failure to support increased T cell trafficking in response to IL-6–

dependent therapies. Complementary in vitro studies revealed that MDSC directly influence and downreg-

ulate trafficking molecule expression on endothelial cells. Taken together, these findings identify a novel

role of MDSC in subverting antitumor immunity by limiting T cell trafficking at tumor vascular loci. Sup-

ported by NIH (R01CA79765, R01AI082039, 2T32CA085183), the Breast Cancer Coalition of Rochester,

the Mark Diamond Research Fund and the Jennifer Linscott Tietgen Family Foundation.

Poster 27

53.

Pretreatment Peripheral Blood Monocyte Subset Signature is Predictive of Patient Response to

Dendritic Cell Vaccination

Anand Sharda1, Alexander Wald1, Mohammad Habiby Kermany1, Katja Koeppen2, Thomas Hampton2,

Jan Fisher3, Camilo Fadul3, Marc Ernstoff4, Thomas Schwaab1, Jason Muhitch1,5

Department of Urology1, Medicine4, and Immunology5 Roswell Park Cancer Institute, Buffalo, NY De-

partments of Microbiology and Immunology2, and Medicine3 Geisel School of Medicine at Dartmouth,

Hanover, NH

Clinical trials have demonstrated that dendritic cell (DC) vaccination can initiate durable anti-

tumor immunity in a subset of cancer patients resulting in complete responses, even in stage IV Renal Cell

Carcinoma (RCC). The influence of monocytes, the starting material for conventional DC vaccines, on

patient responses remains under-investigated. Recently, three subsets of monocytes have been described

(classical, intermediate, and non-classical), each with distinct functional properties. However, their roles

in anti-tumor immunity, particularly in the context of DC vaccination, are unclear. The goal of this study

was to determine whether the circulating pretreatment monocyte subset gene expression and composition

from Stage IV RCC patients prior to DC vaccination predicted responses to treatment in a completed

phase II clinical trial (NCT00085436). Pretreatment circulating classical (CD14++, CD16-), intermediate

(CD14++, CD16+), and non-classical (CD14+, CD16++) monocyte subsets were isolated from patients. Pre-

treatment peripheral blood from complete responders (2 of 3 have no observable disease > 5 years follow-

ing therapy) contained fewer classical monocytes (57.5% ± 6.4) compared to all other groups (P < 0.05).

Interestingly, a higher percentage of DC derived from non-classical monocytes expressed costimulatory

molecules (CD80; 96.4%, CD86; 91.1%, HLA-DR; 99.1%) compared to classical monocyte-derived DC

(64.6%, 54.4%, 60.2%, respectively). DC derived from non-classical monocytes were also superior in

their ability to induce allogeneic T cell proliferation compared to DC originating from classical mono-

cytes. Additional gene expression analysis by unsupervised hierarchical clustering clearly distinguished

the transcriptional profile of classical, intermediate, and non-classical monocytes from RCC patients to

healthy controls. Further investigation revealed that monocytes from long term survivors (> 10 years)

could be distinctly segregated from other RCC patients. These findings demonstrate that DC-derived from

the minor CD16+ monocyte subset may represent a superior product for use in vaccination protocols. Gene

expression profiling of circulating monocytes may provide an accessible biomarker for patient responsive-

ness to immunotherapy. Future studies will address whether pretreatment levels of intermediate and non-

classical monocytes are prognostic indicators for response to additional immunotherapies, including

checkpoint blockade inhibitors.

Poster 48

54.

Patrick Murphy, Ph.D.

Technical Application Scientist

BioLegend

Flex-T™: Produce MHC Tetramers with Your Peptide of Interest

Identification of antigen-specific cytotoxic T-lymphocytes (CTLs) is important to understanding T

cell responses to infection and in both vaccine and immune therapy development. Production of MHC

class I tetramers however is a time-consuming, difficult process and remains a barrier to rapid identifica-

tion of antigen-specific CTLs. To simplify the generation of MHC class I tetramers, BioLegend has devel-

oped the Flex-T™ flexible MHC tetramer system. Flex-T™ is composed of MHC monomers loaded with a

UV-labile peptide that is degraded via use of a UV light source. When UV irradiation is performed in the

presence of the peptide of interest, peptide exchange occurs and the peptide of interest binds to the MHC I

peptide-binding groove. MHC monomers containing your peptide of interest are then made into tetramers

using fluor-conjugated streptavidin. In addition to being simple to use, Flex-T™ technology allows for

identification of up to 15 peptide targets per sample via combinatorial color coding. This advance reduces

the need for large volumes of blood to identify multiple rare CTL populations. Flex-T™ technology expe-

dites large-scale screening studies and allows for simple, cost-effective MHC class I tetramer production in

the lab.

55.


Session C

Lymphocyte Immunobiology II

Chairs: Drs. Nicholas Mantis and

Gary Winslow

56.

Autoantigen Availability Determines the Innate Sensing Requirement During

Self-Antigen Driven Germinal Center Responses in Autoimmunity

Stephanie L. Schell, Chetna Soni, and Ziaur S.M. Rahman

Penn State College of Medicine, Hershey, PA

Systemic lupus erythematosus (SLE)-associated germinal center (GC) responses are driven by syn-

ergistic BCR, TLR, and IFN signaling. Aberrant selection processes within these GCs lead to the escape

of high-affinity, class-switched autoreactive B cells. The environmental factors that stimulate TLRs and

the regulation of TLR and IFN signaling events during autoreactive GC responses in vivo are incompletely

defined. Previous studies in our lab established that MerTK-deficient (Mer-/-) mice, which lack a critical

receptor expressed by macrophages and DCs that is involved in apoptotic cell clearance and TLR/IFN im-

munoregulatory signaling cascades, exhibit increased GC responses and T cell activation. Here, we used a

kinetic immunization-based approach in Mer-/- mice to drive apoptotic cell accumulation in GCs, allowing

us to evaluate the contribution of TLR-dependent self-ligand sensing and dampened immunoregulation on

GC formation and selection processes, under conditions of high and low autoantigen availability. As antic-

ipated from previous study of spontaneous GC responses in autoimmune-prone mice, TLR7-MyD88 sig-

naling significantly contributes to enhanced GC responses observed in Mer-/- mice, independent of autoan-

tigen load. Unexpectedly, there was also a kinetic dependence on TLR9 in autoantigen-driven GC re-

sponses, suggesting that self-DNA sensing by TLR9 may have a more complex and context-dependent role

in GC responses than initially postulated. Further, the deficiency of MyD88 under conditions of high auto-

antigen availability did not diminish GC response, potentially indicating a novel regulatory function for

MyD88 or its dependent pathways whereby the sensing of ligand present in cytosolic spillover caused by

high autoantigen load is inhibited. Mechanistically, Mer-deficiency promotes enhanced GC responses in

GC B cell-intrinsic and APC-dependent manners, with process/cell-type dependent requirements for TLR

signaling. As a result of altered immune activation, the loss of Mer compounded aberrant GC selection

and downstream kidney pathology in autoimmune-prone B6.Sle1b mice deficient for Mer (B6.Sle1b.Mer-/-

). Altogether, these results suggest that TLR signaling plays a complex role in self-antigen driven GC re-

sponses, whereby both kinetics and autoantigen load determine the requirement for different self-ligands

and the activation of their relevant receptors during response. Mer-deficiency also promotes autoimmunity

by synergizing with genetic susceptibility loci to dysregulate selection processes within GCs.

Poster 13

57.

IgM Memory B cells Reconstitute Multiple B Cell Lineages and Provide Protection

Kevin Kenderes, Amber Papillion, and Gary Winslow


IgM memory B cells are now recognized as an important component of immunological memory.

They have been proposed to be a reservoir of broadly-reactive B cells that differentiate, in germinal cen-

ters, into high affinity class-switched B cells following antigen encounter. We provide evidence that a

highly-purified IgM memory B cell population can follow multiple pathways of differentiation after chal-

lenge infection, and demonstrate that the antibodies produced by these cells can provide protective immun-

ity. Our experimental model uses Ehrlichia muris, an intracellular tick-borne bacterium that generates a

robust CD11c+ T-bet+ IgM memory B cell population in C57BL/6 mice. Due to the presence of pre-

existing antibodies, investigation of the secondary IgM memory B cell response to ehrlichial infection had

not been possible. We are able to monitor EYFP-labeled IgM memory B cells after transfer of splenocytes

into naïve mice and observed differentiation EYFP-labeled cells into all effector and memory B cell line-

ages following secondary infection. This was accompanied by a 4-fold increase in IgM production, relative

to infected mice that did not receive memory cells. However, a small population of EYFP-labeled switched

memory cells was also found in the donor spleen cells. Therefore, to determined if IgM memory cells were

solely responsible for the reconstitution of the memory and effector B cell lineages we monitored highly

purified EYFP-labeled spleen IgM memory B cells following their transfer into naïve recipient mice. After

challenge infection, some donor memory B cells differentiated into IgM-producing plasmablasts and CD19

-negative plasma cells. Other donor B cells entered germinal centers, down-regulated CD11c, underwent

class switching, and generated switched memory B cells. Yet other donor cells were maintained as IgM

memory cells. Donor IgM memory B cells also protected the recipient mice from the fatal erhlichial infec-

tion, Ixodes ovatus ehrlichia (IOE), demonstrating the importance of IgM memory cells for protective sec-

ondary responses. Thus, during secondary responses, IgM memory cells can differentiate into IgM-

producing plasmablasts, switched germinal center cells, or switched memory cells, or undergo self-

renewal. These data reveal that IgM memory B cells are capable of generating protective secondary re-

sponses that can replenish many, if not all, effector and memory B cell lineages, thereby contributing to

long-term immunity to pathogens.

Poster 22

58.

CD28 Induces Metabolic Fitness in LLPCs through NFkB-Mediated Irf4 Expression and ROS-

Dependent Survival

Adam Utley, James Cooper, Louise Carlson, Peng Peng, Amin Mahpour, and Kelvin Lee.

Roswell Park Cancer Institute, Buffalo NY

Sustained humoral immunity is dependent upon the continual production of antigen-specific anti-

bodies by plasma cells. Upon activation, B cells differentiate into short-lived plasma cells (SLPCs) that

traffic to secondary lymphoid organs such as the spleen where they live for days to weeks then die by

apoptosis. In a second non-mutually exclusive model, B cells can differentiate into long-lived plasma cells

(LLPCs) that home to specialized survival niches in the bone marrow and live indefinitely. Much work

has gone into describing the competitive BM survival niche; however, the cellular and molecular interac-

tions which govern this survival program are incompletely understood.

We have published that CD28, the canonical T-cell costimulatory molecule, is required for LLPC

survival. In T cells CD28 is known to induce glycolysis at the expense of mitochondrial respiration. To

our great surprise, CD28 increased mitochondrial respiration in LLPCs whilst not affecting glycolysis di-

rectly. CD28 increased Glut1 expression and subsequent LLPC glucose uptake, as well as the glycolytic

capacity. This suggests that CD28 regulates the ability of LLPCs to successfully compete for nutrients in

the BM niche for long term survival and antibody production.

Mechanistically, CD28 induces NFkB-dependent IRf4 upregulation, known to regulate LLPC sur-

vival. Furthermore, inhibition of NFkB abrogates the CD28-induced increases in glucose uptake and mito-

chondrial mass. We recently published that the Grb2/Vav binding domain on the CD28 cytoplasmic tail is

required for LLPC survival. In mice wherein this domain is mutated (AYAA mutants), LLPCs have de-

creased Irf4 expression, glucose uptake and mitochondrial mass. This facilitates a model wherein CD28

induces NFkB dependent Irf4 through Grb2/Vav for metabolic fitness. We also describe an NFkB super-

enhancer element upstream of the Irf4, suggesting that CD28 may govern direct Irf4 promoter activity as

well as DNA folding. Irf4 knock down decreased PC mitochondrial mass, demonstrating that Irf4 may

directly regulate LLPC metabolic fitness.

One byproduct of mitochondrial respiration is the production of reactive oxygen species (ROS).

CD28 increased ROS specifically in LLPCs. Paradoxically, ROS inhibition prevented CD28-mediated

survival. Taken together these results suggest a model wherein CD28 through its Grb2/Vav binding do-

main induces NFkB dependent upregulation of Irf4 directly through the promoter region, and augments

further Irf4 production through a previously undescribed NFkB superenhancer element. Irf4 then goes on

to increase mitochondrial respiration-dependent ROS for CD28-mediated LLPC survival and metabolic

fitness. Targeting CD28 with FDA-approved drugs may augment vaccine design as well as alleviate anti-

body mediated auto-immunity.

Poster 23

59.

Commensal Microbes Drive the Generation of Systemic IgA Responses

Joel R. Wilmore, Brian Gaudette, Wenzhao Meng, Eline T. Luning Prak, and David Allman

Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of

Pennsylvania, Philadelphia, PA

It is well known that IgA functions as a critical component of the mucosal barrier in the gut by vir-

tue of its ability to be secreted into the intestinal lumen. However, little is known about the origin, function

and regulation of IgA in the serum. Mucosal IgA responses have been generally thought of as short-lived

and restricted locally to mucosal tissues. However, we find that IgA-secreting PCs make up the majority of

the BM PC pool in standard C57BL/6 mice bred in our colony. The IgA+ BM PCs are predominantly in the

long-lived pool and express gut homing factors such as CCR9 and the integrin α4β7 (alpha4 beta7), sug-

gesting a mucosal origin. The extent to which the commensal microflora influences the BM plasma cell

pool is evident by mice bred in germ free isolators that lack IgA+ PCs in their BM. Additionally, mice

from standard vendors such as Jackson labs (Jax), have extremely low levels of IgA+ PCs in the BM and

significantly lower serum IgA. Exposing Jax mice to a disparate microflora or Helicobacter sp. led to the

generation of IgA-secreting BM cells, while also inducing increases in serum IgA antibodies enriched for

binding to several commensal bacterial taxa. Moreover, BM IgA-secreting plasma cells exhibited a com-

mon clonal ancestry with intestinal IgA+ plasma cells, and both populations possessed unique gene expres-

sion signatures compared to other long-lived BM plasma cells. We conclude that commensal microbes

overtly influence the BM plasma cell pool, and suggest that select commensal microbes can facilitate the

induction of systemic humoral immunity.

Poster 38

60.

Regulation of IgM Memory B cell Pool Size by the Inhibitory receptor FcRIIb

Amber Papillion and Gary Winslow

Upstate Medical University, Syracuse, NY

Ehrlichia muris infection generates a long-term CD11c/T-bet-positive IgM memory population in

the spleen (J. Immunol. 191:1240). Among the many surface markers that distinguish the IgM memory

cells from canonical B cells is the inhibitory Fc receptor, FcRIIb, which exhibited a 2-fold higher expres-

sion. We hypothesized FcRIIb negatively regulates IgM memory cells by binding immune complexes pre-

sent during low-level chronic infection. To investigate this question, we monitored the IgM memory cell

population in infected FcRIIb-deficient mice. Thirty days post-infection, the IgM memory B cells were

generated earlier, and were found at three-fold higher frequencies in FcRIIb-deficient mice, compared to

wild-type mice. This increase in the frequency of spleen IgM memory cells was due to an increase in cell

number, and was in turn associated with an increase in antigen-specific IgG. These data indicate that Fc

RIIb plays an important role in regulating the expansion and/or persistence of IgM memory cells in wild-

type mice under conditions where antigen-specific IgG is sufficient to control infection. Other studies re-

vealed that the IgM memory population in FcRIIb-deficient mice exhibited much lower expression of

FAS, CD40, BAFF-R, and TACI. We therefore proposed that FcRIIb signaling, likely via immune com-

plexes, acts in wild-type cells to regulate the size of the IgM memory cell pool, by maintaining the expres-

sion of FAS and other receptors that regulate cell survival. These data suggest a novel regulatory role for

FcRIIb in B cell memory.

Poster 44

61.


Session D

Immunoregulation

Chairs: Drs. Eyal Amiel and

William O’Connor

62.

Cytokine-mediated Regulation of Oligodendrocyte Metabolism

Scott B. Minchenberg, Anthony F. Paredes, and Paul T. Massa

SUNY Upstate Medical University, Department of Microbiology & Immunology, Syracuse NY

Multiple Sclerosis (MS) is a debilitating autoimmune disease characterized by inflammatory-

mediated demyelination in the central nervous system. Our primary goal is to understand how inflammato-

ry processes can lead to oligodendrocyte dysfunction, axonal degeneration, and demyelination in the CNS.

Our lab discovered that the protein tyrosine phosphatase SHP-1, a major negative regulator of the immune

system, is expressed in oligodendrocytes and plays an essential role in regulating STAT1 and STAT3 acti-

vation in oligodendrocytes. Importantly, it has recently been discovered that both STAT1 and STAT3 are

important transcriptional regulators of metabolic genes. Metabolic regulation is particularly important for

oligodendrocytes because lipid synthesis is required for the production and maintenance of the myelin

sheath and energetic support for axons. Using freshly isolated O4+ oligodendrocytes, we determined that

SHP-1 deficient oligodendrocytes have significantly reduced glycolysis and oxidative phosphorylation

(OXPHOS) relative to oligodendrocytes of wild type mice. We were able to show that in vivo treatment

with IFN- (gamma), a potent activator of STAT1, is able to down regulate wild type oligodendrocyte me-

tabolism without diminishing oligodendrocyte viability. Further, when oligodendrocytes of SHP-1-

deficient mice were treated with IL-10, a potent STAT3 activator, we were able to rescue metabolic defi-

cits in SHP-1-deficient oligodendrocytes suggesting that SHP-1 is a major regulator of oligodendrocyte

metabolism via regulation of STAT1 and STAT3 activation. The understanding of how cytokines such as

IFN- (gamma) and IL-10 modulate metabolism in oligodendrocytes is extremely relevant in the context of

inflammatory mediated demyelinating diseases.

Poster 8

63.

Inflammation, Androgens and Macrophages in the Prostate: Are we missing the link?

Camila Rosat Consiglio1 and Sandra Gollnick1,2

Departments of Immunology1 and Cell Stress Biology2, Roswell Park Cancer Institute,

Buffalo, NY

Prostate cancer (PCa) has the highest malignancy incidence rates in men and is the second leading

cause of male cancer mortality. While the underlying causes of PCa still remain largely unknown, it is

known that androgens, inflammatory mediators and inflammatory cells, including macrophages, are im-

portant players in prostate tumorigenesis. Interestingly, it has been shown that androgen receptor (AR) ex-

pression by macrophages enhances prostatic intraepithelial neoplasia formation in PTEN+/- mice, suggest-

ing that macrophage promotion of tumorigenesis is linked to macrophage AR signaling. Although it is

known that androgens influence immunity, the consequences of AR signaling in macrophages are largely

unknown. Furthermore, the activity and regulation of macrophage AR in the context of prostate tumorigen-

esis is still unclear. To address this gap in the field, the current study aims on analyzing the role of AR in

macrophage homeostasis and in prostate tumorigenesis. Initial results have shown that macrophages ex-

press higher levels of AR protein when compared to monocytes. Polarization of macrophages with either

IFN-γ (gamma) or LPS led to an increase in AR levels. Interestingly, macrophages in G2/M phase of the

cell cycle express higher levels of AR than macrophages in G1 phase, indicating that not only is AR in-

volved in macrophage differentiation and M1 polarization, but also in macrophage cell cycle. Imagestream

analysis of macrophages indicated that AR has strong nuclear localization. In addition, we have confirmed

AR transcriptional activity in bone marrow-derived macrophages using a luciferase assay. Since our results

show that AR is involved in macrophage homeostasis, we next investigated its role in tumor-bearing ani-

mals. We observed an increase in AR levels in tumor-associated macrophages (TAMs) of TRAMP C2

prostate tumors when compared to macrophages from other tissues. This finding was also observed in a

head and neck cancer model (MTERL). This evidence points to a role of the tumor microenvironment in

modulating macrophage AR levels and potentially its activity. Since it is known that macrophage AR facil-

itates prostate tumorigenesis and that macrophage AR levels are higher in the tumor microenvironment, it

is possible that early stages of tumorigenesis could induce macrophage AR signaling, leading to a pro-

tumorigenic phenotype in these cells. Future aims for this project will focus on elucidating macrophage AR

transcriptional targets in homeostasis and in cancer, as well as characterizing macrophage AR function dur-

ing prostate tumorigenesis.

Poster 18

64.

Cell-Intrinsic Glycogen Metabolism Supports Early Activation and Maintains

Metabolite Homeostasis in Dendritic Cells

Phyu Thwe1, Angelo D’ Alessandro2, Princess Rodriguez1, and Eyal Amiel1,3

1University of Vermont, Cellular Molecular and Biomedical Science Program 2University of Colorado Denver

3University of Vermont, College of Nursing and Health Sciences.

As professional antigen presenting cells of the immune system, dendritic cells (DCs) serve as a

bridge between innate and adaptive immune responses. Activation of DCs by a stimulus through toll-like

receptors (TLRs) is coupled with an increase in metabolic demand that is fulfilled by a TLR-driven burst in

glycolytic reprogramming. Up-regulation of glycolysis in activated DCs provides metabolites required for

DC effector function, and inhibition of glycolysis impairs the post-activation survival and effector function

of these cells.

TLR-driven glycolysis is thought to be sustained primarily by increased glucose uptake via the in-

ducible glucose transporter 1 (GLUT1). However, whether glucose is sourced from extracellular or intra-

cellular stores during early glycolytic reprogramming in DCs is still not well-defined. We propose that cell

-intrinsic glycogen metabolism in DCs supports early glycolytic burst that is essential for TLR-driven acti-

vation. The functional importance of glycogen metabolism in the context of DC effector responses has not

been previously described. Our data indicate that glycogen metabolism supports the activation of DCs, par-

ticularly during early activation before the up-regulation of GLUT1 expression. We show that DCs express

the enzymes essential for glycogen metabolism and that glycogen metabolism is regulated upon TLR stim-

ulation. Inhibition of glycogen utilization in DCs impairs the expression of costimulatory molecules CD40

and CD86 in these cells. The ability of DCs to uptake antigens and stimulate T cells is also compromised

upon disruption of glycogen metabolism. In addition, our metabolomics data indicate that glycogen metab-

olism in DCs generates both glycolytic and TCA cycle intermediates and that glycogen-derived carbons

may support metabolic pathways distinct from free glucose catabolism. These data demonstrate that the

glycogen metabolism plays a significant role in metabolic homeostasis in DCs and define a novel metabol-

ic regulatory pathway that supports DC immune function.

Poster 21

65.

Macrophages Negatively Regulate Hematopoietic Stem Cells in Murine

Aplastic Anemia

Angelica Costello, Amanda McCabe, PhD, Julianne N.P. Smith, PhD, & Katherine C. MacNamara, PhD

Albany Medical College, Albany, NY

Aplastic anemia (AA) is a rare bone marrow (BM) failure syndrome characterized by T cell-

mediated bone marrow destruction and pancytopenia. AA can be genetic or acquired, with the latter caused

by such insults as radiation, chemicals, drugs, and infection. T cell-derived interferon-γ (IFN-γ) has been

implicated in driving disease, but the mechanisms of IFN-γ-mediated pathogenesis in hematopoietic failure

during AA are not well understood. We recently identified a role for IFN-γ signaling specifically in macro-

phages (Mϕs) in driving the transient loss of hematopoietic stem cells (HSCs) in murine ehrlichiosis. Here,

in a mouse model of AA, we find that BM-resident Mϕs are maintained in the BM despite the loss of other

hematopoietic cell types, and that this requires IFN-γ. Moreover, Mϕ depletion with clodronate liposomes

or abrogation of IFN-γ signaling in Mϕs during AA rescues the HSC pool and significantly improves sur-

vival. We observed similar numbers of T-bet+ T cells and comparable IFN-γ production in AA mice that

were depleted of Mϕs, suggesting that Mϕs are critical sensors of IFN-γ and drivers of disease during AA.

Despite this, we did not detect a significant reduction in inflammatory proteins when IFN-γ signaling is

abrogated or when Mϕs were depleted. The one notable exception, however, was the chemokine CCL5

(RANTES), which was highly expressed in the BM during AA and significantly reduced in mice depleted

of Mϕs or when Mϕs were unable to respond to IFN-γ. CCL5 production in BM fibroblastic cells is in-

duced via podoplanin (PDPN) signaling. In support of a role of PDPN signaling in driving increased CCL5

during AA, we detected a significant increase in PDPN+ BM-resident Mϕs during AA. Neutralization of

either CCL5 or PDPN during AA rescued HSC numbers. Moreover, anti-PDPN treatment rescued platelet

numbers and significantly improved survival. Altogether, we demonstrate a novel role for IFN-g in AA

pathogenesis whereby IFN-γ promotes increased PDPN+ Mϕs and enhanced production of CCL5, thus

driving HSC loss and thrombocytopenia.

Poster 40

66.

Antagonistic Control of Intestinal Wnt Expression by IBD-Associated Cytokines

Travis Walrath, Stephen Sharp M.D., Shanti D'Souza, Prabhu Tewari, and William O'Connor Jr, Ph.D.

Albany Medical College, Albany NY

Homeostasis of the intestinal epithelium depends on the complex interplay of a variety of mediators

including cytokines that control, among other processes, a gradient of Wnt proteins. Homeostasis can be

disrupted by acute and chronic inflammation such as Inflammatory Bowel Disease (IBD). During IBD a

high concentration of the TH1 and TH17 associated cytokines, interferon gamma (IFN-γ) and interleukin-

17A (IL-17A) respectively, is observed within the colonic lamina propria. IFN-γ has previously been

shown to be cytotoxic to the intestinal epithelium, in part through interference with canonical Wnt signal-

ing; this is associated with increased disease severity in IBD. In murine models of disease, IL-17A has

been shown to lessen the severity of induced IBD.

One important question in the field is how these cytokines govern the intestinal epithelial and adja-

cent lymphoid compartments to either amplify inflammation or support mucosal healing. Toward under-

standing how these cytokines regulate the colonic microenvironment, we undertook a study using murine

knockout strains, primary tissue treated ex vivo, and primary spheroid/organoid cultures treated with IFN-γ,

IL-17A, or a combination of both. We found that IFN-γ reduced canonical Wnt expression while inducing

expression of non-canonical Wnts in primary murine colon tissue. Additionally we found that this altered

balance of wnt expression is dependent on the presence of lymphocytes and further, that wnt5a is specifi-

cally induced in CD19+ cells in the colon. Interestingly, IL-17A was able to oppose the increased expres-

sion of wnt5a. Moreover, mice deficient in IL-17A exhibit enhanced colonic wnt5a and TH1 cell marker

expression in response to Citrobacter rodentium infection, suggesting that IL-17A is opposing infection-

induced wnt5a and the TH1 bias during active colonic inflammation.

Canonical wnts, such as Wnt3a, are necessary for maintaining the potency of stem cells in the in-

testinal stem cell niche and the proliferative capacity of the intestinal epithelium. Non-canonical wnts such

as Wnt5a, have been shown to inhibit canonical wnt signaling and support tissue damage during colitis by

supporting increased IFN-γ production. Therefore, our data suggest that IFN-γ promotes intestinal epitheli-

al dysfunction in part by promoting a shift from canonical to non-canonical Wnt expression and function in

the colon. Furthermore, we hypothesize that IL-17A interferes with the IFN- mediated shift in Wnt ex-

pression, and therefore may reduce IFN- mediated epithelial toxicity and facilitate mucosal healing.

Poster 49

67.

Workshop I

Wayne M. Yokoyama, M.D.

“Opportunities to Study the Immunology of Human Diseases”

Much is known about basic immunology from the study of mouse immune responses. But how do

we translate these basic concepts to the human immune response. I will present examples where the trans-

lation has been achieved in less than obvious ways, highlighting the challenges and opportunities to under-

stand human diseases.

68.

Arielle Ginsberg

Technical Application Specialist

BD Biosciences

“No Title or Abstract Provided”

69.

Symposium III

Cancer Immunity

Chair: Dr. Edith Lord

70.

Exploring the Mechanisms of Neurotrophin-mediated Immune Tolerance and Their Implications

for Autoimmunity and Cancer

Joseph Barbi1, Paolo D. A. Vignali2, Hong Yu2, Fan Pan2, Drew Pardoll2 1Roswell Park Cancer Institute, Department of Immunology, Buffalo, NY 2Johns Hopkins

University School of Medicine, Department of Oncology, Baltimore, MD

Regulatory T cells (Tregs) enforce immune homeostasis and self-tolerance and inhib-

it both natural and induced anti-tumor immunity. Consequently, there is considerable inter-

est in therapeutic Treg enhancement or antagonism to treat inflammatory/autoimmune dis-

eases and cancer, respectively. Targeting Tregs requires a comprehensive understanding of

the factors important for their maintenance, differentiation and function. Here we report that

neuritin, a conserved, gpi-anchored molecule important for the development, survival and

function of neurons, is highly expressed by induced and natural Tregs. Neuritin expression

was also found to promote the maintenance and function of the Treg pool. Additionally,

neuritin ablation undercut Treg suppression in vivo, and tumors grew poorly in in the ab-

sence of neuritin, which permitted the mobilization of more robust anti-tumor immunity.

This was marked by enhanced pro-inflammatory cytokine production by tumor-associated

leukocytes and reduced expression of the checkpoint inhibitor PD-1. Importantly, we also

found that in Tregs, neuritin expression was closely linked to the functional differentiation

of Tregs into an activated, peripheral tissue-homing phenotype. As such, neuritin deficiency

resulted in an imbalance between “central”- and “effector”-like Treg populations and func-

tions. A role for neuritin in the modulation of co-stimulation pathways in favor of tolero-

genic outcomes was also identified. These findings characterize this neurotrophin as a hith-

erto unappreciated immunoregulatory molecule and a potential target for therapies aimed at

the fine-tuning of Treg function in cancer and inflammatory/autoimmune diseases.

71.

In situ analysis of multiple immune cells in tumor and microenvironment

Fiona Ginty, Ph.D.

GE Global Research, Niskayuna, NY 12309

Tumor immune response is highly complex, involving several distinct cell lineages,

cell-cell interactions and regulation. Variable cell density, infiltration and functional status

can determine tumor progression and immunotherapy response. Routine IHC methodolo-

gies are limited in their ability to quantify cellular diversity, and genomic methods may not

account for spatial location or cell activation status. Multiplexed analysis methods provide a

way to combine multiple markers and distinguish/quantify different cell types, and poten-

tially decipher immunotherapy response mechanisms. This presentation will discuss novel

approaches for multiplexed analysis of immune cell markers, quantifying cell type and

number in the tumor and microenvironment.

72.

Symposium IV

Immune Mediated Disease

Chair: Dr. Wayne Yokoyama

73.

Reactive Oxygen Species Induce Virus-independent MAVS-oligomerization in

Systemic Lupus Erythematosus

Andreas Koenig1, Theresa Montgomery1, Michael Murphy2, Richard Hartley3, Ryan Kelly4,

Andras Perl4, Ralph Budd5, and Iwona A. Buskiewicz1*

Department of 1Pathology and 5Medicine, Vermont Center for Immunology and Infectious

Diseases, University of Vermont, Burlington, USA; 2MRC Mitochondrial Biology

Unit, Wellcome Trust/MRC, Cambridge, UK; 3WestCHEM School of Chemistry,

University of Glasgow, Glasgow, UK; 4Upstate University Hospital, State University of

New York, Syracuse, USA

Systemic lupus erythematosus (SLE) is a complex autoimmune disease associated

with multiple immunologic abnormalities, among which upregulation of type I interferon

(IFN-I) genes correlates strongly with disease activity. Here we show that mitochondrial an-

tiviral signaling protein (MAVS), which normally forms a complex with retinoic acid gene

I (RIG-I)-like helicases during viral infection, can be activated by oxidative stress

alone. We observe that MAVS oligomerization on the surface of the mitochondrial outer

membrane by oxidative stress leads to mitochondrial hyperpolarization, decreased ATP pro-

duction, and increased spare respiratory capacity. This virus-independent oligomerization of

MAVS also leads to the secretion of IFN-I and proinflammatory cytokines. Consistent with

this, inhibition of mitochondrial reactive oxygen species (ROS) by the mitochondria-targeted

antioxidant MitoQ prevents oligomerization of MAVS. Patients with SLE are known to mani-

fest hyperpolarized mitochondria and increased ROS in peripheral blood lymphocytes. Our

findings reveal that MAVS is spontaneously oligomerized in peripheral blood mononuclear

cells of SLE patients, but not in matched healthy control individuals. Furthermore, ROS-

mediated MAVS oligomerization and IFN-I production was greatly reduced in cells expressing

a MAVS-C79F variant that occurs in 30% of sub-Saharan Africans and has been linked with

reduced expression of IFN-I and milder SLE. Our findings suggest that spontaneous redox-

induced MAVS oligomerization in SLE patients contributes to the IFN-I signature characteris-

tic of this syndrome.

74.

Increasing Regulatory Tone to Suppress Allergic Inflammatory Responses

in the Lung

Timothy J Chapman PhD, Sara E Hillman, Sara A Knowlden PhD, Jason A Emo MS,

Steve N Georas MD

Division of Pulmonary and Critical Care Medicine, Department of Medicine, University of

Rochester Medical Center, Rochester NY

Allergic inflammation and asthma are the result of inflammatory immune responses

raised against otherwise innocuous inhaled allergens. To prevent these unwanted respons-

es, mucosal immune cells establish an anti-inflammatory state in the lung that must be over-

come in order to induce inflammation. We term this phenomenon ‘regulatory tone’. In a

mouse model of allergic inflammation, low-dose bacterial lipopolysaccharide (LPS) admin-

istered with ovalbumin (OVA) via inhalation to naïve mice resulted in Th2-type allergic in-

flammation. However, prior tolerizing exposure to OVA alone prevented subsequent LPS/

OVA-induced allergic inflammation. Raising the dose of adjuvant in tolerized mice result-

ed in breakdown of tolerance and allergic inflammation with a mixed Th2/Th17 phenotype.

Therefore, increasing regulatory tone in the lung can prevent sensitizing exposures to aller-

gens. However, allergen exposure in the context of more severe inflammation can result in

breakdown of tolerance and a qualitative difference in the resulting immune response.

These data suggest that the balance between regulatory and inflammatory cells in the lung

is a critical feature in determining the outcome of inhaled allergen exposure.

75.

Industry Panel

“Careers Outside of Academia”

Participants

Patrick Murphy, Ph.D. (BioLegend, Inc.)

Fiona Ginty, Ph.D. (GE Global)

Erik Puffer, Ph.D. (BD Biosciences)

Olivia Schneider, Ph.D. (Shenandoah)

76.

Workshop II

Thomas J. Braciale , M.D., Ph.D.

Carter Immunology Center, Department of Pathology and Molecular Medicine

University of Virginia Health Systems

“The Business End of Academic Research”

This workshop is aimed at junior investigators in the process of starting up (or contemplating the

startup) an independent biomedical research laboratory. We will track the process of starting up a laborato-

ry from reviewing an offer of employment letter, negotiating a startup package, to hiring a technician, and

recruiting graduate students and fellows. We will use as a starting point for discussion, a video by a junior

investigator recently appointed to an independent academic position, who will describe her thoughts and

impressions about the business of starting up a research laboratory.

77.

Workshop III

“Granstmanship, Funding, and Mock Peer Review”

NIH Members

Timothy Gondre-Lewis, Ph.D. (Program Officer/COR/ DAIT Training Officer)

Thomas J. Palker, Ph.D. (Program Officer)

B. Duane Price, Ph.D. (Senior Scientific Review Officer)

James Snyder, Ph.D. (Scientific Review Officer)

Mock Peer Review Panelists

Margaret Bynoe, Ph.D. (Cornell University)

Richard I. Enelow, M.D. (Dartmouth Medical College)

Jonathan A. Harton, Ph.D. (Albany Medical College)

Nicholas J. Mantis, Ph.D. (Wadsworth Center/NYSDOH)

78.

Symposium V

Immunity in Infectious Disease

Chair: Dr. Thomas Braciale

79.

Control of the CD8+ Effector Population in Influenza Infection

Richard I. Enelow, M.D.

Dartmouth Medical College

The controlled contraction of immune responses during the clearance of respiratory virus infection

is critical to resolution of inflammation and the limitation of lung injury. The kinetics of antiviral effector

CD8+ T cell contraction, and dampening of effector function, appears to be programmed during the initial

activation of naive T cells, and possibly prior. Among other negative regulators of immune responses, it

has long been appreciated that TNF-alpha signaling plays an important role in contraction of CD8+ effec-

tor T cells, though the precise mechanisms involved remain unclear. We've shown that the critical timing

of TNF-programmed contraction occurs within the first 24-48 hours after initial antigen recognition, and

plays little immunoregulatory role thereafter. Furthermore, the critical source of this early burst of TNF

production is the naive CD8+ T cell upon initial antigen recognition. We hypothesize that the ability of the

naive CD8+ T cell to produce an early TNF burst is critically dependent upon homeostatic type I interferon

signaling in the host milieu prior to infection. The IFN-mediated regulation of the early burst of TNF-a by

naive CD8+ T cells is quite complex, and appears dependent upon post-thymic peripheral T cell

"licensing", during which the impact of constitutive low-level IFN production confers the tendency to

mount this important early TNF response. Together theses activities have a direct impact on the kinetics of

the expansion and contraction of the T cell responses and effector activities during and after viral infection,

serving to limit lung injury and immunopathology.

80.

Mast Cells in Kaposi’s Sarcoma – A Model for the Pathogenesis of

KSHV-driven Oncogenesis

Arturo Barbachano-Guerrero and Christine A. King

Department of Microbiology and Immunology, SUNY Upstate Medical University,

Syracuse, NY, USA

Kaposi sarcoma (KS) is a multicentric, inflammatory-driven, angioproliferative tumor of endotheli-

al cell (EC) origin where KS-associated herpesvirus (KSHV) is the etiological agent. This “hemorrhagic

sarcoma” is characterized by, and dependent upon, KSHV viral infection and inflammatory mediators for

proliferation and survival. Sources for infectious KSHV and inflammatory factors required for initiation,

expansion, and maintenance of KSHV-induced KS lesions are not fully understood. Here we demonstrate

for the first time that mast cells (MCs), key innate immune cells with potent inflammatory and angiogenic

responses, are key components of KS lesions. In this study, we analyzed cutaneous, gut, lymph node and

lung lesions from patients with classic KS for the presence of MCs by IHC staining using the MC specific

anti-tryptase (a major mast cell granule product) and KSHV LANA-1 antibodies. We found that MCs

were present in all lesions, independent of location or stage and localized in large numbers to highly vascu-

lar areas. IHC clearly demonstrated a sub-population of MCs in lesions that were positive for KSHV LA-

NA. MCs exhibited extensive degranulation that resulted in the release of potent pre-formed pro-

inflammatory tryptase and histamine. MCs were fully permissive to KSHV infection in vitro with de novo

viral gene expression observed as early as 6h post-infection (p.i) and culminating in release of infectious

progeny capable of establishing latent infection in primary human endothelial cells by 24 h p.i.. Taken to-

gether, we demonstrate for the first time that human MCs latently infected with KSHV are associated with

KS in all locations and stages of lesions development, in vivo. This suggests a model where MCs accumu-

late at the vascular interface between healthy and malignant KS tissue, are a potential source of infectious

virus and also act as inflammatory drivers needed to maintain and advance lesions.

81.

Toxoplasma Infections of the Nervous System

Ira J. Blader

Department of Microbiology and Immunology


Toxoplasma gondii infections can be severe and even fatal after they reactivate in immunocompro-

mised individuals. In most cases, these patients develop toxoplasmic encephalitis who clinically present

with dizziness, headaches, and seizures. Why patients develop these symptoms is unknown and has been

the focus of work in our laboratory for the past several years. To address this question, we used a murine

model of toxoplasmic encephalitis and found that the mice develop spontaneous seizures that are long last-

ing and severe. Since seizures can develop due to decreased inhibitory neuronal synaptic activity, we in-

vestigated the structure and function of GABAergic synapses, which are the major inhibitory synapses in

the brain. We found that the localization of GAD67, which is the primary GABA biosynthetic enzyme in

the brain, changes from being clustered at synaptic termini to be diffusely localized throughout the axon

and synapse. We further find that Toxoplasma decreases host neuronal GABAergic signaling, which is

consistent with the hypothesis that GAD67 mislocalization reduces host GABA receptor signaling. Toxo-

plasma triggering of seizures and GAD67 mislocalization occurs in mice infected with type II but not type

III strain parasites indicating the involvement of a polymorphic parasite factor . Given that these strains

generate significant differences in inflammatory responses, we compared the activation state of microglia/

monocytes in the brains of these mice and observed that strains that triggered seizures also had more severe

microglial activation. Finally, we find that a microglia inhibitor reduces both seizures and GAD67 mislo-

calization. Taken together, these data indicate that differences in inflammatory responses within the brain

in Toxoplasma-infected mice lead to specific changes in host GABAergic signaling that likely underlie sei-

zures in toxoplasmosis patients.

82.

Keynote Speaker

Thomas J. Braciale, M.D., Ph.D. Director, Emeritus

Carter Immunology Center

Professor of Pathology and Molecular Medicine

University of Virginia

“Dendritic Cells in the Host Response to Stress or Dendritic Cells Can Do More Than Present Antigens”

Dendritic cells (DC) have long been appreciated as the predominant antigen presenting

cells for the induction of primary adaptive immune responses to many invading microorganisms.

In this capacity, DC represent important first responders to the "stress" of microbial infection.

This presentation will describe recent studies analyzing the role of distinct subsets of respiratory

DC in initiating and regulating the adaptive immune CD8+ T cell response to influenza A virus

infection. However microbial infection is only one of many types of stress encountered in the en-

vironment. Related studies will describe a novel role for DC in regulating stress erythropoiesis,

that is the production of erythrocytes following hypoxia and tissue inflammation. (Supported by

USPHS grants AI-15608 and HL-33391).

Dr. Braciale’s lab is interested in the host immune response to virus infection—specifically, studying the role of the

adaptive immune response in the clearance of both virus and virus-infected cells from the body with much of the

work focused on infection of the respiratory tract by Influenza Virus and Respiratory Syncytial Virus.

83.

2015 NYIC Poster Numbers

AAI Young Investigator Award and Oral Poster Presentation eBioscience Travel Award and Oral Poster Presentation

Poster No:

1 Safiehkhatoon Moshkani

2 Megan Peppenelli

3 Lauren M. Webb

4 Kelly L. Singel

5 Oyebola Oyesola

6 Weishan Huang

7 James Emo

8 Scott B. Minchenberg

9 Abhinit Nagar

10 Taylor J. Moon

11 Christina M. Post

12 Alicia Soucy

13 Stephanie L. Schell

14 Danielle Y.F. Twum

15 Praneet Kaur Sandhu

16 Tariq A. Bhat

17 Jennifer Yates

18 Camila Rosat Consiglio

19 Bhuvana Katkere

20 Victoria L. DeVault

21 Phyu Thwe

22 Kevin Kenderes

23 Adam Utley

24 Colin A. Powers

25 Olesea Cojohari

Poster No:

26 Adaobi Amobi

27 Amy Ku

28 Catherine G. Burke

29 Aditi Murthy

30 Kathy A. Green

31 Arturo Barbachano-Guerrero

32 Tiffany Coupet/Clair Palmer

33 Janell Veazey

34 Danielle E. Baranova

35 Donald Steiner

36 Chenyang Jiang

37 Sadikshya Bhandari

38 Joel R. Wilmore

39 Angelene F. Richards

40 Angelica Costello

41 Amanda Poon

42 Kirsten E. Dostie

43 Amy Thees

44 Amber Papillion

45 Jennifer Vella

46 Margaret L. Barlow

47 Shivana M. Maharaj

48 Anand Sharda

49 Travis Walrath

84.

Poster 1.

Highly Attenuated Vesicular Stomatitis Virus-Based Vectors as Therapeutic Vaccines for Chronic

Hepatitis B

Safiehkhatoon Moshkani1, Sabine Lang2, John K. Rose2, Michael D. Robek1 1Department of Immunology & Microbial Disease, Albany Medical College, Albany, NY

2Department of Pathology, Yale University, New Haven, CT

Despite the availability of an effective preventative vaccine, chronic hepatitis B virus (HBV) infec-

tion affects over 240 million people worldwide and causes serious diseases such as liver cirrhosis and

hepatocellular carcinoma. Current treatments for chronic hepatitis B typically control but do not eliminate

the infection. The immune response to HBV in chronically infected people is typically weak and ineffec-

tive; therefore, therapeutic vaccines that elicit a functional antiviral immune response may be an important

component of a treatment regimen that is capable of curing chronic HBV. Virus-based vectors are highly

immunogenic compared to non-replicating vaccine platforms, but safety concerns can limit their use. We

previously demonstrated that recombinant wild-type vesicular stomatitis virus (VSV) expressing the HBV

middle surface envelope protein (MHBs) elicits functional virus-specific T cell and antibody responses in

mouse models of acute and chronic HBV replication. However, VSV has some undesirable pathogenic

properties, and the use of this platform in humans will require further attenuation of the vector. We there-

fore generated a VSV-based vector that expresses MHBs and contains two attenuating mutations that syn-

ergistically reduce virus replication: translocation of the nucleocapsid gene to the fourth genome position

and truncation of the glycoprotein cytoplasmic tail. This vector was evaluated for immunogenicity, patho-

genesis, and anti-HBV function in mice. Compared to wild-type VSV, the highly attenuated VSV vector

displayed markedly reduced pathogenesis but induced similar MHBs-specific CD8+ T cell and antibody

responses. The memory CD8+ T cell responses elicited by the highly attenuated VSV vector prevented

HBV replication in mice that were subsequently challenged by transduction with adeno-associated virus

(AAV) encoding the HBV genome (AAV-HBV). In mice in which persistent HBV replication was first

established by AAV-HBV transduction, treatment with the highly attenuated VSV induced MHBs-specific

CD8+ T cell responses that corresponded with a reduction in serum and liver HBV antigens and nucleic

acids, respectively. The ability of VSV to induce a robust multi-specific T cell response capable of control-

ling HBV replication combined with the improved safety profile of the highly attenuated vector suggests

this platform offers a potential strategy for HBV therapeutic vaccination.

85.

Poster 2.

HCMV Mediated Signaling Induces The Synthesis Of Select Akt-Dependent Antiapoptotic Proteins

During Entry To Promote Survival Of Short-Lived Monocytes.

Megan Peppenelli (1), Kyle Arend (2), Olesea Cojohari (1), Nathaniel Moorman (2) and Gary Chan (1).

(1): SUNY Upstate Medical University, Syracuse, New York, United States

(2): University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States

HCMV infection of immunocompromised individuals often leads to multi-system organ failure.

The development of multi-system organ failure is dependent on the ability of HCMV to spread to peripher-

al organs, which is mediated by blood monocytes. In order for monocytes to mediate spread, we have pre-

viously shown HCMV to extend the short 48-hour lifespan of monocytes. Mechanistically, HCMV upregu-

lated two specific cellular antiapoptotic proteins, myeloid leukemia sequence 1 (Mcl-1) and heat shock

protein 27 (HSP27), to block two proteolytic cleavages necessary for the formation of fully active caspase

3 and the subsequent initiation of apoptosis. We now show that HCMV more robustly upregulated Mcl-1

when compared to normal myeloid growth factors and that HCMV was the only myeloid survival factor to

rapidly induce HSP27 prior to the 48-h cell fate checkpoint. We determined that HCMV glycoproteins gB

and gH signal through the cellular epidermal growth factor receptor (EGFR) and αvβ3 integrin, respective-

ly, during viral entry in order to drive the increase of Mcl-1 and HSP27 in an Akt-dependent manner. Alt-

hough Akt is known to regulate protein stability and transcription, we found that gB- and gH-initiated sig-

naling preferentially and cooperatively stimulated the synthesis of Mcl-1 and HSP27 through mTOR-

mediated translation. Overall, these data suggest that the unique signaling network generated during the

viral entry process stimulates the upregulation of select antiapoptotic proteins allowing for the differentia-

tion of short-lived monocytes into long-lived macrophages, a key step in the viral dissemination strategy.

Contact: Gary Chan, [email protected]

86.

Poster 3.

Role of Notch signaling in basophils in Type 2 inflammation

Lauren M. Webb1, Rebecca L. Cubitt1, Everett Henry2, Mark C. Siracusa2 and Elia D. Tait Wojno1

1Baker Institute for Animal Health and Department of Microbiology and Immunology, College of Veteri-

nary Medicine, Cornell University, Ithaca, NY, USA. 2Center for Immunity and Inflammation, New Jersey

Medical School, Rutgers Biomedical and Health Sciences, Rutgers, The State University of New Jersey,

Newark, New Jersey, USA.

Basophils are a rare granulocyte population, making up just 1% of circulating immune cells. De-

spite their rarity, basophils expand and contribute to the development of host-protective type 2 inflamma-

tion and worm expulsion following infection with Trichuris muris, a murine model of human whipworm.

However, the pathways that regulate basophil expansion and function in this setting remain unclear. Previ-

ous work has shown that the Notch signaling pathway is important in lymphopoiesis and particularly in the

function of T helper 2 cells, but little is known about its role in innate cell function. Here, we have investi-

gated the importance of the Notch signaling pathway in controlling the function of basophils during Type 2

inflammation. We show for the first time that basophils selectively upregulate Notch under Type 2 inflam-

matory conditions. Further, while Notch is not required for the steady-state development of basophils, it is

required for their optimal cytokine production in response to such damage signals as IL-33. We are now

actively investigating how other facets of basophil function are affected by inhibition of Notch signaling

and are characterizing how the selective deletion of Notch signaling in basophils affects anti-helminth re-

sponses and worm expulsion during T. muris infection. These findings have wide reaching implications

both for our continued understanding of innate cell function in Type 2 immunity but also in the potential

development of therapeutics aimed at targeting Type 2 inflammation.

87.

Poster 4.

A Novel Barrier to Endogenous Anti-Tumor Immunity: Ovarian cancer ascites-activated

neutrophils suppress T cell proliferation in a contact-dependent mechanism

Kelly L. Singel1, ANM Nazmul H. Khan2, Kirsten B. Moysich3, Kunle Odunsi4,5, Brahm H. Segal1,2,6

Departments of 1Immunology, 2Medicine, 3Cancer Prevention and Control, 4Gynecologic Oncology, and

the 5Center for Immunotherapy, Roswell Park Cancer Institute, Buffalo, NY 6Department of Medicine, University at Buffalo Jacobs School of Medicine and Biomedical Sciences,

Buffalo, NY

Neutrophils are the first responders to infection and injury, and critical for antimicrobial host de-

fense. Through the generation of reactive oxidants, activation of granular constituents, and neutrophil ex-

tracellular traps (NETs), neutrophils target microbes and prevent their dissemination. While these path-

ways are beneficial in the context of trauma and infection, their roles in the context of tumor are less un-

derstood. Ovarian cancer (OC) is often diagnosed at advanced stages and presents with ascites. Necrosis is

a hallmark of advanced cancer and releases DAMPs that activate innate immune responses. Cytotoxic T

lymphocyte (CTL) immunity is critical in OC, and barriers to durable anti-tumor immunity include TAMs,

MDSCs, and Tregs. While activated neutrophils can kill tumor cells, knowledge is limited on the role of

activated neutrophils in the tumor microenvironment.

Our prior studies showed that granulocytic cells from ascites of patients with newly diagnosed OC

variably suppressed stimulated normal donor T cell proliferation ex vivo. In ascites, ~90% of cells were

inflammatory (CD45+) with varying proportions of neutrophils, monocytes, and lymphocytes. Neutrophils

comprised ~15% of CD45+ cells and the neutrophil:CD8+ T cell ratio was 1.5:1. In ex vivo studies, cell-free

ascites (CFA) attracted normal donor neutrophils (NDN) and induced NETs. We next evaluated the effects

of CFA-treated NDN on T cell proliferation. In co-culture studies, a subset of CFA (5/10 tested) activated

NDN to completely suppress CD3/CD28-stimulated T cell proliferation. Neither CFA nor NDN alone im-

paired T cell proliferation as measured by [3H] thymidine at a 1:1 target:effector cell ratio. Suppression did

not affect T cell viability or induce apoptosis, but required T cell contact with NDN in the presence of

CFA. In addition, we found that PD-1 expression on the suppressed T cells phenocopied the unstimulated

controls. When T cells were CD3/CD28-stimulated overnight before exposure to CFA and NDN, T cell

proliferation was not inhibited by the addition of CFA and NDN, suggesting that CFA-treated NDN inter-

rupt T cell proliferation at an early stage of activation.

These results support a model in which neutrophils in the ascites of a subset of OC patients sup-

press T cell responses, which may be an important barrier to endogenous anti-tumor immunity and immu-

notherapy. Further studies will identify ascites constituents that activate neutrophils and mechanisms for

neutrophil-mediated T cell suppression. This work may lead to novel prognostic biomarkers regarding in-

flammatory responses in the OC tumor microenvironment and therapeutic approaches that target neutro-

phils.

Supported by the NIH (R01, CA188900; T32, CA085183), the RPCI-UPCI Ovarian Cancer SPORE (P50

CA159981), and the Cancer Center Support Grant to RPCI (CA016056).

88.

Poster 5.

The prostaglandin D2 receptor CRTH2 mediates interleukin-33-elicited group 2 innate lymphoid cell

accumulation in tissues

Oyebola Oyesola1, Lauren M. Webb1, Rebecca Cubitt1, Elia Tait Wojno1 1 Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca NY 14853,

United States

Group 2 innate lymphoid cells (ILC2s) are rare innate immune cells that contribute to the develop-

ment of type 2 inflammation. These cells and associated type 2 inflammation promote allergic lung in-

flammation, which affects nearly 8% of the general population in the United States. Previous studies have

shown that the accumulation of ILC2s at inflammatory sites is mediated by epithelial cell-derived cyto-

kines and alarmins, particularly interleukin-33 (IL-33). In addition, prostaglandin D2 (PGD2), a bioactive

lipid produced by mast cells, induces accumulation of ILC2s at inflammatory sites by binding to its recep-

tor chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2), which is expressed

by ILC2s. However, whether the IL-33-IL-33 receptor (IL-33R) and PGD2-CRTH2 pathways coordinate or

intersect to regulate ILC2 responses during type 2 lung inflammation has not fully been explored in vivo.

In this study, we investigated the role of the PGD2-CRTH2 pathway during IL-33-elicited ILC2 accumula-

tion and type 2 inflammation in the lung. When wild-type mice were treated either systemically by intra-

peritoneal injection or locally by intranasal injection with recombinant murine IL-33 (rmIL-33) to induce

type 2 inflammation, ILC2 frequency and number increased in the lung parenchyma. In contrast, ILC2s

accumulated in the lung of mice deficient in CRTH2 (CRTH2KO) following intranasal but not intraperito-

neal treatment with rmIL-33. The defect in ILC2 accumulation in the CRTH2KO lung following systemic

rmIL-33 treatment was not due to differences in expression of the IL-33R, cell death, apoptosis, or cell

proliferation. Therefore, our data suggest that the PGD2-CRTH2 pathway acts downstream of IL-33 to par-

tially mediate ILC2 accumulation in the lung during type 2 inflammation that originates in the periphery,

acting via pathways other than those controlling cell responsiveness to IL-33, death, or proliferation. Fu-

ture studies will focus on determining if ILC2 accumulation in the lung in response to IL-33 is mediated

via CRTH2-dependent migration to or retention in the lung. A better understanding of how cytokines and

bioactive lipid mediators interact to regulate ILC2 responses will be important in informing the use and

development of drugs that target these pathways to treat ILC2-associated type 2 airway inflammation.

89.

Poster 6.

ITK signaling via IRF4 regulates the development and function of type 1 regulatory T cells

Weishan Huang*, Sabrina Solouki, Nicholas Koylass, and Avery August*

Department of Microbiology and Immunology, Cornell University, Ithaca, NY 14853.

Type 1 regulatory T (Tr1) cells lack the expression of Foxp3 but have significant regulatory func-

tion in suppressing inflammation and promoting tolerance, in part via their expression of the immunosup-

pressive cytokine IL-10. Tr1 cells differentiate in response to signals engaging T cell receptor (TCR) and/

or the regulatory cytokine milieu. The non-receptor tyrosine kinase ITK is a key modulator downstream of

TCR, playing critical roles in T cell development and function. Using mouse models carrying Foxp3RFP

and IL-10GFP dual reporters, we found that, in the absence of ITK, TCR activation-driven development of

Foxp3- IL-10+ Tr1 cells is severely impaired in various organs (spleen, blood, lung, gut, and fat). Itk-/-

mice were also deficient in mounting a mucosal Tr1 cell response during parasitic (Nippostrongylus Brasil-

iensis) and viral (Influenza A) infections.

Naïve Itk-/- thymic and splenic Foxp3- CD4+ T cells also exhibited severe deficiency in Tr1 differ-

entiation under Tr1 polarizing condition. Although Itk-/- CD4+ T cells proliferated under Tr1 differentiating

conditions, they failed to up-regulate IL-10, and Tr1 cell markers LAG3, CD49b, ICOS, PD-1, c-Maf,

AHR, and IRF4 to levels observed in WT cells, suggesting that ITK is critical for Tr1 cell fate program-

ming. Utilizing a transgenic mouse model carrying an allele sensitive mutant of ITK (ITKas) that allows

ITK kinase specific blockade by a small molecule 3MB-PP1, we determined that the expression of the

aforementioned markers, as well as the balance between IL-10 and IFN-g (gamma) production during Tr1

differentiation, are dependent on ITK kinase activity. Furthermore, using cells from an ITKas-Foxp3RFP/IL

-17AGFP dual reporter mouse model, we find that ITK kinase activity is required for optimal Th17 trans-

differentiation to Tr1 cells. We also find that inhibiting ITK kinase activity diminished Tr1 differentiation

by human CD4+ T cells. The requirement for ITK function during Tr1 cell development can be restored by

the expression of the transcription factor IRF4. Finally, specifically targeting ITK kinase activity in already

differentiated Tr1 cells diminished their suppressive function.

We conclude that the TCR/ITK/IRF4 pathway is required for the development and function of Tr1

cells, which may be targeted to modulate regulatory immunity for clinical benefit.

*Correspondence: Weishan Huang ([email protected]) and Avery August

([email protected]).

mailto:[email protected]


90.

Poster 7.

Enrichment for TNF-α Producing Naïve T Cells in Premature Infants Is Not Secondary to Bystander

Activation from Clinical Chorioamnionitis

Jason Emo, Natalie Martinez, Heidie Hyuck, Gloria Pryhuber, Dave Topham, Kristin Scheible

University of Rochester School of Medicine and Dentistry, Rochester, NY, United States.

Background: Newborn T cells are predominantly naïve, but interestingly naïve T cells from preterm infants

have a higher capacity to make TNF-α than full term babies. In-utero infection (chorioamnionitis), a com-

mon condition causing premature delivery, exposes the fetus to inflammatory cytokines such as IL-6, IL-1-

β, IL-8, and TNF-α. These cytokines are known to promote bystander activation of CD4+T cells.

Hypothesis: We hypothesized that enhanced TNF-α-producing T cells observed in premature infants at

birth, is due to inflammatory cytokine conditions present in chorioamnionitis.

Objective: To identify plasma cytokine profiles at birth that associate with prematurity or chorioamnionitis,

and determine the relationship between these profiles and frequency of TNF-α naïve CD4+ T cells.

Design/Methods: Umbilical cord blood plasma and mononuclear cells (UCMC) from 26 chorio-exposed

and 26 non-exposed infants was collected from 32 preterm (<32 0/7 weeks gestational age) and 20 full

term (>37 0/7 weeks gestational age) infants. Multiplex assays, customized to detect T-cell activating and

secreted cytokines, were performed on plasma samples. Functional profiles of CD4+T cells in matched

isolated UCMC was performed using flow cytometry.

Results: CD4+CD45RA+TNF-α+ T cells trended higher in infants exposed to chorioamnionitis. Naïve

CD4+ T cells from preterm babies showed higher frequencies of TNF-α+ producers at lower gestational

ages (p<0.0001), which was still significant when controlling for chorioamnionitis. Principle Component

Analysis (PCA) of plasma cytokines separated subjects into several clusters. Prematurity associated with

PCA1, which represented the greatest (22%) variability. PCA5, associated with chorioamnionitis, but con-

tributed to only 7% of the variability. CD4+CD45RA+TNF-α+ T cells were significantly lower (p=0.05) at

birth in subjects that clustered with PCA1.

Conclusion: Our results do not support the hypothesis that enrichment for naïve TNF-α+ producing T cells

in preterm infant cord blood is a result of bystander activation from in utero infection. Naïve TNF-α-

producing T cells are highly correlated with younger gestational age, which suggests this may be a devel-

opmentally-related finding. Plasma cytokine profiles in preterm infants are associated with reduced TNF-

α+ T cells, and may be protective against propagation of inflammation by T cells.

91.

Poster 8.

Cytokine-mediated regulation of oligodendrocyte metabolism

Scott B. Minchenberg, Anthony F. Paredes, and Paul T. Massa

SUNY Upstate Medical University, Department of Microbiology & Immunology, Syracuse NY

Multiple Sclerosis (MS) is a debilitating autoimmune disease characterized by inflammatory-

mediated demyelination in the central nervous system. Our primary goal is to understand how inflammato-

ry processes can lead to oligodendrocyte dysfunction, axonal degeneration, and demyelination in the CNS.

Our lab discovered that the protein tyrosine phosphatase SHP-1, a major negative regulator of the immune

system, is expressed in oligodendrocytes and plays an essential role in regulating STAT1 and STAT3 acti-

vation in oligodendrocytes. Importantly, it has recently been discovered that both STAT1 and STAT3 are

important transcriptional regulators of metabolic genes. Metabolic regulation is particularly important for

oligodendrocytes because lipid synthesis is required for the production and maintenance of the myelin

sheath and energetic support for axons. Using freshly isolated O4+ oligodendrocytes, we determined that

SHP-1 deficient oligodendrocytes have significantly reduced glycolysis and oxidative phosphorylation

(OXPHOS) relative to oligodendrocytes of wild type mice. We were able to show that in vivo treatment

with IFN-g (gamma), a potent activator of STAT1, is able to down regulate wild type oligodendrocyte me-

tabolism without diminishing oligodendrocyte viability. Further, when oligodendrocytes of SHP-1-

deficient mice were treated with IL-10, a potent STAT3 activator, we were able to rescue metabolic defi-

cits in SHP-1-deficient oligodendrocytes suggesting that SHP-1 is a major regulator of oligodendrocyte

metabolism via regulation of STAT1 and STAT3 activation. The understanding of how cytokines such as

IFN-g (gamma) and IL-10 modulate metabolism in oligodendrocytes is extremely relevant in the context of

inflammatory mediated demyelinating diseases.

92.

Poster 9.

NLRP3 inflammasome activation is modulated by a non-Pyrin domain cysteine

Abhinit Nagar, Tabassum Rahman, Jonathan A. Harton, PhD

Department for Immunology and Microbial diseases


NLRP3 inflammasome activation results in activation of caspase-1 that aids in eliciting inflamma-

tory response by cytokine maturation. While structurally divergent agonists activate the NLRP3 inflam-

masome, the specific mechanism is still unresolved. The crystal structure of NLRP3 highlights the pres-

ence of a disulfide bond between conserved cysteines 8 and 108 in and adjacent to the N-terminal pyrin

domain (PYD) which may be important for NLRP3:ASC interaction and NLRP3 processing of IL-1β

(Beta). Many NLRP3 agonists trigger production of reactive oxygen species (ROS) which has been specu-

lated to drive the C8-108 disulfide bond. C8 is highly conserved across species. We explored the contribu-

tion of these cysteines to the NLRP3 inflammasome by evaluating multiple cysteine substitution mutants.

We evaluated formation of a perinuclear, oligomeric NLRP3:ASC complex (the inflammasome ‘speck’) by

Time Of Flight Inflammasome Evaluation (TOFIE) and NLRP3 processing of IL-1β by inflammasome re-

constitution. Here, we provide evidence that C8 and C108 are not important for NLRP3:ASC interaction.

However, NLRP3 activation of IL-1β processing is regulated by C108. These results highlight potential

inflammasome regulatory role for C108 adjacent to NLRP3 PYD and suggest that the spacer sequence con-

taining C108 likely comprises an uncharacterized regulatory domain of NLRP3. Further, as C108 appear to

differentially regulate the inflammasome response to distinct agonists, these results discount the hypothesis

that NLRP3 inflammasome agonists share a unified activation mechanism.

93.

Poster 10.

P2Y2 Regulation of CXCR4 Function

Taylor J. Moon and Michael R. Elliott

David H. Smith Center for Vaccine Biology & Immunology

University of Rochester, Rochester, NY

CXCR4 is a G-protein-coupled chemokine receptor with critical roles in immune cell development,

survival, and tissue homing. CXCR4 is also one of two major co-receptors for HIV-1 infection of human

cells. Therefore, a better understanding of the molecular regulation of CXCR4 expression and function is

important for developing therapeutic strategies to target this receptor as a means for altering immune cell

motility and HIV-1 infection. Here we demonstrate a novel role for the purinergic receptor P2Y2 in regu-

lating CXCR4 signaling in lymphocytes and myeloid cells. P2Y2 is a G-protein-coupled receptor respon-

sive to extracellular nucleotides including ATP and UTP, and P2Y2 signaling has recently been reported to

enhance HIV-1 infection of CD4+ T cells. Our work shows that human Jurkat T cells stably expressing

P2Y2 have decreased CXCR4-dependent migration, and concurrently human THP-1 monocyte cells with

siRNA knockdown of P2Y2 exhibit increased CXCR4 function. P2Y2 regulation of migration is specific

to CXCR4, because migration to other chemoattractants is not affected by over expression or knock down

of P2Y2. Additionally, after testing HIV-1 infection of human Jurkat T cells, we found that P2Y2 expres-

sion also significantly decreases infection over time compared to P2Y2 null controls. Interestingly, mouse

P2Y2 does not inhibit HIV-1 infection, nor does a truncated form of human P2Y2 lacking the cytoplasmic

tail (aa 1-310). From these data, we hypothesize that human P2Y2 signaling negatively regulates the ex-

pression and function of CXCR4 at the surface of lymphocytes. Elucidating the specific mechanisms re-

sponsible for P2Y2 regulation of CXCR4 will help determine whether P2Y2 is an attractive target for in-

hibiting HIV-1 entry as well as for therapeutic strategies aimed at modulating immune cell mobilization.

94.

Poster 11.

The ancestral environment shapes antiviral CD8+ T cell responses across generations

Christina M. Post, Jason Myers, and B. Paige Lawrence

Department of Environmental Medicine, University of Rochester School of Medicine & Dentistry,

Rochester NY

Recent studies have linked health fates of great-grandchildren to environmental exposures of their

great grandparents. However, few studies have considered whether ancestral exposures influence the im-

mune system across generations. Here we report novel findings regarding transgenerational transmission of

altered T cell responses resulting from maternal (F0) exposure to an environmentally relevant aryl hydro-

carbon receptor (AHR) ligand. The AHR is a transcriptional regulator that plays diverse roles in cellular

function, including modulating immune responses. AHR ligands comprise several classes of pollutants,

such as dioxins and PCBs, as well as molecules from foods and other sources. AHR-binding pollutants

cross the placenta and are excreted in breast milk. In animal models and human populations, early life ex-

posure to dioxins and PCBs is associated with persistent defects in the offspring’s immune function. Using

a mouse model, maternal exposure to the AHR ligand and pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin

(TCDD) results in a significantly reduced CD8+ T cell response to influenza A virus (IAV) in the adult off-

spring (F1), compared to the response of infected offspring of control-treated dams. Specifically, there are

significantly fewer cytotoxic T lymphocytes (CTL; CD44hiCD62Llo), virus-specific CD8+ T cells, and

CD8+ T cells that produce interferon gamma (IFNγ). Transcriptomic analyses using sorted CD8+ T cells

from F1 offspring of TCDD and control dams support new evidence that triggering AHR during develop-

ment changes programming of senescence or exhaustion regulatory pathways. Follow up studies show in-

creased expression of proteins associated with hindered T cell responses, such as CTLA-4 and KLRG1 on

CD8+ T cells. We next asked whether the diminished CD8+ T cell response in the F1 generation was ob-

served in the F3 generation. We detected fewer CTL and virus-specific CD8+ T cells in the TCDD F3 line-

age, as well as increased expression of CTLA-4 and KLRG1 compared to control F3 lineage following

IAV infection. These data indicate that F0 maternal exposure to AHR ligands is capable of disrupting im-

mune function not only via direct activation of the AHR in the F1 generation, but also by reprogramming

immune responses in subsequent generations. This has broad implications for understanding how the envi-

ronment of prior generations shapes susceptibility to pathogens and antiviral immunity in later generations.

95.

Poster 12

Immune tolerance against pulmonary F. tularensis infection

Alicia Soucy and Dennis W. Metzger, Ph.D.

Department of Immunology and Microbial Disease, Albany Medical College, Albany, NY

F. tularensis, a CDC Tier 1 agent, is the causative agent of tularemia, and inhalation of only

10 CFU can result in fatal disease. Currently, there is no licensed vaccine against this pathogen; however,

it is known that vaccinating BALB/c mice with a less virulent F. tularensis holartica strain (LVS) can pro-

tect against pulmonary challenge with the highly virulent SchuS4 strain. Vaccination does not protect

C57Bl/6 mice. We hypothesize that, as a result of vaccination, BALB/c mice establish a strong TH17-

lik erecall immune response, which results in tolerance to the SchuS4 strain. To test this, we primed

and boosted mice intranasally with LVS. After three weeks, we challenged LVS-vaccinated mice with

SchuS4, and analyzed bacterial burden. BALB/c mice had a lower bacterial burden in their lungs compared

to C57Bl/6 mice at days 8 and 10. Interestingly, while BALB/c mice survived challenge, they still had a

high bacterial burden in their lungs three weeks after infection. We also analyzed bronchoalveolar lavage

fluid for cytokine secretion; BALB/c mice produced greater levels of IL-17 and IL-22, compared to unpro-

tected C57Bl/6 mice. As expected, we saw decreased survival of vaccinated IL-17R knockout mice follow-

ing SchuS4 infection. In contrast, C57Bl/6 mice expressed greater levels various pro‑inflammatory cyto-

kines, including IFNγ, IL-6, IL-1α, and IL-1β. Histological analysis showed that BALB/c mice had less

lung necrosis at day 8 following SchuS4 challenge compared to C57Bl/6 mice. BALB/c mice also had less

total protein in the bronchoalveolar lavage fluid by day 10 post-infection. These findings suggest that

BALB/c mice mount a strong IL-17 and IL-22 recall response compared to C57Bl/6 mice, which aids in

enhancing survival by limiting tissue damage, but not bacterial clearance. Future studies are being conduct-

ed to determine the precise mechanisms responsible for tolerance in BALB/c mice from pulmonary

SchuS4 infection. (Supported by NIH P01AI056320)

96.

Poster 13.

Autoantigen Availability Determines the Innate Sensing Requirement During Self-Antigen Driven

Germinal Center Responses in Autoimmunity

Stephanie L. Schell, Chetna Soni and Ziaur S.M. Rahman

Penn State College of Medicine, Hershey, PA

Systemic lupus erythematosus (SLE)-associated germinal center (GC) responses are driven by syn-

ergistic BCR, TLR, and IFN signaling. Aberrant selection processes within these GCs lead to the escape

of high-affinity, class-switched autoreactive B cells. The environmental factors that stimulate TLRs and

the regulation of TLR and IFN signaling events during autoreactive GC responses in vivo are incompletely

defined. Previous studies in our lab established that MerTK-deficient (Mer-/-) mice, which lack a critical

receptor expressed by macrophages and DCs that is involved in apoptotic cell clearance and TLR/IFN im-

munoregulatory signaling cascades, exhibit increased GC responses and T cell activation. Here, we used a

kinetic immunization-based approach in Mer-/- mice to drive apoptotic cell accumulation in GCs, allowing

us to evaluate the contribution of TLR-dependent self-ligand sensing and dampened immunoregulation on

GC formation and selection processes, under conditions of high and low autoantigen availability. As antic-

ipated from previous study of spontaneous GC responses in autoimmune-prone mice, TLR7-MyD88 sig-

naling significantly contributes to enhanced GC responses observed in Mer-/- mice, independent of autoan-

tigen load. Unexpectedly, there was also a kinetic dependence on TLR9 in autoantigen-driven GC re-

sponses, suggesting that self-DNA sensing by TLR9 may have a more complex and context-dependent role

in GC responses than initially postulated. Further, the deficiency of MyD88 under conditions of high auto-

antigen availability did not diminish GC response, potentially indicating a novel regulatory function for

MyD88 or its dependent pathways whereby the sensing of ligand present in cytosolic spillover caused by

high autoantigen load is inhibited. Mechanistically, Mer-deficiency promotes enhanced GC responses in

GC B cell-intrinsic and APC-dependent manners, with process/cell-type dependent requirements for TLR

signaling. As a result of altered immune activation, the loss of Mer compounded aberrant GC selection

and downstream kidney pathology in autoimmune-prone B6.Sle1b mice deficient for Mer (B6.Sle1b.Mer-/-

). Altogether, these results suggest that TLR signaling plays a complex role in self-antigen driven GC re-

sponses, whereby both kinetics and autoantigen load determine the requirement for different self-ligands

and the activation of their relevant receptors during response. Mer-deficiency also promotes autoimmunity

by synergizing with genetic susceptibility loci to dysregulate selection processes within GCs.

97.

Poster 14.

The myeloid transcription factor IRF8 is integral for macrophage-mediated control of tumor

metastasis

Danielle Y.F. Twum1, Michael Nemeth1, Austin Miller2 & Scott I. Abrams1 1Department of Immunology; 2Department of Biostatistics & Bioinformatics,

Roswell Park Cancer Institute, Elm and Carlton Streets, Buffalo, NY 14263

The ‘M1’ (immune stimulatory) or ‘M2’ (immune suppressive) macrophage response in neoplasia

is governed by transcriptional pathways likely regulated by stromal- or tumor-derived factors bathing the

tumor microenvironment. In neoplasia, notably breast cancer, a high infiltration of macrophages is associ-

ated with reduced overall survival as well as progression to metastasis, which is thought to involve an M1

to M2 shift that favors tumor growth. Although several transcriptional regulators that drive the M2 pheno-

type in the tumor microenvironment have been identified, transcription factors that drive or enforce the M1

phenotype are not as well defined. IRF8 (Interferon Regulatory Factor-8) is a transcriptional regulator of

multiple aspects of myeloid biology, including differentiation and functional status. In the case of func-

tion, it is well-known that IRF8 is critical for the expression of genes such as iNOS and IL-12p40, hall-

mark features of an M1 phenotype. However, the role of IRF8 in macrophage-tumor biology is unknown.

Based on this rationale, we hypothesized that IRF8 is important in governing the ‘M1 vs M2’ macrophage

response in malignancy, namely mammary cancer whereby tumor-associated macrophages have been

linked to disease progression. To test this hypothesis, we utilized an IRF8 knockout model (IRF8fl/fl)

whereby IRF8 is specifically deleted in the macrophage compartment (LysM-Cre) and compared the rate of

tumor growth, as well as propensity for lung metastasis. Importantly, using an implantable mouse mam-

mary tumor model, we observed an increased propensity of lung metastasis in IRF8-/- mice compared to the

appropriate counterpart controls. Altogether, these data indicate that IRF8 expression in macrophages can

impact tumor progression to metastasis, likely through altering the tumor-suppressing (M1) vs. tumor-

promoting (M2) phenotypes. Thus, IRF8 may represent a potentially novel therapeutic target to modulate

the macrophage phenotype in neoplastic settings, such as mammary cancer, whereby this myeloid response

is a critical determinant of outcome.

98.

Poster 15.

Differentiation of iPSC-derived T cells specific to HBV

Praneet Kaur Sandhu, Mohammad Haque, Kristin Fino, Jianxun (Jim) Song

Pennsylvania State University College of Medicine, Hershey PA

Hepatitis B virus (HBV) is a double stranded DNA virus that infects hepatocytes to cause acute or

chronic infection in humans. Approximately 240 million people suffer from chronic HBV infection, which

involves persistence of virus for more than 6 months in infected individuals and can result in development

of hepatocellular carcinoma (HCC) and liver cirrhosis. Current treatment of HBV involves administration

of nucleoside/nucleotide inhibitors and cytokines but these have various side effects and prone to escape

mutations by HBV. Cellular immunotherapy using T cells primed to target disease-specific antigen could

prove to be an effective strategy to target hepatocytes that are chronically infected with HBV. Induced plu-

ripotent stem cells (iPSCs) can serve as a source of T cells as they have the ability to differentiate into any

cell type within the body. The objective of this project is to develop HBV-specific T cells from iPSCs in

vitro, characterize the underlying in vitro T cell differentiation mechanisms and utilize these iPSC-derived,

HBV-specific T cells for therapy in a HBV mouse model.

HBV-specific TCR was cloned into dsRed containing retroviral vector and successfully used for

retroviral transduction into murine iPSCs to establish a stable HBV-TCR expressing iPSC cell line. HBV-

TCR iPSCs were co-cultured with OP9-DL1-DL4 cells, a stromal cell line expressing delta ligand 1 (DL1)

and delta ligand 4 (DL4) for Notch signaling, to induce T cell differentiation.

In vitro differentiation of iPSCs into T cells can be utilized as effective method for generating T

cells for immunotherapy. Moreover, the development of iPSC-derived, HBV-specific T cells would pave

way for a therapeutic intervention for HBV.

99.

Poster 16.

Smoke Exposure Impairs Immunity To Lung Infection

Tariq A. Bhat1, Amit A. Lugade1, Suresh G. Kalathil1, Paul N. Bogner2,Thomas H. Thatcher3, Richard P.

Phipps3, Patricia J. Sime3,4, and Yasmin M.Thanavala1 1Department of Immunology and 2Department of Pathology, Roswell Park Cancer Institute, Buffalo, NY,

USA, 3Department of Medicine and 4Department of Environmental Medicine, University of Rochester,

Rochester, NY 14620

Tobacco smoke is recognized to have a detrimental impact on public health. Despite aggressive ef-

forts to prevent smoking, approximately a billion individuals continue to smoke worldwide. Cigarette

smoke (CS) exposure is a key initiator of chronic pulmonary inflammation and a major risk factor in the

development of respiratory disorders including chronic obstructive pulmonary disease (COPD). Strikingly,

in developed nations secondhand smoke (SHS) exposure is the main risk factor for nonsmokers to develop

emphysema and COPD. While exposure to SHS is common, the magnitude of this problem is not fully ap-

preciated.

To understand the connections among tobacco smoke exposure, chronic pulmonary infection,

chronic inflammation, and changes in immunity, we developed a mouse model. A major goal of our studies

is to understand the increased susceptibility to lung infection following exposure to either mainstream CS

or SHS, and to evaluate therapies that can restore normal immune function.

Using this model, we have shown that CS exposure has a profound immunosuppressive effect on

the generation of innate and adaptive immune responses to chronic infection with nontypeable Haemophi-

lus influenzae (NTHI). Our studies provide an understanding of the mechanism by which prior and ongo-

ing CS exposure predisposes COPD patients to recurrent infections that lead to exacerbations and contrib-

ute to mortality.

Recognizing the detrimental effects of SHS on human health we also utilized our mouse model to

interrogate its impact on pulmonary inflammation, immunity to pulmonary infection and to vaccination.

We observed that prior chronic SHS exposure also induces pulmonary inflammation and significantly

dampens the development of innate and adaptive immunity against chronic infection. We further investi-

gated whether judicious therapeutic intervention with an anti-inflammatory and pro-resolving lipid media-

tor may help reverse the inflammatory damage caused by prior smoke exposure and showed that it is possi-

ble to mitigate the effects caused by SHS.

It is well accepted that lung inflammation caused by cigarette smoke exposure persists long-term.

We therefore examined if cessation of exposure to SHS alone or in combination with treatment can reduce

inflammatory changes in the lung and augment immune function.

By taking a multi-pronged approach we have clearly shown that prior exposure to either CS or SHS

worsens respiratory infection-mediated inflammatory milieu and leads to the generation of defective innate

and adaptive immune responses. Furthermore, our results demonstrate the dramatic beneficial effects of

cessation and therapy in limiting pulmonary injury and augmenting immune responses to infection and

vaccination. Our results have considerable clinical relevance.

These studies were supported by a FAMRI Clinical Innovators Award.

100.

Poster 17.

Early IL-10 signals favor regulatory B cell over memory B cell development during cognate iNKT cell help

Jennifer Yates1,4, Emilie Vomhof-Dekrey1, Paula Lanthier1, Katja Mohrs1, Thomas Hägglöf 2, Natacha Veerapen3, Gurdyal Besra3, Mikael Karlsson2, and Elizabeth Leadbetter1,5

1Trudeau Institute, Saranac Lake, NY 12983; 2Karolinska Institutet, Stockholm, Sweden; 3School of Bio-sciences University of Birmingham, Birmingham, UK; 4Wadsworth Center, NYSDOH, Albany, NY; 5Uni-versity of Texas Health Science Center at San Antonio, San Antonio, TX

Effective generation of humoral B cell memory is dependent upon help from CD4+ T cells. We and

others have found that invariant natural killer T (iNKT) cells can provide both cognate and non-cognate

helper signals to enhance B cell responses. While both cognate and non-cognate iNKT cell help induce

class-switched, antigen-specific humoral immune responses - only non-cognate iNKT cell help drives the

formation of humoral memory. Rather, cognate iNKT cell help drives an early, un-sustained expansion of

germinal center B cells and antigen-specific antibody production. Therefore, we posit that cognate help

provided to B cells by iNKT cells is fundamentally different from the help provided by conventional CD4+

T cells. We now find that glycolipid immunization drives considerable IL-10 transcription by many differ-

ent spleen cell populations including dendritic cells, plasmablasts, iNKT cells, and B regulatory cells. Cog-

nate iNKT cell help expands antigen-specific IL-10 producing B regulatory cells upon primary immuniza-

tion, and IL-10 producing iNKT10 cells following secondary antigen challenge. Early, but not late, block-

ade of the IL-10 receptor resulted in a significant, and sustained increase in antigen-specific antibody titers

during cognate iNKT cell help, but had no effect when traditional CD4+ T cell help was present. We con-

clude that the early composite cytokine environment is critical for dictating the long-term course of the B

cell response. Based on these data, we suggest that B cell antigens which recruit only cognate help from

iNKT cells experience a regulatory rather than inflammatory environment.

101.

Poster 18.

Inflammation, androgens and macrophages in the prostate: Are we missing the link?

Camila Rosat Consiglio1 and Sandra Gollnick1,2

Departments of Immunology1 and Cell Stress Biology2, Roswell Park Cancer Institute,

Buffalo, NY

Prostate cancer (PCa) has the highest malignancy incidence rates in men and is the second leading

cause of male cancer mortality. While the underlying causes of PCa still remain largely unknown, it is

known that androgens, inflammatory mediators and inflammatory cells, including macrophages, are im-

portant players in prostate tumorigenesis. Interestingly, it has been shown that androgen receptor (AR) ex-

pression by macrophages enhances prostatic intraepithelial neoplasia formation in PTEN+/- mice, suggest-

ing that macrophage promotion of tumorigenesis is linked to macrophage AR signaling. Although it is

known that androgens influence immunity, the consequences of AR signaling in macrophages are largely

unknown. Furthermore, the activity and regulation of macrophage AR in the context of prostate tumorigen-

esis is still unclear. To address this gap in the field, the current study aims on analyzing the role of AR in

macrophage homeostasis and in prostate tumorigenesis. Initial results have shown that macrophages ex-

press higher levels of AR protein when compared to monocytes. Polarization of macrophages with either

IFN-γ (gamma) or LPS led to an increase in AR levels. Interestingly, macrophages in G2/M phase of the

cell cycle express higher levels of AR than macrophages in G1 phase, indicating that not only is AR in-

volved in macrophage differentiation and M1 polarization, but also in macrophage cell cycle. Imagestream

analysis of macrophages indicated that AR has strong nuclear localization. In addition, we have confirmed

AR transcriptional activity in bone marrow-derived macrophages using a luciferase assay. Since our results

show that AR is involved in macrophage homeostasis, we next investigated its role in tumor-bearing ani-

mals. We observed an increase in AR levels in tumor-associated macrophages (TAMs) of TRAMP C2

prostate tumors when compared to macrophages from other tissues. This finding was also observed in a

head and neck cancer model (MTERL). This evidence points to a role of the tumor microenvironment in

modulating macrophage AR levels and potentially its activity. Since it is known that macrophage AR facil-

itates prostate tumorigenesis and that macrophage AR levels are higher in the tumor microenvironment, it

is possible that early stages of tumorigenesis could induce macrophage AR signaling, leading to a pro-

tumorigenic phenotype in these cells. Future aims for this project will focus on elucidating macrophage AR

transcriptional targets in homeostasis and in cancer, as well as characterizing macrophage AR function dur-

ing prostate tumorigenesis.

102.

Poster 19.

Selenoproteins regulate B cell functions by targeting B cell receptor-mediated antigen

presentation pathway

Bhuvana Katkere1, Rachel L. Markley1, David R. Williamson1, Ashley E. Shay1, Bradley A. Carlson2.

Kumble S. Prabhu1 and Girish S. Kirimanjeswara1

1The Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park,

PA,16802

2Molecular Biology of Selenium Section, Mouse Cancer Genetics Program, National Cancer Institute, Na-

tional Institutes of Health, Bethesda, MD, 20892

The essential micronutrient selenium (Se) has been shown to influence the immune functions in

both animals and humans. A vast majority of biological effects of Se is mediated by selenoproteins, a class

of proteins that contain 21st amino acid selenocysteine. A few recent studies have revealed that selenopro-

teins modulate the functions of T cells and macrophages. However, few studies have investigated the di-

rect effect of Se or selenoproteins on B cell functions. We therefore sought to establish the role of Se and

selenoproteins in B cell activation and functions, which are dependent on B cell receptor (BCR) signaling.

Supplementation of B cell line A20uWT with Se as sodium selenite at 200 nmol concentration resulted in

increased rate of BCR endocytosis, calcium signaling, antigen trafficking to MIIC compartment, antigen

degradation and antigen presentation compared to the cells maintained on deficient (0 nmol) or adequate

levels (50 nmol) of sodium selenite. Similar results were observed when B cells were supplemented with

other selenium containing compounds that can be metabolized and incorporated as selenocysteine. Howev-

er, supplementation with selenium compounds that cannot be metabolized and incorporated as selenocyste-

ine did not alter BCR-mediated signaling suggesting that the BCR-mediated signaling is regulated by sele-

noproteins. These data were reproducible in primary B cells isolated from the spleens of mice fed with 0.4

ppm sodium selenite (supplemented) compared to the mice fed with 0.01 ppm (deficient) or 0.08 ppm

(adequate) sodium selenite. This phenomenon was specific to BCR as other cell surface receptors such as

transferrin receptor internalization was not affected by the Se status of cells. Further studies revealed that

Se supplemented B cells had significantly lower levels of reactive oxygen species (ROS) compared to defi-

cient and adequate cells. Pharmacological reduction of ROS in Se deficient and adequate B cells resulted in

increased rate of BCR endocytosis. In contrast, pharmacological induction of ROS in Se supplemented

cells significantly decreased the BCR endocytosis. Together, these data suggest that one or more seleno-

proteins regulate the redox status of B cells that in turn has effect on B cell antigen processing and presen-

tation. Our preliminary studies suggest that Glutathione peroxidase (Gpx) family of selenoproteins such as

Gpx1 may be directly involved in the regulation of BCR-mediated signaling. These data together suggest

that Se, via selenoproteins, regulate the B cell receptor-antigen trafficking and presentation to impact the

secondary immune responses providing novel insights into molecular mechanisms of immune regulation.

103.

Poster 20.

SLAMf6 modulates the NKT cell death threshold

Victoria L. DeVault1, Oliver Dienz1, Graham W.J. Lilley1, Patrick Benoit1,

Pamela L. Schwartzberg2, Jonathan E. Boyson1 1Department of Surgery, University of Vermont College of Medicine, Burlington, VT 2National Human Genome Research Institute (NHGRI), National Institutes of Health,

Bethesda, MD

Signaling lymphocyte activation marker family member 6 (SLAMf6) is a cell surface signaling

receptor that plays a critical role in NKT cell development. Surprisingly, the exact mechanisms through

which SLAMf6 regulates NKT cell development and function remain unclear. To investigate the function

of SLAMf6 on peripheral NKT cells, we challenged C57BL/6 (B6) or B6.Slamf6-/- mice with the NKT

cell agonist, aGalCer. While we detected no difference between B6 and B6.Slamf6-/- mice in NKT cell

IFN-g, IL-4, and TNFa production, we did find a significant difference in NKT cell numbers in the pe-

riphery. Three days after challenge, we observed a 50-fold increase in NKT cells in B6 mice over vehi-

cle-treated controls. In contrast, we found a 220-fold increase in NKT cells in B6.Slamf6-/- mice versus

controls. A comparison between B6 and B6.Slamf6-/- mice of in vivo BrdU uptake by NKT cells re-

vealed no significant differences in proliferation. We then compared NKT cell apoptosis using Annexin

V staining and live/dead discriminator 3 h after aGalCer administration. This analysis revealed a signifi-

cantly lower percentage of apoptotic NKT cells in B6.Slamf6-/- versus B6 mice, suggesting that SLAM-

f6 expression on NKT cells was associated with activation-induced cell death (AICD). Consistent with

this observation, we found significantly diminished expansion of sorted B6 NKT cells when they were

stimulated by SLAMf6+ antigen-presenting cells in an in vitro cell culture system. We conclude that in

the presence of a strong, high-affinity agonist, SLAMf6 contributes to significantly increased NKT cell

AICD and concomitant diminished NKT cell expansion. Interestingly, when we conducted similar com-

parisons in naïve mice under homeostatic conditions, we observed an increased percentage of apoptotic

NKT cells in B6.Slamf6-/- mice versus their B6 counterparts, which was associated with lower NKT cell

numbers in B6.Slamf6-/- mice. Taken together, these data support a model in which SLAMf6 regulates

NKT cell activation and death thresholds depending on the strength of activation. These data also suggest

that SLAMf6 blockade could be a useful tool to manipulate NKT cell populations in vivo.

104.

Poster 21.

Cell-Intrinsic Glycogen Metabolism Supports Early Activation and Maintains Metabolite

Homeostasis in Dendritic Cells

Phyu Thwe1, Angelo D’ Alessandro2, Princess Rodriguez1, and Eyal Amiel1,3

1. University of Vermont, Cellular Molecular and Biomedical Science Program

2. University of Colorado Denver

3. University of Vermont, College of Nursing and Health Sciences.

As professional antigen presenting cells of the immune system, dendritic cells (DCs) serve as a

bridge between innate and adaptive immune responses. Activation of DCs by a stimulus through toll-like

receptors (TLRs) is coupled with an increase in metabolic demand that is fulfilled by a TLR-driven burst in

glycolytic reprogramming. Up-regulation of glycolysis in activated DCs provides metabolites required for

DC effector function, and inhibition of glycolysis impairs the post-activation survival and effector function

of these cells.

TLR-driven glycolysis is thought to be sustained primarily by increased glucose uptake via the in-

ducible glucose transporter 1 (GLUT1). However, whether glucose is sourced from extracellular or intra-

cellular stores during early glycolytic reprogramming in DCs is still not well-defined. We propose that cell

-intrinsic glycogen metabolism in DCs supports early glycolytic burst that is essential for TLR-driven acti-

vation. The functional importance of glycogen metabolism in the context of DC effector responses has not

been previously described. Our data indicate that glycogen metabolism supports the activation of DCs, par-

ticularly during early activation before the up-regulation of GLUT1 expression. We show that DCs express

the enzymes essential for glycogen metabolism and that glycogen metabolism is regulated upon TLR stim-

ulation. Inhibition of glycogen utilization in DCs impairs the expression of costimulatory molecules CD40

and CD86 in these cells. The ability of DCs to uptake antigens and stimulate T cells is also compromised

upon disruption of glycogen metabolism. In addition, our metabolomics data indicate that glycogen metab-

olism in DCs generates both glycolytic and TCA cycle intermediates and that glycogen-derived carbons

may support metabolic pathways distinct from free glucose catabolism. These data demonstrate that the

glycogen metabolism plays a significant role in metabolic homeostasis in DCs and define a novel metabol-

ic regulatory pathway that supports DC immune function.

105.

Poster 22.

IgM memory B cells reconstitute multiple B cell lineages and provide protection

Kevin Kenderes, Amber Papillion, Gary Winslow


IgM memory B cells are now recognized as an important component of immunological memory.

They have been proposed to be a reservoir of broadly-reactive B cells that differentiate, in germinal cen-

ters, into high affinity class-switched B cells following antigen encounter. We provide evidence that a

highly-purified IgM memory B cell population can follow multiple pathways of differentiation after chal-

lenge infection, and demonstrate that the antibodies produced by these cells can provide protective immun-

ity. Our experimental model uses Ehrlichia muris, an intracellular tick-borne bacterium that generates a

robust CD11c+ T-bet+ IgM memory B cell population in C57BL/6 mice. Due to the presence of pre-

existing antibodies, investigation of the secondary IgM memory B cell response to ehrlichial infection had

not been possible. We are able to monitor EYFP-labeled IgM memory B cells after transfer of splenocytes

into naïve mice and observed differentiation EYFP-labeled cells into all effector and memory B cell line-

ages following secondary infection. This was accompanied by a 4-fold increase in IgM production, relative

to infected mice that did not receive memory cells. However, a small population of EYFP-labeled switched

memory cells was also found in the donor spleen cells. Therefore, to determined if IgM memory cells were

solely responsible for the reconstitution of the memory and effector B cell lineages we monitored highly

purified EYFP-labeled spleen IgM memory B cells following their transfer into naïve recipient mice. After

challenge infection, some donor memory B cells differentiated into IgM-producing plasmablasts and CD19

-negative plasma cells. Other donor B cells entered germinal centers, down-regulated CD11c, underwent

class switching, and generated switched memory B cells. Yet other donor cells were maintained as IgM

memory cells. Donor IgM memory B cells also protected the recipient mice from the fatal erhlichial infec-

tion, Ixodes ovatus ehrlichia (IOE), demonstrating the importance of IgM memory cells for protective sec-

ondary responses. Thus, during secondary responses, IgM memory cells can differentiate into IgM-

producing plasmablasts, switched germinal center cells, or switched memory cells, or undergo self-

renewal. These data reveal that IgM memory B cells are capable of generating protective secondary re-

sponses that can replenish many, if not all, effector and memory B cell lineages, thereby contributing to

long-term immunity to pathogens.

106.

Poster 23.

CD28 Induces Metabolic Fitness in LLPCs through NFkB-Mediated Irf4 Expression and

ROS-Dependent Survival.

Adam Utley, James Cooper, Louise Carlson, Peng Peng, Amin Mahpour, and Kelvin Lee.

Roswell Park Cancer Institute, Buffalo NY

Sustained humoral immunity is dependent upon the continual production of antigen-specific anti-

bodies by plasma cells. Upon activation, B cells differentiate into short-lived plasma cells (SLPCs) that

traffic to secondary lymphoid organs such as the spleen where they live for days to weeks then die by

apoptosis. In a second non-mutually exclusive model, B cells can differentiate into long-lived plasma cells

(LLPCs) that home to specialized survival niches in the bone marrow and live indefinitely. Much work has

gone into describing the competitive BM survival niche; however, the cellular and molecular interactions

which govern this survival program are incompletely understood.

We have published that CD28, the canonical T-cell costimulatory molecule, is required for LLPC

survival. In T cells CD28 is known to induce glycolysis at the expense of mitochondrial respiration. To our

great surprise, CD28 increased mitochondrial respiration in LLPCs whilst not affecting glycolysis directly.

CD28 increased Glut1 expression and subsequent LLPC glucose uptake, as well as the glycolytic capacity.

This suggests that CD28 regulates the ability of LLPCs to successfully compete for nutrients in the BM

niche for long term survival and antibody production.

Mechanistically, CD28 induces NFkB-dependent IRf4 upregulation, known to regulate LLPC sur-

vival. Furthermore, inhibition of NFkB abrogates the CD28-induced increases in glucose uptake and mito-

chondrial mass. We recently published that the Grb2/Vav binding domain on the CD28 cytoplasmic tail is

required for LLPC survival. In mice wherein this domain is mutated (AYAA mutants), LLPCs have de-

creased Irf4 expression, glucose uptake and mitochondrial mass. This facilitates a model wherein CD28

induces NFkB dependent Irf4 through Grb2/Vav for metabolic fitness. We also describe an NFkB superen-

hancer element upstream of the Irf4, suggesting that CD28 may govern direct Irf4 promoter activity as well

as DNA folding. Irf4 knock down decreased PC mitochondrial mass, demonstrating that Irf4 may directly

regulate LLPC metabolic fitness.

One byproduct of mitochondrial respiration is the production of reactive oxygen species (ROS).

CD28 increased ROS specifically in LLPCs. Paradoxically, ROS inhibition prevented CD28-mediated sur-

vival. Taken together these results suggest a model wherein CD28 through its Grb2/Vav binding domain

induces NFkB dependent upregulation of Irf4 directly through the promoter region, and augments further

Irf4 production through a previously undescribed NFkB superenhancer element. Irf4 then goes on to in-

crease mitochondrial respiration-dependent ROS for CD28-mediated LLPC survival and metabolic fitness.

Targeting CD28 with FDA-approved drugs may augment vaccine design as well as alleviate antibody me-

diated auto-immunity.

107.

Poster 24.

Tumor-induced myeloid-derived suppressor cells act via remote control to inhibit L-selectin-

dependent adaptive immunity in lymph nodes

Amy W. Ku1, Jason B. Muhitch1, Colin A. Powers1, Michael Diehl1, Anand P. Sharda1,

Kieran O'Loughlin1, Hans Minderman1, Joseph J. Skitzki1, Suzanne Ostrand-Rosenberg2, Scott I. Abrams1,

Sharon S. Evans1

1Roswell Park Cancer Institute, Buffalo, NY 14263, USA; 2University of Maryland Baltimore County,

Baltimore, MD 21250, USA

Myeloid derived suppressor cells (MDSC) are potent immunomodulatory cells that play an exten-

sive role in cancer progression and immune evasion. These immature myeloid cells are known to accumu-

late within the spleen and tumor, and their ability to suppress effector T cell functions within these tissues

is well described. In contrast, the impact of MDSC on naive T cells within lymph nodes (LN) has been

largely overlooked as MDSC are rare within these critical sites of immune priming. Previous reports have

shown that peripheral MDSC from tumor-bearing mice downregulate the LN homing receptor L-selectin

on naive CD4 and CD8 T lymphocytes, but the molecular mechanisms and target cell-specificity has been

unclear. Furthermore, the biological relevance of moderate fluctuations of L-selectin is questionable as the

high density of L-selectin molecules normally present on T cells could theoretically buffer against the ef-

fects of such loss during trafficking. Using stringent murine mammary tumor models of high and low

MDSC burden (4T1 and AT-3, respectively), we demonstrate that MDSC downregulate L-selectin on na-

ive T and B cells post-transcriptionally via a contact-dependent mechanism. MDSC-driven loss of L-

selectin occurs within 24 hours both in vitro and in vivo, and does not appear to be species-restricted as L-

selectin on human lymphocytes can also be targeted by MDSC. By employing real-time intravital micros-

copy and immunofluorescence histology to visualize and assess naive CD8 T cell trafficking within vascu-

lar gateways for lymphocyte trafficking known as high endothelial venules (HEV), we found that even

moderate losses of L-selectin mediated by MDSC causes a profound reduction in the quality of lymphocyte

-HEV interactions. Ultimately, this results in significantly fewer T cells trafficking and infiltrating into the

LN parenchyma. In an in vivo vaccination model, MDSC-mediated loss of L-selectin on naive CD8 T cell

and subsequent reduction in lymphocyte trafficking severely diminishes antigen-driven T cell expansion

within draining LN. These data reveal a novel mechanism by which tumor-induced MDSC localized out-

side of the LN shape the magnitude of T cell responses within the intranodal compartment, which has un-

anticipated implications for systemic immunity in cancer. Supported by the NIH (CA79765, AI082039,

T32CA085183, 5T32CA108456, 5P30 CA016056), the Breast Cancer Coalition of Rochester, the Mark

Diamond Research Fund and the Jennifer Linscott Tietgen Family Foundation.

108.

Poster 25.

Small Molecule Inhibitors of the PI3K/SHIP-1/Akt Pathway Sensitize Human

Cytomegalovirus-infected Monocytes to Apoptosis

Olesea Cojohari, Megan A. Peppenelli, and Gary C. Chan

SUNY Upstate Medical University, Syracuse, NY

Blood monocytes are responsible for systemic human cytomegalovirus (HCMV) dissemination,

which can cause multiorgan failure in immunodeficient hosts, such as transplant recipients and patients un-

dergoing chemotherapy. Current anti-HCMV drugs target viral replication, rendering them ineffective in

monocytes where the virus does not replicate; thus, novel therapeutic targets are needed to eliminate

HCMV-infected monocytes and prevent viral spread. Monocytes are naturally short-lived cells. We previ-

ously showed HCMV manipulates monocytes to survive past their 48-hour life-span. Activation of Akt

(antiapoptotic protein) is critical for M-CSF (myeloid growth factor) induced monocyte survival. Howev-

er, Akt’s role in HCMV-infected monocytes is unknown. We now show that treatment with the Akt inhibi-

tor, MK-2206, resulted in infected-monocytes apoptosis, suggesting Akt is central to HCMV induction of

monocyte viability. Interestingly, Akt activation was enhanced and kinetically different with HCMV than

M-CSF, implying the virus uses a unique mechanism to regulate Akt. Using PI3K isoform inhibitors, we

show HCMV induces a switch from p110δ, the main PI3K isoform positively regulating Akt in leukocytes,

to p110β, to mediate Akt-dependent survival. We next examined SHIP-1, a negative Akt regulator, shown

to also mediate positive (non-canonical) Akt regulation in cancer cells. Treatment with a novel SHIP-1

inhibitor (3AC) annulled HCMV-induced Akt signaling and triggered apoptosis, suggesting HCMV ex-

ploits a two-pronged approach, through PI3K-p110β and SHIP-1, to stimulate infected-monocyte survival.

Finally, HCMV entry elicited phosphorylation-mediated inactivation of PTEN (another negative Akt regu-

lator), ensuring maximum Akt activity. Overall, our data indicate that targeting Akt, p110β, or SHIP-1

with small molecule inhibitors could be effective in eliminating HCMV-infected monocytes and prevent-

ing viral spread and pathogenesis.

109.

Poster 26.

Tumor-Derived Indoleamine 2,3- Dioxygenase Regulates Density of Tumor Infiltrating CD8+ T cells

and Myeloid Derived Suppressor Cells in a Murine Model of Ovarian Cancer

Adaobi Amobi1,3, and Kunle Odunsi1,2,3

Departments of Immunology1, Gynecologic Oncology2 and Center for Immunotherapy3

Roswell Park Cancer Institute, Buffalo, NY

Amino-acid withdrawal is an important molecular mechanism regulating anti-tumor immune re-

sponses. The catabolism of the essential amino-acid tryptophan (TRP) by indoleamine 2,3-dioxygenase

(IDO1) is a central pathway that contributes to the immunosuppressive microenvironment in many types of

cancer. IDO1 enzymatic activity results in depletion of TRP and the generation of immunosuppressive me-

tabolites, such as kynurenine. Our lab has previously shown that IDO1 expression in human ovarian tumor

correlates with poor prognosis and poor tumor infiltration by CD8+ T cells. Moreover, our lab demonstrat-

ed that increased infiltration of CD8+ T cells into the tumor is associated with improved survival. Thus,

IDO1 inhibition represents an attractive target for cancer immunotherapy.

To establish the mechanism by which IDO1 inhibition augments immune responses in a murine

model of metastatic ovarian cancer, we utilized a murine ovarian surface epithelial cancer cell line, IE9-

mp1. We generated a stable IDO1-overexpressing cell line (IE9mp1-mIDO1) by transfecting murine IDO

cDNA into parental IE9-mp1 cells and confirmed functional IDO1 enzyme activity. C57BL/6 mice were

challenged intraperitoneally with either parental IE9mp1-Empty Vector (IE9mp1-EV) or IE9mp1-mIDO1

tumor cells. Syngeneic immunocompetent mice inoculated with IE9mp1-mIDO1 cells displayed earlier

onset of tumor burden and decreased overall survival compared with IE9mp1-EV challenged mice.

To delineate the role of host- and tumor-derived IDO1 on immune cell infiltration to the tumor site,

we utilized the IDO1 genetic knockout (IDO1KO) mouse model. IDO1KO and C57BL/6 mice were chal-

lenged intraperitoneally with either the IE9mp1-EV or IE9mp1-mIDO1 tumor cells. C57BL/6 and

IDO1KO mice challenged with IE9mp1-mIDO1 demonstrate reduced CD8+ T cell infiltration within the

tumor. Interestingly, IDO1KO mice challenged with IE9mp1-EV tumor cells demonstrate increased tumor

infiltration by CD8+ T cells compared to C57BL/6 mice. Moreover, tumor-derived IDO1 mediates in-

creased frequency in the CD11b+Gr1+ myeloid derived suppressor cell (MDSC) population in ascites fluid

early-on along tumor burden in C57BL/6 and IDO1KO tumor-bearing animals.

From these data, we conclude that regulation of IDO1 will promote anti-tumor immune responses,

by permitting increased frequency of effector T cells in tumor tissues. Moreover, tumor-derived IDO1 inhi-

bition may decrease the frequency of CD11b+Gr1+ MDSCs in ascites fluid. Future studies are ongoing to

further delineate the specific contribution of IDO1 by tumor cells, host cells, or both mutually to the regu-

lation of immunosuppressive MDSCs in ovarian cancer. Experiments are ongoing to characterize the

mechanism by which IDO1 inhibition may augment vaccine-induced immune responses in a murine model

of ovarian cancer.

Supported by: NCI SPORE P50 CA159981

110.

Poster 27.

Negative impact of myeloid-derived suppressor cells on CD8 effector T cell trafficking within the tu-

mor microenvironment

Amy Ku1, Michelle Appenheimer1, Jason Muhitch2, Scott I. Abrams1, and Sharon S. Evans1

Departments of Immunology1 and Urology2, Roswell Park Cancer Institute, Buffalo, NY

The success of T cell-based immunotherapy and, unexpectedly, thermal therapy, standard chemo-

therapy and radiation hinges on cytotoxic T cells gaining access to tumor targets. These observations have

prompted interest in strategies to improve T cell trafficking to tumors although the mechanisms that posi-

tively or negatively regulate extravasation at tumor vascular checkpoints are poorly understood. Here, we

report that the ability of tumor vessels to respond to IL-6-dependent preconditioning regimens that boost

CD8 effector T cell homing is temporally and inversely related to the accumulation of myeloid-derived

suppressor cells (MDSC) within the tumor microenvironment. Using real-time intravital imaging and im-

munofluorescence histology, IL-6 therapies were shown to convert vessels from T cell-low to -high recruit-

ment sites in murine tumors with minimal MDSC infiltration (i.e., CT26 colorectal, B16 melanoma, EMT6

mammary tumors). This conversion requires induction of the ICAM-1 trafficking molecule on tumor ves-

sels. Conversely, mammary (4T1, AT-3 and PyMT-MMTV) and pancreatic (Pan02) tumors with high

MDSC burdens were refractory to IL-6 therapies, but became responsive after acute MDSC depletion. To

further investigate contributions of MDSC to poor trafficking, IL-6-responsive tumors were admixed with

syngeneic CD11b+Gr-1+ MDSC isolated from spleens of tumor-bearing mice at a ratio of 2:1, thus mimick-

ing the high MDSC burden detected in IL-6-refractive tumors. Sustained intratumoral elevation of MDSC

in admixed tumors resulted in failure to support increased T cell trafficking in response to IL-6–dependent

therapies. Complementary in vitro studies revealed that MDSC directly influence and downregulate traf-

ficking molecule expression on endothelial cells. Taken together, these findings identify a novel role of

MDSC in subverting antitumor immunity by limiting T cell trafficking at tumor vascular loci. Supported

by NIH (R01CA79765, R01AI082039, 2T32CA085183), the Breast Cancer Coalition of Rochester, the

Mark Diamond Research Fund and the Jennifer Linscott Tietgen Family Foundation.

111.

Poster 28.

Triggering the aryl hydrocarbon receptor during development durably programs

CD4+ T cell responses

Catherine G. Burke1, Lisbeth A. Boule2, Jason R. Myers3, and B. Paige Lawrence1,2 1Department of 1Microbiology and Immunology, 2Department of Environmental Medicine, and

3Genomics Research Center, University of Rochester Medical Center, Rochester, NY

Emerging evidence shows that early life environmental exposures lead to lasting changes in im-

mune function, including increased incidence and severity of respiratory infections. Among the environ-

mental factors associated with altered immune function are ligands of the aryl hydrocarbon receptor

(AHR). Yet, how inappropriate triggering of AHR during development changes immune responses later in

life remains poorly understood. Recently, we showed that developmental activation of the AHR using the

prototypical ligand 2,3,7,8-tetrachlorodibenzo-p- dioxin (TCDD) results in an impaired response to influ-

enza A virus infection (IAV) in adult offspring. Changes include significantly fewer conventional CD4+

T cell effector subsets in infected offspring of TCDD treated dams, compared to infected offspring of con-

trol-treated dams. Adoptive transfer experiments show that this poorer response reflects developmentally

induced events intrinsic to the CD4+ T cell lineage. The activation and differentiation of naïve CD4+ T

cells into functionally distinct conventional subsets involves the integration of multiple signaling path-

ways, providing many different genes and cellular processes that could be affected by AHR activation dur-

ing development. To address this, we used an unbiased transcriptomic approach to identify differentially

expressed genes (DEG) in naïve and activated CD4+ T cells from infected adult offspring of dams treated

with TCDD or control. Specifically, we sorted CD44lo (naïve) and CD44hi (activated) CD4+ T cells, and

used next generation RNA sequencing (RNA-seq). After alignment and mapping to the genome, we used

DESeq2 to identify DEGs, and Ingenuity Pathways Analysis (IPA) to identify cellular pathways changed

by developmental AHR activation. Developmental exposure altered several signaling pathways critical for

CD4+ T cell differentiation, cell cycle regulation checkpoints, and metabolism. Activated CD4+ T cells

had more DEGs compared to unactivated cells. In addition, many DEGs and pathways were unique to

CD44hi cells, indicating that some developmentally programmed changes in gene expression are cryptic

(i.e. only revealed after CD4+ T cells are activated). These results indicate that early life AHR activation

influences several pathways critical for CD4+ T cells to mount appropriate responses to immune challeng-

es later in life. As humans are regularly exposed to AHR ligands, this has broad spanning implications for

human health, and gives insight into putative mechanisms by which early life environmental exposures

result in long lasting deficits in immune function.

112.

Poster 29.

Tumor Hypoxia alters Anti-tumor Immune Responses: Implications for Radiotherapy

Aditi Murthy1, Scott A. Gerber1, Cameron J. Koch2, and Edith M. Lord1

1University of Rochester Medical Center, Rochester, NY 2 University of Pennsylvania, Perelman School of Medicine, Philadelphia, PA

Tumor hypoxia occurs due to the increase in demand for oxygen by the rapidly growing tumor cells

together with reduction in the supply of oxygen due to malformed and non-functional tumor vasculature.

Tumor hypoxia offers resistance to radiotherapy (RT) and chemotherapy. Interestingly, a new paradigm has

emerged suggesting that hypoxia may also suppress immunotherapy, however the mechanisms behind this

observation remain undetermined. Our laboratory and others have shown that IFN conditions the tumor

milieu and is important for the efficacy of RT. As a result, we hypothesized that hypoxia could inhibit IFN

mediated anti-tumor responses resulting in decreased RT efficacy. Hypoxia could be modulating the pro-

duction of intratumoral IFN, and/or reducing the ability of cells to respond to IFN. To test the first possi-

bility we utilized Colon-38, a murine colon adenocarcinoma tumor model, and measured intratumoral hy-

poxia by both flow cytometry and fluorescence microscopy using a monoclonal antibody that detects hy-

poxia-induced 2-nitroimidazole adducts from the drug EF5. We used this drug to demonstrate a time de-

pendent increase of hypoxia within untreated Colon-38 tumors. Unexpectedly, irradiation resulted in a de-

crease in total tumor hypoxia. We also detected various sub-populations of hypoxic immune cells in both

the untreated and irradiated groups, with macrophages being the majority population. Importantly, CD8+ T

cells, which are an important source of IFN and mediate effector anti-tumor responses, were present in

these tumors but were not co-localized with hypoxic regions. This deficiency of effector T cells in the hy-

poxic regions indicates that tumor cells in these areas may also escape from immunotherapy. To address the

second possibility of hypoxia induced inhibition we tested whether hypoxia could modulate the responsive-

ness to IFN in vitro on various cells that make up the tumor microenvironment, in particular tumor cells,

and immune cells. Indeed, we were able to demonstrate that hypoxia inhibited the induction of IFN-

stimulated genes in multiple mouse and human tumor cell lines as well as peripheral blood mononuclear

cells suggesting hypoxia could inhibit the responsiveness to IFN. We propose that tumors that are less hy-

poxic (e.g. after RT and immunotherapy) are more conducive to IFN and T cell responses, resulting in en-

hanced tumor control. Funded by NIH R01 CA28332 and AAI fellowship 058232-002.

113.

Poster 30

LP-BM5 retrovirus-augmented Monocytic Myeloid Derived Suppressor Cells, Checkpoint Regulator

VISTA, and miR-155, are all players in a profound murine Immunodeficiency-causing syndrome

Kathy A. Green 1, Li Wang1,2, Randolph J. Noelle1, and William R. Green1

1 The Geisel School of Medicine at Dartmouth, 2 Medical College of Wisconsin

Inhibition by myeloid derived suppressor cells (MDSC) against T-cell responses is well established

in tumor microenvironments. We demonstrated (Green et al., J. Virology, 2013) augmentation of mono-

cytic MDSCs (M-MDSCs) during infection of susceptible B6 mice by LP-BM5 retrovirus, which causes a

profound immunodeficiency. These M-MDSCs inhibited not only T-, but also the much less reported B-,

cell responsiveness in ex vivo suppression assays. The M-MDSC inhibition of stimulated T-cell prolifera-

tion and IFN-gamma production was almost completely iNOS/NO dependent, whereas M-MDSC suppres-

sion of B-cell responses was only ~50% iNOS/NO dependent.

In exploration of additional suppressive mechanism(s) in the unique M-MDSC inhibition of B-cell

responsiveness, we have recently reported that VISTA (Green et al., J. Virology, 2015), a newly described

negative checkpoint regulator, is involved in LP-BM5-augmented M-MDSC-mediated suppression of

stimulated ex vivo B-cell responses. We found that combining the use of reagents to block both iNOS/

NO and VISTA lead to an additive, if not synergistic, abrogation of M-MDSC suppression of B-cell re-

sponsiveness. In additional recent studies, in light of the reported significant involvement of miR-155 in B

-cell and myeloid cell function, we have focused on the use of M-MDSCs from 5-7 week LP-BM5 infected

miR-155 k.o. mice. Our results have consistently shown these M-MDSCs to be substantially less (30-

50%) suppressive compared to M-MDSCs from LP-BM5 infected Wt. mice. Also, at 10-12 weeks post

infection, miR-155 k.o. mice exhibit a selectively different kind of immunodeficiency compared to the dis-

ease of Wt. mice. Thus, spleen cells from miR-155 k.o. infected mice consistently present as significantly

more responsive to stimulation with B-cell polyclonal activators, such as anti-CD40 and IL-4, and have

significant less splenomegaly than LP-M5 infected Wt. mice. The infected spleens of 10-12 week miR-

155 k.o.s also contain substantially fewer M-MDSCs as compared to Wt. infected spleens. In an initial

experiment, involving adoptive transfer of M-MDSCs from donor LP-BM5 infected Wt. B6 to infected

miR-155 k.o. recipients, at 10 weeks post infection, we were able to significantly augment in the recipient

miR-155 k.o.: 1) splenomegaly, 2) splenic B-and T-cell immunodeficiency, and 3) the number of residing

splenic M-MDSCs, to levels almost comparable to those of Wt. infected mice.

These results highlight involvement of multiple and unique suppressive pathways in the under-

studied area of MDSC suppression of B-cell responses, and were compatible with a role for M-MDSC in

LP-BM5-induced immunodeficiency.

114.

Poster 31.

Dengue virus non-structural protein 1 activates mast cells and endothelial cells

Arturo Barbachano-Guerrero1, Timothy Endy2 and Christine A. King1 1Department of Microbiology and Immunology; 2Department of Medicine, Upstate Medical University,

Syracuse NY

Arboviruses are an emerging worldwide heath concern. Transmitted by the mosquito vector Aedes

ageypti, dengue virus, family Flaviviridae, is estimated to infect 390 million individuals annually and is

associated with enormous morbidity worldwide.

Dengue infection results in severe disease in a subset of patients characterized by pathological vascular

permeability and high levels of systemic inflammatory mediators. A small RNA virus, dengue encodes

ten proteins, of which non-structural protein one (NS1) is secreted as a hexamer during infection and re-

mains in circulation after the virus is cleared. The role of this highly conserved hexameric viral protein in

the pathogenesis of severe dengue disease is unknown.

Previous work in our laboratory and others has shown that pathogenesis of severe disease involves

activation of both innate immune mast cells (MCs) and endothelial cells (ECs). The location of MCs adja-

cent to and lining the endothelium allow for direct cross talk facilitating MC modulation of vascular he-

mostasis. Mast cells directly contribute to the systemic pro-inflammatory environment by release of large

amounts of inflammatory cytokines and chemokines in response to virus and more recent work demon-

strated modulation of EC permeability by NS1 directly.

We hypothesized that secreted NS1 promotes viral pathogenesis by directly activating MCs and

ECs leading to altered homeostasis and inflammation. To address our hypothesis we used a series of

complementary in vitro assays to asses MC and EC responses. Our data demonstrated NS1 induced acti-

vation of both MCs and ECs at multiple levels. At the transcriptional level, NS1 stimulation of ECs in-

duced the select upregulation of CCL5, E-selectin and IL-11, all markers of EC activation. Further-

more, western blot analysis of MC and ECs stimulated with increasing concentrations of NS1 demon-

strated a dose dependent increase in Akt activation at S473, a cellular hub of several regulatory and in-

flammatory pathways, including angiogenesis, vascular permeability and pro- inflammatory cytokine

production. To determine the downstream effect of NS1 activation on EC function we evaluated the

ability of NS1 to directly modulate EC intercellular organization via the ability to organize into complex

structures and to migrate. Using the tube formation and scratch assays we demonstrate NS1 potently

modifies by inducing tube formation and migration. Overall, our results demonstrate that dengue NS1

can induce MC and EC activation and potently modify basic EC functions. Together, our data suggest

that NS1 may contribute to the pathogenesis of dengue disease by directly key cellular components

linked to severe disease pathogenesis.

115.

Poster 32.

NKG2D ligand targeted Bispecific T Cell Engagers (BiTE) lead to robust antitumor activity against

diverse human tumors

Tiffany Coupet, Claire Palmer, and Charles L. Sentman

Center for Synthetic Immunity, Department of Microbiology & Immunology, Geisel School of Medicine

at Dartmouth, Lebanon, NH

Two new bispecific T cell engager (BiTE) molecules with specificity for NKG2D ligands were de-

veloped. One, designated huNKG2D-OKT3, was derived from the extracellular portion of the human

NKG2D receptor fused to a CD3ε (epsilon) binding single chain variable fragment (scFv). NKG2D is a

receptor expressed by both NK and T cells that has the ability to bind multiple ligands, including MICA,

expressed by a variety of malignant cells. A second molecule, B2-OKT3, was created in the tandem scFv

BiTE format that targets MICA on tumor cells and CD3 on T cells. The data show that these BiTEs specif-

ically activate T cells to kill several human tumor cell lines, representing both liquid and solid tumors. The

huNKG2D-OKT3 BITE had a lower affinity for recombinant MICA (rMICA) but induced greater T cell

cytotoxicity and cytokine production compared with B2-OKT3. Both BiTEs were able to trigger activated

human T cells and resting PBMCs. Culture of plate-bound rMICA with T cells and these BiTEs showed a

ligand density dependent production of IFNγ (gamma) by both CD4+ and CD8+ T cells. There was two-

fold more IFNγ (gamma) produced per CD8+ T cell and five-fold greater percentage of CD8+ T cells that

produced IFNγ (gamma) compared to CD4+ T cells. The data show robust anti-tumor activity using scFv

or receptor-based bispecific constructs and support their further development for clinical use against can-

cer.

116.

Poster 33.

Novel role for protein kinase D in regulating airway epithelial cytokine secretion and barrier

integrity

Janelle Veazey1, Tim Chapman2, Tim Smyth3, Sara Hillman2, Steve Georas1,2

Department of Microbiology and Immunology1, Department of Pulmonary and Critical Care

Medicine2, Department of Toxicology3, University of Rochester, Rochester NY

Background: Previous work from our lab identified the serine/threonine kinase protein kinase D (PKD)

as a key regulator of barrier integrity in virally-infected bronchial epithelial cells. Although PKD has been

implicated in various cellular functions including cell growth, motility, and cytokine secretion, little is

known about the expression or function of PKD in the airway.

Methods: Human bronchial epithelial cells (16HBE) cells were pre-treated with vehicle or 5 µM of the

competitive PKD inhibitor CRT0066101 (CRT) or 50 µM of the noncompetitive PKD inhibitor

CID755673 (CID) (Tocris). 2 hours later, cells were stimulated in vitro with or without the double strand-

ed RNA polyI:C (5 µg/ml). Cell supernatants were analyzed for IL-8 and IFN-lambda by ELISA (R&D

BioScience).

Wild-type 6-10 week old C57BL/6 mice were treated with either vehicle or CRT (100

µg) prior to challenge with 10 µg polyI:C. 24 hours later, bronchoalveolar lavage (BAL) fluids were col-

lected, and cell counts determined by hemacytometry and cytospin. BAL supernatants were analyzed by

ELISA for KC and IFN-lambda. PKD isoenzyme protein level in whole cell extracts of different organs

was analyzed by Western blot.

Results: Inhibiting PKD in 16HBE cells with 5 mM CRT prior to 5 µg/ml polyI:C treatment markedly

attenuated the production of both IL-8 and IFN-lambda. Using wild-type mice, we found that PKD inhi-

bition reduced polyI:C-induced KC levels and neutrophilia in BAL fluid across multiple doses of polyI:C

administered and in a dose-dependent manner with the PKD-inhibitor. The PKD inhibitor CRT also at-

tenuated polyI:C-induced IFN-lambda levels in BAL fluid. Finally, we determined that PKD3 protein

levels are more highly expressed in the lung, lymph node and spleen than PKD1, suggesting PKD3 is the

isoform driving cytokine release.

Conclusions: PKD3 may play a previously unsuspected role in regulating epithelial chemokine release

and neutrophil influx into the airway during respiratory viral infections. PKD3-mediated cytokine release

might also contribute to induction of an

anti-viral state, specifically via IFN-lambda regulation. Further study of the mechanism of PKD3-

mediated cytokine release in the airway epithelium may lead to new therapeutics in combating respirato-

ry pathogens.

117.

Poster 34.

Intestinal Immunity to Vibrio cholerae Involves Antibody-Mediated Effects on Multiple Bacterial

Cell Processes

Danielle E. Baranova, Kara J. Levinson, Nicholas J. Mantis

Wadsworth Center, New York State Department of Health, Albany NY; University at Albany, Albany NY

Secretory IgA, and to a lesser extent serum IgG, serves as a primary line of defense against enteric patho-

gens in the intestinal lumen, although exactly how antibodies protect against non-invasive bacteria like

Vibrio cholerae is not well understood. It is generally assumed that secretory antibodies function solely by

promoting cell-cell cross-linking, agglutination and subsequent clearance via peristalsis of antibody-

pathogen complexes from the intestinal lumen, a process referred to as immune exclusion. To better define

the immunological processes associated with intestinal immunity, we are investigating the interactions be-

tween V. cholerae and ZAC-3, a monoclonal antibody that recognizes a conserved surface exposed epitope

within lipopolysaccharide (LPS) coat. We have found that mAbs have direct effects on bacteria in vitro,

independent of antibody-mediated crosslinking. These effects include rapid motility arrest and the coinci-

dent appearance of surface blebs and deformations that are suggestive of outer membrane (OM) stress. We

have also shown that ZAC-3 treatment stalls V. cholerae motility in liquid and viscous environments. To-

gether this indicates that the bacterial response to secretory antibodies is multifaceted and goes beyond ag-

glutination. We now demonstrate by scanning electron microscopy that exposure of V. cholerae strain

O395 to ZAC-3 promotes the appearance of exopolysaccharide (EPS)-like extensions on the bacterial sur-

face. This EPS production is both time and dose dependent, as detected by a crystal violet assay. ZAC-3

Fab fragments did not induce EPS production, indicating that agglutination is necessary, but (from other

studies) not sufficient to trigger exopolysaccharide expression. While the exact composition of the EPS

remains unknown, we provide preliminary results that indicate that EPS production shields the bacterium

from complement-mediated killing, suggesting that V. cholerae “goes on the defensive” in response to

ZAC-3 treatment. Collectively, these results suggest a complex interplay between pathogen and secretory

antibodies in the intestinal lumen, which ultimately determines efficiency of infection.

118.

Poster 35.

Innate control of pneumonic Francisella tularensis infection through macrophage depletion

and polarization

Donald Steiner and Dennis W. Metzger

The bacterium Francisella tularensis is a Tier 1 select agent and the cause of tularemia. F. tularen-

sis is a facultative intracellular pathogen capable of replicating within the cytoplasm of macrophages, and

in the highly lethal respiratory form of tularemia, alveolar macrophages are the primary hosts in the early

stages of infection. We have found that depletion of alveolar macrophages protects mice against respirato-

ry tularemia induced using a low infectious dose of the virulent type A strain Schu S4. Mice protected in

this fashion do not develop a systemic inflammatory response, suggesting that macrophage depletion pre-

vents establishment of infection. Clodronate treatment followed by infection with antibody-opsonized F.

tularensis protects mice even against greater infectious doses. F. tularensis has been recovered from the

blood of protected mice 5 days after infection, but not from the spleens of the same mice 21 days after in-

fection, indicating that these mice develop a productive, systemic infection but ultimately clear the infec-

tion and survive. Neutrophil depletion abrogates this protection. This work represents evidence that im-

munologically naïve mice can be protected against type A F. tularensis infection through innate immunity.

Supported by NIH grant PO1 A1056320

119.

Poster 36.

House Dust Mites-induced Allergic Asthma in Sickle Cell Disease Mice

Chenyang Jiang and Steven M. Szczepanek

Department of Pathobiology and Veterinary Science, University of Connecticut, Storrs CT

House dust mites (HDM) are human allergens and the leading cause of allergic asthma in the US and

worldwide. Sickle cell disease (SCD), as the most common monogenic blood disorder, is accompanied by

massive co-morbid diseases, including allergic asthma. Asthma confers a higher rate of mortality in indi-

viduals with SCD, especially in children. Some studies demonstrated it that co-morbid allergic airway dis-

ease (AAD) in transgenic SCD mice is more severe than in wild-type (WT) mice when they are exposed to

ovalbumin (OVA) and aluminum hydroxide (alum), which is characterized by increasingly intense vaso-

occlusion and pulmonary inflammation. Given this information, we questioned if SCD mice exhibit as

fierce of co-morbid AAD when they are exposed to HDM extract, as do OVA/alum sensitized SCD mice.

To study this, SCD mice were intranasally inoculated with HDM for acute (5 weeks) and chronic (11

weeks) periods. Unexpectedly, when SCD mice were instilled with a low dose (1 ug (microgram)) of

HDM for an acute time period, the amount of total white blood cells per ml of bronchoalveolar lavage flu-

id (BALF) was similar to the amount found in WT mice. BALF leukocyte differential counting was not

statistically different between SCD mice and WT mice, with eosinophils approximating 23% and macro-

phages 77% of cells. Consistent with this, inflammation scoring for histologic slides of lungs from SCD

mice was not significantly different with scores from WT mice. After an acute exposure of high dose

HDM (25 ug(microgram)), the amount of total white blood cells per ml BALF, BALF leukocyte differen-

tial counting and inflammation scoring for histologic slides of lungs from SCD mice were same as those

from WT mice. In this case, eosinophils were the majority at approximately 81% instead of macrophages

at 17%. As with acute disease, chronic exposure of SCD and WT mice to a high dose of HDM resulted in

statistically similar results for all parameters. Acute inflammation turned into immune tolerance, as eosino-

phils decreased to approximately 40% and macrophages increased to 52% of BALF leukocytes in both

groups. These data indicate that HDM exposure causes similar AAD in SCD mice and WT mice, which is

in stark contrast to the reports of severe AAD in SCD mice when they are run through the OVA model.

HDM are much more complicated allergens than OVA/alum, and our results highlight the need to better

understand the dynamics of SCD lung disease and co-morbid allergic asthma.

120.

Poster 37.

Stress Mediated Immunomodulation: Role of Metallothionein in the extracellular environment

Sadikshya Bhandari, Michael A. Lynes

University of Connecticut, Storrs, CT

Metallothioneins (MTs) are highly conserved stress response proteins, which are up-regulated after

exposure to heavy metals or reactive oxygen and nitrogen species. Of the 61 amino acids present in a con-

sensus mammalian MT sequence, 20 are cysteine residues that are integral to this protein’s biological prop-

erties. Traditionally, research on MT focused on its intracellular roles as a free radical scavenger, metal

reservoir and regulator of NF-kB and Sp-1 transcription factor activities. Though MT lacks a signal peptide

sequence, its presence in extracellular spaces such as milk, bronchoalveolar and prostatic fluids, bile, liver

sinusoids, serum etc. has been well documented. MT has also been detected at sites of inflammation and

tissue wounding. We have previously shown that MT can bind to the surface of lymphocytes, is a potent

chemoattractant for T cells and this effect can be blocked with antagonists of G-protein coupled receptors.

When T cells are pre-incubated with MT, there is a significant and substantial decrease in their chemotac-

tic response towards SDF-1a. This pre-incubation does not interfere with internalization of the cognate

SDF-1a receptor (CXCR4) that accompanies SDF-1a induced chemotaxis. Pre-incubation of MT with T

cells also does not change intracellular reactive oxygen species (ROS) levels. MT can also block SDF-1a

induced chemotactic responses of a both primary splenocytes and a breast cancer cell line, MDA-MB 231

(ATCC HTB-26), suggesting that this is not a cell-specific phenomenon. Our most recent studies have

used specific MT peptides to identify those MT sequences that can interfere with SDF-1a mediated chem-

otaxis. The presence of MT in the extracellular environment and its novel chemotactic properties suggest a

possible immunomodulatory role in inflammatory diseases and in the progression of metastatic cell move-

ment. Hence, MT manipulation under these conditions may provide a possible avenue of therapeutic inter-

vention.

121.

Poster 38.

Commensal microbes drive the generation of systemic IgA responses

Joel R. Wilmore, Brian Gaudette, Wenzhao Meng, Eline T. Luning Prak, and David Allman

Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of

Pennsylvania, Philadelphia, PA

It is well known that IgA functions as a critical component of the mucosal barrier in the gut by vir-

tue of its ability to be secreted into the intestinal lumen. However, little is known about the origin, function

and regulation of IgA in the serum. Mucosal IgA responses have been generally thought of as short-lived

and restricted locally to mucosal tissues. However, we find that IgA-secreting PCs make up the majority of

the BM PC pool in standard C57BL/6 mice bred in our colony. The IgA+ BM PCs are predominantly in the

long-lived pool and express gut homing factors such as CCR9 and the integrin α4β7 (alpha4 beta7), sug-

gesting a mucosal origin. The extent to which the commensal microflora influences the BM plasma cell

pool is evident by mice bred in germ free isolators that lack IgA+ PCs in their BM. Additionally, mice from

standard vendors such as Jackson labs (Jax), have extremely low levels of IgA+ PCs in the BM and signifi-

cantly lower serum IgA. Exposing Jax mice to a disparate microflora or Helicobacter sp. led to the genera-

tion of IgA-secreting BM cells, while also inducing increases in serum IgA antibodies enriched for binding

to several commensal bacterial taxa. Moreover, BM IgA-secreting plasma cells exhibited a common clonal

ancestry with intestinal IgA+ plasma cells, and both populations possessed unique gene expression signa-

tures compared to other long-lived BM plasma cells. We conclude that commensal microbes overtly influ-

ence the BM plasma cell pool, and suggest that select commensal microbes can facilitate the induction of

systemic humoral immunity.

122.

Poster 39.

Elucidating the Mechanism of Protective Antibodies Against the Enteric Pathogen Salmonella

Typhimurium at Epithelial Surfaces

Angelene F. Richards1, 2, John J. Varrone1, Jennifer E. Westfall1, and Nicholas J. Mantis1, 2

1Division of Infectious Diseases, Wadsworth Center, New York State Department of Health, Albany, NY

12208; 2Department of Biomedical Sciences, University at Albany School of Public Health,

Albany, NY 12208

Salmonella enterica serovar Typhimurium (ST) is a leading cause of gastroenteritis in the United

States and an emerging cause of invasive non-typhoidal salmonella (NTS) in sub-Saharan Africa. Inva-

sion of the intestinal mucosa by ST involves a complex series of events in which the bacteria utilize flagel-

la-based motility and type 3 secretion systems (T3SS) to adhere to and invade Peyer’s patch tissues before

spreading systemically to the liver and spleen. Here we confirm in an adult mouse model that Sal4, an IgA

monoclonal antibody (mAb) against ST lipopolysaccharide (LPS), blocks bacterial entry into Peyer’s

patch tissues. We show for the first time, however, that an IgG1-derivative of Sal4 when co-gavaged with

ST does not prevent bacterial invasion of Peyer’s patch tissues, even though the same mAb did limit ST

spread to the liver and spleen when administered systemically prior to ST injection into the peritoneal cav-

ity. These results illustrate the importance of IgA in mucosal protection against ST infection and provide a

tractable model to begin to define mechanisms by which secretory antibodies limit bacterial uptake into

Peyer’s patch tissues. With this long goal in mind we generated a second IgA mAb against ST LPS,

named PeA3, which binds an epitope similar to but distinct from Sal4. PeA3 was as effective as Sal4 IgA

at limiting ST infection of mouse Peyer’s patch tissues, indicating that Sal4’s biological activities are not

unique. We have now embarked on a comparative in vitro analysis of Sal4 IgA, Sal4 IgG and PeA3 IgA

mAbs in an effort to define specific properties associated with protective immunity. Sal4 IgA and IgG, as

well as PeA3 IgA, are each potent inhibitors of ST-flagellum based motility in semi-solid agar and liquid

media indicating that they are effective at limiting early steps in the infectious process. However, in vitro

analysis did demonstrate that Sal4 IgA and PeA3 IgA mAbs were more effective than Sal4 IgG at promot-

ing ST agglutination. It has been hypothesized that IgA-mediated agglutination of bacterial pathogens in

the intestinal lumen results in the formation of immune complexes that are readily trapped in mucus and

cleared via peristalsis. Our future goals are to test this hypothesis using in vitro and ex vivo assays, in-

cluding organoids to mimic the intestinal lumen. A better understanding of the mechanisms by which se-

cretory IgA functions in mucosal immunity has important implications for oral vaccine development

against an array of enteric pathogens.

123.

Poster 40.

Macrophages Negatively Regulate Hematopoietic Stem Cells in Murine Aplastic Anemia

Angelica Costello, Amanda McCabe, PhD, Julianne N.P. Smith, PhD, & Katherine C. MacNamara, PhD


Aplastic anemia (AA) is a rare bone marrow (BM) failure syndrome characterized by T cell-

mediated bone marrow destruction and pancytopenia. AA can be genetic or acquired, with the latter caused

by such insults as radiation, chemicals, drugs, and infection. T cell-derived interferon-γ (IFN-γ) has been

implicated in driving disease, but the mechanisms of IFN-γ-mediated pathogenesis in hematopoietic failure

during AA are not well understood. We recently identified a role for IFN-γ signaling specifically in macro-

phages (Mϕs) in driving the transient loss of hematopoietic stem cells (HSCs) in murine ehrlichiosis. Here,

in a mouse model of AA, we find that BM-resident Mϕs are maintained in the BM despite the loss of other

hematopoietic cell types, and that this requires IFN-γ. Moreover, Mϕ depletion with clodronate liposomes

or abrogation of IFN-γ signaling in Mϕs during AA rescues the HSC pool and significantly improves sur-

vival. We observed similar numbers of T-bet+ T cells and comparable IFN-γ production in AA mice that

were depleted of Mϕs, suggesting that Mϕs are critical sensors of IFN-γ and drivers of disease during AA.

Despite this, we did not detect a significant reduction in inflammatory proteins when IFN-γ signaling is ab-

rogated or when Mϕs were depleted. The one notable exception, however, was the chemokine CCL5

(RANTES), which was highly expressed in the BM during AA and significantly reduced in mice depleted

of Mϕs or when Mϕs were unable to respond to IFN-γ. CCL5 production in BM fibroblastic cells is in-

duced via podoplanin (PDPN) signaling. In support of a role of PDPN signaling in driving increased CCL5

during AA, we detected a significant increase in PDPN+ BM-resident Mϕs during AA. Neutralization of

either CCL5 or PDPN during AA rescued HSC numbers. Moreover, anti-PDPN treatment rescued platelet

numbers and significantly improved survival. Altogether, we demonstrate a novel role for IFN-g in AA

pathogenesis whereby IFN-γ promotes increased PDPN+ Mϕs and enhanced production of CCL5, thus

driving HSC loss and thrombocytopenia.

124.

Poster 41.

Neutralizing Camelid Antibodies Target the Binding Subunit of Ricin by Interfering with

Receptor Attachment and Intracellular Trafficking

Amanda Y. Poon, David J. Vance, Yinghui Rong, Nicholas J. Mantis Wadsworth

Center, New York State Department of Health, Albany NY 12208 Department of Bio-

medical Sciences, University at Albany, Albany NY 12208

Ricin toxin is a ribosome-inactivating protein (RIP) and classified by the Center for Disease Control

and Prevention as a potential bioterrorism agent. Ricin’s B-subunit (RTB) is a lectin that mediates toxin

attachment, internalization, and intracellular trafficking of ricin in mammalian cells, and is therefore

indispensable for ricin’s cytotoxic activity. RTB (32 kDa) consists of two homologous domains (1 and

2), each divided into three subdomains (a, b, g), with lectin activity restricted to subdomains 1a

and 2g.

Fig.1: Linear depiction of RTB

Theoretically, blocking RTB binding and/or trafficking activity is an ideal means by which to neu-

tralize ricin. However, analysis of dozens of RTB-specific monoclonal antibodies (mAbs) indicate

that toxin-neutralizing antibodies are extremely rare and likely target a very limited number of

epitopes, possibly in subdomains 1a and 2g. To better understand RTB-antibody interactions we

have embarked on a concerted effort to generate a comprehensive B cell epitope map of RTB using a

highly diverse ricin-specific camelid single domain antibody (VHH) library. Currently, we have isolated

a panel of RTB-specific VHHs with a range of affinities and in vitro toxin-neutralizing activities. Using

a novel biotinylated-ricin competition ELISA, we observed that VHHs with the best toxin-neutralizing

activity tend to recognize epitopes within the vicinity of subdomain 2g, whereas VHHs with weak neu-

tralizing activity cluster near 1a. We next tested the hypothesis, using a solid phase receptor bind-

ing assay, as well as THP-1 cells in a flow cytometric based readout, that ricin neutralizing activity

correlates with the ability of VHHs to interfere with toxin attachment to mammalian cell surfaces. The

hypothesis proved incorrect, as neutralizing and non-neutralizing VHHs interfered with ricin attach-

ment to similar degrees. Based on these and other results, we are now investigating the possibility that

toxin-neutralizing activity by RTB-specific VHHs is not due to interference of cell attachment, but rather

effects on toxin endocytosis and/or intracellular trafficking. At the same time we have initiated efforts

to perform more detailed epitope mapping studies by X-ray crystallography (X-TAL) and hydrogen-

deuterium exchange coupled with mass spectrometry (HX-MS). As RTB is the prototypic member

of a large family of toxin and bacterial R-type lectins we expect our results will afford insights into

basic mechanisms of pathogen-host interactions.

125.

Poster 42.

The lymphoproliferative effects of the small stress response protein, metallothionein

Kristen E. Dostie, Michael A. Lynes.

University of Connecticut, Storrs CT

Metallothioneins are small metal-binding proteins that can alter a variety of immune responses,

and, for example, can influence the progression of autoimmune diseases and the response to bacterial in-

fection. MTs are biochemically unique proteins due to their low molecular weight and unusually high cys-

teine content, which accounts for ~33 mol% of the total amino acid composition. These cysteines within

MT provide sulfhydryl groups which bind essential metal cations such as zinc and copper and can also

serve to regulate the local redox environment. Although MT lacks an N-terminal signal peptide and has

conventionally been considered an intracellular protein, it has been found in extracellular compartments

where it can act as a potential “danger signal” that can mediate both innate and adaptive immune respons-

es. Previous work of our laboratory has shown that MT binds to the surface of lymphocytes and can lead to

proliferation of lymphocytes. Interestingly, MT can synergistically enhance the proliferative response of

lymphocytes when co-administered with either T- or B-cell mitogens. The aim of this research is to further

characterize the mechanism responsible for the enhanced proliferative response to MT of various lympho-

cyte subsets. Our work profiles the relative binding affinity of MT to surface molecules of specific lym-

phocyte populations. These relative binding affinities of MT for different lymphocyte populations may

provide insight to potential MT receptors on the cell surface.

126.

Poster 43.

The Role of Pseudomonas aeruginosa Metallothionein, PmtA, in Immune Modulation

Amy Thees, Kathryn Pietrosimone, Brian Greco, Michael A. Lynes

Pseudomonas aeruginosa is a gram-negative, opportunistic pathogen, frequently associated with hospital-

acquired infections. Chronic infections of Pseudomonas aeruginosa are common in cystic fibrosis patients

due to exaggerated and ineffective airway inflammatory processes that fail to eradicate this pathogen. A

novel bacterial stress protein, PmtA, has been shown to contribute to bacterial survival by influencing the

host immune response. Disruption of the PmtA gene production causes a decrease in the production of a

redox-active, secondary metabolite, pyocyanin. It has also been demonstrated that PmtA blocks the migra-

tion of cells in response to SDF-1 alpha and decreases SDF-1 alpha-induced internalization of CXCR4 on

Jurkat T cells. Here we show that PmtA can be found on the surface of leukocytes in culture after expo-

sure to exogenous MT, which suggests that leukocytes may express one or more receptors for PmtA. Ex-

posure to PmtA also enhances proliferation of Jurkat T cells. Pseudomonas aeruginosa is known for the

multitude of virulence factors that it can produce. PmtA may represent an additional tool that Pseudomo-

nas aeruginosa can employ to sustain an infection.

127.

Poster 44.

Regulation of IgM Memory B cell Pool Size by the Inhibitory receptor FcgRIIb

Amber Papillion and Gary Winslow

Upstate Medical University, Syracuse, NY

Ehrlichia muris infection generates a long-term CD11c/T-bet-positive IgM memory population in

the spleen (J. Immunol. 191:1240). Among the many surface markers that distinguish the IgM memory

cells from canonical B cells is the inhibitory Fc receptor, FcgRIIb, which exhibited a 2-fold higher expres-

sion. We hypothesized FcgRIIb negatively regulates IgM memory cells by binding immune complexes

present during low-level chronic infection. To investigate this question, we monitored the IgM memory

cell population in infected FcgRIIb-deficient mice. Thirty days post-infection, the IgM memory B cells

were generated earlier, and were found at three-fold higher frequencies in FcgRIIb-deficient mice, com-

pared to wild-type mice. This increase in the frequency of spleen IgM memory cells was due to an increase

in cell number, and was in turn associated with an increase in antigen-specific IgG. These data indicate

that FcgRIIb plays an important role in regulating the expansion and/or persistence of IgM memory cells in

wild-type mice under conditions where antigen-specific IgG is sufficient to control infection. Other studies

revealed that the IgM memory population in FcgRIIb-deficient mice exhibited much lower expression of

FAS, CD40, BAFF-R, and TACI. We therefore proposed that FcgRIIb signaling, likely via immune com-

plexes, acts in wild-type cells to regulate the size of the IgM memory cell pool, by maintaining the expres-

sion of FAS and other receptors that regulate cell survival. These data suggest a novel regulatory role for

FcgRIIb in B cell memory

128.

Poster 45.

Skin Microbiome and the Development of Autoimmune Vitiligo

Jennifer Vella, Walburga Croteau, Yina Huang

Dartmouth College, Hanover NH

Skin cancer is the most common form of cancer and can be classified into two groups: melanoma

and non-melanoma, with melanoma being the more dangerous of the two. Melanoma arises when our pig-

ment producing cells, called melanocytes, turn cancerous, gaining the ability to rapidly divide and metasta-

size to other organs throughout the body. Melanoma patients who simultaneously develop autoimmune

vitiligo have a more favorable prognosis, suggesting therapies increasing vitiligo may be beneficial. While

studies have shown that autoimmune vitiligo is a CD8 T cell mediated disease, the mechanisms responsi-

ble for its development remain unclear. By utilizing a mouse model we are able to mimic melanoma-

induced vitiligo allowing us to study potential triggers of this autoimmune disease. With this method we

compared the development of vitiligo in response to various treatments. We found that mice treated with

broad-spectrum antibiotics were less likely to develop vitiligo compared to their untreated littermates. In

addition to the decrease vitiligo incidence, the mice that did develop vitiligo only developed it locally,

without dissemination of depigmentation. Our studies indicate that the skin microbiome may play an im-

portant role in the development and progression of autoimmune vitiligo.

129.

Poster 46

Hypofractionated vs. hyperfractionated radiotherapy treatment alters immune cell viability and

function

Margaret L. Barlow1, Nicholas Battaglia1, Scott A. Gerber1, and Edith M. Lord1

1Department of Microbiology and Immunology, University of Rochester, Rochester, NY 14642

Radiotherapy is one of the most effective means of treating solid tumors in cancer patients. This is

due to the radiation acting in two ways to control tumor growth. First, by inducing cell death due to direct

or indirect damage to the DNA, then by the killed tumor cells releasing tumor antigens and inflammatory

signals that boost the antitumor immune response. Unfortunately, radiotherapy may also damage the

healthy tissue surrounding the tumor, causing adverse side effects in patients. To limit this damage, radio-

therapy in the clinic is generally delivered as multiple small doses (hyperfractionation) over the course of

several weeks. However, in preclinical radiotherapy studies, radiation is often given as a single or a few

high ablative doses (hypofractionation). We previously observed improved tumor growth control and an

increased number of tumor infiltrating lymphocytes (TIL) in tumors given a single ablative dose when

compared to a hyperfractionated scheme. As TILs are radiosensitive, we hypothesize that the repeated dos-

es of a hyperfractionated scheme may result in increased killing of the TILs as they infiltrate the tumor,

damaging them as they attempt to carry out their effector response and thereby reducing the effectiveness

of the anti-tumor immune response. To further investigate this, we created two radiation dosing schedules

based on current clinical protocols. Our hypofractionated scheme used either one or two doses of 10 Gy,

while our hyperfractionated scheme used five doses of 2 Gy. We compared the responses of the mouse

Colon38 tumor model to these schemes. In vitro, the hypofractionation treatment was more effective at

killing tumor cells and inducing the tumor cells to produce the inflammatory cytokines IL-β(beta), TNFa

(alpha), IL-6, as well as the chemoattractant IP-10. In vivo, the hypofractionated treatment resulted in a sig-

nificant delay in tumor outgrowth and improved survival rates when compared to the hyperfractionated

treatment. We also observed fewer apoptotic or dead effector CD8+ T cells before the second dose of 10 Gy

in the hypofractionated scheme, compared to 24 hours after the second 10 Gy dose, suggesting that some

of these cells were damaged by the repeated radiation. Therefore, hypofractionated radiotherapy may in-

crease immunity and preserve immune cell viability and function. Funded by NIH R01CA28332 and

T32AI007285.

130.

Poster 47.

The role of Indoleamine 2,3-dioxygenase (IDO) in the survival of bone marrow resident long lived

Plasma cells

Shivana M. Maharaj, Louise M. Carlson, and Kelvin P. Lee

Department of Immunology, Roswell Park Cancer Institute; Buffalo, NY

Long lived plasma cells (LLPCs) are essential for sustained antibody responses and protective hu-

moral immunity. How these cells maintain longevity and a durable antibody response is largely dependent

on the complex nature of the bone marrow microenvironment in which these cells reside, and the pro sur-

vival factors produced in this niche. Work in our lab has shown that CD28 is required for LLPC survival,

and that Indoleamine 2,3-dioxygenase (IDO), an enzyme that catabolizes tryptophan, is produced as a re-

sult of back signaling from CD28:CD80/86 interactions between plasma cells and bone marrow myeloid

derived DCs. IDO is classically known to contribute to an immunosuppressive environment through its

depletion of tryptophan (specifically with respect to T cells). However, to our great surprise we found that

there are fewer LLPCs present in the bone marrow of IDO knockout mice in comparison to wild type

mice. Additionally, live cell numbers of purified plasma cells in vitro in tryptophan free media and IDO

conditioned media are greater than in media alone. These conditions also caused autophagy to occur in

these plasma cells, a process shown to be required for LLPC survival. This leads us to propose a model

where CD28, through back signaling to CD80/86, induces IDO production which leads to depletion of

tryptophan and causes induction of autophagy, a mechanism LLPCs use to compete and survive within

the bone marrow microenvironment.

Funding: T32 CA085183

131.

Poster 48.

Pretreatment Peripheral Blood Monocyte Subset Signature is Predictive of Patient Response to

Dendritic Cell Vaccination

Anand Sharda1, Alexander Wald1

, Mohammad Habiby Kermany1, Katja Koeppen2

, Thomas Hampton2, Jan

Fisher3, Camilo Fadul3

, Marc Ernstoff4, Thomas Schwaab1

, Jason Muhitch1,5

Department of Urology1, Medicine4

, and Immunology5 Roswell Park Cancer Institute, Buffalo, NY Depart-

ments of Microbiology and Immunology2, and Medicine3 Geisel School of Medicine at Dartmouth,

Hanover, NH

Clinical trials have demonstrated that dendritic cell (DC) vaccination can initiate durable anti-tumor

immunity in a subset of cancer patients resulting in complete responses, even in stage IV Renal Cell Carci-

noma (RCC). The influence of monocytes, the starting material for conventional DC vaccines, on patient

responses remains under-investigated. Recently, three subsets of monocytes have been described (classical,

intermediate, and non-classical), each with distinct functional properties. However, their roles in anti-

tumor immunity, particularly in the context of DC vaccination, are unclear. The goal of this study was to

determine whether the circulating pretreatment monocyte subset gene expression and composition from

Stage IV RCC patients prior to DC vaccination predicted responses to treatment in a completed phase II

clinical trial (NCT00085436). Pretreatment circulating classical (CD14++, CD16-), intermediate (CD14++,

CD16+), and non-classical (CD14+, CD16++) monocyte subsets were isolated from patients. Pretreatment

peripheral blood from complete responders (2 of 3 have no observable disease > 5 years following therapy)

contained fewer classical monocytes (57.5% ± 6.4) compared to all other groups (P < 0.05). Interestingly, a

higher percentage of DC derived from non-classical monocytes expressed costimulatory molecules (CD80;

96.4%, CD86; 91.1%, HLA-DR; 99.1%) compared to classical monocyte-derived DC (64.6%, 54.4%,

60.2%, respectively). DC derived from non-classical monocytes were also superior in their ability to in-

duce allogeneic T cell proliferation compared to DC originating from classical monocytes. Additional gene

expression analysis by unsupervised hierarchical clustering clearly distinguished the transcriptional profile

of classical, intermediate, and non-classical monocytes from RCC patients to healthy controls. Further in-

vestigation revealed that monocytes from long term survivors (> 10 years) could be distinctly segregated

from other RCC patients. These findings demonstrate that DC-derived from the minor CD16+ monocyte

subset may represent a superior product for use in vaccination protocols. Gene expression profiling of cir-

culating monocytes may provide an accessible biomarker for patient responsiveness to immunotherapy.

Future studies will address whether pretreatment levels of intermediate and non-classical monocytes are

prognostic indicators for response to additional immunotherapies, including checkpoint blockade inhibi-

tors.

132.

Poster 49.

Antagonistic Control of Intestinal Wnt Expression by IBD-Associated Cytokines

Travis Walrath, Stephen Sharp M.D., Shanti D'Souza, Prabhu Tewari, and William O'Connor Jr, Ph. D.

Albany Medical College, Albany NY

Homeostasis of the intestinal epithelium depends on the complex interplay of a variety of mediators

including cytokines that control, among other processes, a gradient of Wnt proteins. Homeostasis can be

disrupted by acute and chronic inflammation such as Inflammatory Bowel Disease (IBD). During IBD a

high concentration of the TH1 and TH17 associated cytokines, interferon gamma (IFN-γ) and interleukin-

17A (IL-17A) respectively, is observed within the colonic lamina propria. IFN-γ has previously been

shown to be cytotoxic to the intestinal epithelium, in part through interference with canonical Wnt signal-

ing; this is associated with increased disease severity in IBD. In murine models of disease, IL-17A has

been shown to lessen the severity of induced IBD.

One important question in the field is how these cytokines govern the intestinal epithelial and adja-

cent lymphoid compartments to either amplify inflammation or support mucosal healing. Toward under-

standing how these cytokines regulate the colonic microenvironment, we undertook a study using murine

knockout strains, primary tissue treated ex vivo, and primary spheroid/organoid cultures treated with IFN-γ,

IL-17A, or a combination of both. We found that IFN-γ reduced canonical Wnt expression while inducing

expression of non-canonical Wnts in primary murine colon tissue. Additionally we found that this altered

balance of wnt expression is dependent on the presence of lymphocytes and further, that wnt5a is specifi-

cally induced in CD19+ cells in the colon. Interestingly, IL-17A was able to oppose the increased expres-

sion of wnt5a. Moreover, mice deficient in IL-17A exhibit enhanced colonic wnt5a and TH1 cell marker

expression in response to Citrobacter rodentium infection, suggesting that IL-17A is opposing infection-

induced wnt5a and the TH1 bias during active colonic inflammation.

Canonical wnts, such as Wnt3a, are necessary for maintaining the potency of stem cells in the in-

testinal stem cell niche and the proliferative capacity of the intestinal epithelium. Non-canonical wnts such

as Wnt5a, have been shown to inhibit canonical wnt signaling and support tissue damage during colitis by

supporting increased IFN-γ production. Therefore, our data suggest that IFN-γ promotes intestinal epitheli-

al dysfunction in part by promoting a shift from canonical to non-canonical Wnt expression and function in

the colon. Furthermore, we hypothesize that IL-17A interferes with the IFN-g mediated shift in Wnt ex-

pression, and therefore may reduce IFN-g mediated epithelial toxicity and facilitate mucosal healing.

133.

Authors Index

CS – Corporate Speaker P - Poster Number

KS - Keynote Speaker S – Speaker

O - Oral Poster Presentation WP – Workshop Presenter

Page (s)

*************************************************************************************

Abrams, Scott I. ................................................................................................................... 50, 52, 97, 107, 110

Allman, David ..................................................................................................................... 59, 119

Amiel, Eyal .......................................................................................................................... 64, 104

Amobi, Adaobi (P26, O) ...................................................................................................... 51, 109

Appenheimer, Michelle ....................................................................................................... 52, 110

Arend, Kyle ......................................................................................................................... 85

August, Avery ...................................................................................................................... 44, 89

Baranova, Danielle E. (P34) ................................................................................................ 117

Barbachano-Guerrero, Arturo (P31) .................................................................................... 79, 114

Barbi, Joseph (S) .................................................................................................................. 72

Barlow, Margaret L. (P46) ................................................................................................... 129

Battaglia, Nicholas ............................................................................................................... 129

Benoit, Patrick ..................................................................................................................... 47, 103

Besra, Gurdyal ..................................................................................................................... 46, 100

Bhandari, Sadikshya (P37) .................................................................................................. 120

Bhat, Tariq A. (P16) ............................................................................................................ 99

Blader, Ira J. (S) ................................................................................................................... 81

Bogner, Paul N. ................................................................................................................... 99

Boule, Lisbeth A. ................................................................................................................. 111

Boyson, Jonathan E. ............................................................................................................ 47, 103

Braciale, Thomas J. (WP, KS) ............................................................................................. 76, 82

Budd, Ralph ......................................................................................................................... 73

Burke, Catherine G.(P28) .................................................................................................... 111

Buskiewicz, Iwona A. (S) .................................................................................................... 73

Carlson, Louise M. .............................................................................................................. 48, 58, 106, 130

Carlson, Bradley A. ............................................................................................................. 102

Chan, Gary C. ...................................................................................................................... 85, 108

Chapman, Timothy J. (S) ..................................................................................................... 74, 116

Consiglio, Camila Rosat (P18, O) ....................................................................................... 63, 101

Cojohari, Olesea (P25) ........................................................................................................ 85, 108

Cooper, James ...................................................................................................................... 58, 106

Costello, Angelica (P40, O) ................................................................................................. 65, 123

Coupet, Tiffany (P32) .......................................................................................................... 115

Croteau, Walburga ............................................................................................................... 128

134.

Cubitt, Rebecca L. ................................................................................................................ 43, 86, 88

D’Alessandro, Angelo .......................................................................................................... 64, 104

Davenport, Miles P. ............................................................................................................. 40

DeVault, Victoria L. (P20, O) .............................................................................................. 47, 103

Diehl, Michael ..................................................................................................................... 50, 107

Dienz, Oliver ........................................................................................................................ 47, 103

Dostie, Kristen E. (P42) ....................................................................................................... 125

D’Souza, Shanti ................................................................................................................... 66

Elliott, Michael R. ................................................................................................................ 93

Emo, Jason A. (P7) .............................................................................................................. 74, 90

Endy, Timothy ..................................................................................................................... 114

Enelow, Richard I. (S) ......................................................................................................... 79

Ernstoff, Marc ...................................................................................................................... 131

Evans, Sharon S. .................................................................................................................. 50, 52, 107, 110

Fadul, Camilo ....................................................................................................................... 53, 131

Fino, Kristin ......................................................................................................................... 98

Fisher Jan ............................................................................................................................. 53, 131

Gaudette, Brian .................................................................................................................... 59, 121

Gerber, Scott A. ................................................................................................................... 112, 129

Georas, Steve N. .................................................................................................................. 74, 116

Ginty, Fiona (S) ................................................................................................................... 71

Gollnick, Sandra .................................................................................................................. 63, 101

Greco, Brian ......................................................................................................................... 126

Green, Kathy A. (P30) ......................................................................................................... 113

Green, William R. ............................................................................................................... 113

Grimson, Andrew ................................................................................................................. 40

Hägglöf, Thomas .................................................................................................................. 46, 100

Hampton, Thomas ................................................................................................................ 53, 131

Harton, Jonathan A. ............................................................................................................. 92

Haque, Mohammad ............................................................................................................. 98

Hartley, Richard .................................................................................................................. 73

Henry, Everett ...................................................................................................................... 86

Hillman, Sara E. ................................................................................................................... 74, 116

Huang, Weishan (P6, O) ....................................................................................................... 44, 89

Huang, Yina ......................................................................................................................... 128

Hyuck, Heidie ...................................................................................................................... 90

Jiang, Chenyang (P36) ......................................................................................................... 119

Kalathil, Suresh G. ............................................................................................................... 99

Karlsson, Mikael .................................................................................................................. 46, 100

Katkere, Bhuvana (P19) ....................................................................................................... 102

135.

Kearsing, Lori ...................................................................................................................... 41

Kelly, Ryan .......................................................................................................................... 73

Kenderes, Kevin (P22, O) .................................................................................................... 57, 105

Kermany, Mohammad Habiby ............................................................................................ 53, 131

Khan, ANM Nazmul H. ....................................................................................................... 49, 87

King, Christine A. (S) .......................................................................................................... 79, 114

Kiramanjeswara, Girish S. ................................................................................................... 102

Knowlden, Sara A. ............................................................................................................... 73

Koch, Cameron J. ................................................................................................................ 111

Koenig, Andreas .................................................................................................................. 73

Koeppen, Katja .................................................................................................................... 53, 131

Koylass, Nicholas ................................................................................................................ 44, 89

Ku, Amy W. (P27, O) .......................................................................................................... 50, 52, 107, 110

Lang, Sabine ........................................................................................................................ 84

Lanthier, Paula ..................................................................................................................... 46, 100

Lawrence, B. Paige .............................................................................................................. 45, 94, 111

Lawrence, David A. (S) ....................................................................................................... 41

Leadbetter, Elizabeth .......................................................................................................... 46, 100

Lee, Kelvin P. ...................................................................................................................... 48, 58, 106, 130

Levinson, Nicholas J. ........................................................................................................... 117

Lilley, Graham W.J. ............................................................................................................ 47, 103

Lord, Edith M. ..................................................................................................................... 112, 129

Lubliner, Jane ...................................................................................................................... 41

Lugade, Amit A. .................................................................................................................. 99

Lynes, Michael A. (S) .......................................................................................................... 36, 120, 125, 126

MacNamara, Katherine C. ................................................................................................... 65, 123

Maharaj, Shivana M. (P47) .................................................................................................. 130

Mahpour, Amin ................................................................................................................... 58, 106

Mantis, Nicholas J. .............................................................................................................. 117, 122, 124

Markley, Rachael L. ............................................................................................................ 102

Martinez, Natalie ................................................................................................................. 90

Massa, Paul T. ..................................................................................................................... 62, 91

McCabe, Amanda ................................................................................................................ 65, 123

Mendoza, Alvaro ................................................................................................................. 41

Meng, Wenzhao ................................................................................................................... 59, 121

Metzger, Dennis W. ............................................................................................................. 95, 118

Miller, A. ............................................................................................................................. 97

Minchenberg, Scott B. (P8, O) ............................................................................................ 62, 91

Minderman, Hans ................................................................................................................ 50, 107

Mohrs, Katja ........................................................................................................................ 46, 100

136.

Mondal, Tapan .................................................................................................................... 41

Montgomery, Theresa ......................................................................................................... 73

Moon, Taylor J. (P10) ......................................................................................................... 93

Moorman, Nathaniel ............................................................................................................ 85

Moshkani, Safiehkhatoon (P1) ............................................................................................ 84

Moysich, Kirsten B. ............................................................................................................ 49, 87

Muhitch, Jason ................................................................................................................... 50, 52, 53, 107, 110, 131

Murphy, Michael ................................................................................................................ 73

Murthy, Aditi (P29) ............................................................................................................. 112

Myers, Jason R. ................................................................................................................... 45, 94, 111

Nagar, Abhinit (P9) ............................................................................................................. 92

Nemeth, M. .......................................................................................................................... 97

Noelle, Randolph J. ............................................................................................................. 113

O’Connor, William .............................................................................................................. 66, 132

Odunsi, Kunle ...................................................................................................................... 50, 51, 87, 109

O’Loughlin, Kieran ............................................................................................................. 50, 107

Ostrand-Rosenberg, Suzanne .............................................................................................. 50, 107

Oyesola, Oyebola (P5, O) .................................................................................................... 43, 88

Papillion, Amber (P44, O) ................................................................................................... 57, 60, 105, 127

Palmer, Claire (P32) ............................................................................................................ 115

Pan, Fan .............................................................................................................................. 70

Pardoll, Drew ...................................................................................................................... 70

Paredes, Anthony F. ............................................................................................................ 62, 91

Peng, Peng ........................................................................................................................... 58, 106

Peppenelli, Megan A. (P2) .................................................................................................. 85, 108

Perl, Andras ......................................................................................................................... 73

Phipps, Richard P. ............................................................................................................... 99

Pietrosimone, Kathryn ......................................................................................................... 126

Poon, Amanda Y. (P41) ...................................................................................................... 124

Post, Christina M. (P11, O) ................................................................................................. 45, 94

Powers, Colin A. (P24, O) ................................................................................................... 50, 107

Prabhu, Kumble S. .............................................................................................................. 102

Prak, Eline T. Luning .......................................................................................................... 59, 121

Pryhuber, Gloria .................................................................................................................. 90

Rahman, Tabassum ............................................................................................................. 92

Rahman, Ziaur S.M. ............................................................................................................ 56, 96

Reynaldi, Arnold ................................................................................................................. 40

Richards, Angelene F. (P39) ............................................................................................... 122

Robek, Michael D. .............................................................................................................. 84

Rodriguez, Princess ............................................................................................................. 64, 104

137.

Rong, Yinghui ..................................................................................................................... 124

Rose, John K. ....................................................................................................................... 84

Rudd, Brian D. (S) ............................................................................................................... 40

Sandhu, Praneet Kaur (P15) ................................................................................................ 98

Scheible, Kristin .................................................................................................................. 90

Schwaab, Thomas ............................................................................................................... 53, 131

Schell, Stephanie L. (P13, O) .............................................................................................. 56, 96

Schneider, Karin (S) ............................................................................................................ 38

Schwartzberg, Pamela L. ..................................................................................................... 47, 103

Segal, Brahm H. ................................................................................................................... 49, 87

Sentman, Charles L. ............................................................................................................. 115

Sharda, Anand P. (P48, O) ................................................................................................... 50, 53, 107, 131

Sharp, Stephen ..................................................................................................................... 66, 132

Shay, Ashley E. ................................................................................................................... 102

Sime, Patricia J. ................................................................................................................... 99

Singel, Kelly L. (P4, O) ....................................................................................................... 49, 87

Siracusa, Mark C. ................................................................................................................ 86

Skitzki, Joseph J. ................................................................................................................. 50, 107

Smith, Julianne N.P. ............................................................................................................ 65, 123

Smith, Norah L. ................................................................................................................... 40

Smyth, Tim .......................................................................................................................... 116

Solouki, Sabrina ................................................................................................................... 44, 89

Song, Jianxun ..................................................................................................................... 98

Soni, Chetna ........................................................................................................................ 56, 96

Soucy, Alicia (P12) .............................................................................................................. 95

Steiner, Donald (P35) .......................................................................................................... 118

Szczepanek, Steven M. ........................................................................................................ 119

Tewari, Prabhu .................................................................................................................... 66, 132

Thanavala, Yasmin .............................................................................................................. 99

Thatcher, Thomas H. ........................................................................................................... 99

Thees, Amy (P43) ................................................................................................................ 126

Thwe, Phyu (P21, O) ........................................................................................................... 64, 104

Topham, David .................................................................................................................... 90

Twum, Danielle Y.F. (P14) ................................................................................................. 97

Utley, Adam (P23, O) .......................................................................................................... 48, 58, 106

Vance, David J. .................................................................................................................... 124

Varrone, John J. ................................................................................................................... 122

Veazey, Janelle (P33) .......................................................................................................... 116

Veerapen, Natacha ............................................................................................................... 46, 100

Vella, Jennifer (P45) ............................................................................................................ 128

138.

Ventro, Daniela .................................................................................................................... 48

Vignali, Paolo D.A. ............................................................................................................. 70

Vomhof-Dekrey, Emilie ...................................................................................................... 46, 100

Wald, Alexander ................................................................................................................. 53, 131

Walrath, Travis (P49, O) ..................................................................................................... 66, 132

Wang, Jocelyn ..................................................................................................................... 40

Wang, Li .............................................................................................................................. 113

Watson, Neva ...................................................................................................................... 40

Webb, Lauren M. (P3) ......................................................................................................... 43, 86, 88

Westfall, Jennifer E. ............................................................................................................ 122

Williamson, David R. .......................................................................................................... 102

Wilmore, Joel R. (P38, O) ................................................................................................... 59, 121

Winslow, Gary ..................................................................................................................... 57, 60, 105, 127

Wissink, Erin ....................................................................................................................... 40

Wojno, Elia Tait ................................................................................................................... 43, 86, 88

Yang, Qi (S) ........................................................................................................................ 37

Yates, Jennifer (P17, O) ...................................................................................................... 46, 100

Yokoyama, Wayne M. (KS, WP) ........................................................................................ 34, 67

Yu, Hong ............................................................................................................................. 70

Zhang, Kangning ................................................................................................................. 37

139.

Amiel, Eyal


802-656-0522

[email protected]

Amobi, Adaobi


716-583-0875

[email protected]

Baranova, Danielle

Wadsworth Center

518-473-3947

[email protected]

Barbachano-Guerrero, Arturo


315-464-9490

[email protected]

Barlow, Margaret

University of Rochester

216-410-5140

[email protected]

Battaglia, Nicholas


716-471-6114

[email protected]

Bellville, Dawn


518-262-5365

[email protected]

Berwin, Brent

Dartmouth College

603-208-7446

[email protected]

Bhandari, Sadikshya


580-647-0228

[email protected]

Bhat, Tariq


716-602-9907

[email protected]

Blader, Ira


716-852-9580

[email protected]

Braciale, Thomas

University of Virginia

434-825-2415

[email protected]

Burke, Catherine


716-949-2802

[email protected]

Buskiewicz, Iwona


802-6568507

[email protected]

Bynoe, Margaret

Cornell University

607-253-4023

[email protected]

Chapman, Timothy


585-314-3935

[email protected]

Attendee Contact Information









140.

Cojohari, Olesea


315-944-5434

[email protected]

Consiglio, Camila Rosat


716-845-8167

[email protected]

Costello, Angelica


518-262-0922

[email protected]

Coupet, Tiffany

Dartmouth College

571-393-1357

[email protected]

DeVault, Victoria


978-895-3204

[email protected]

Dostie, Kristen


860-486-3648

[email protected]

Drake, James (Jim)


518-262-9337

[email protected]

Elliott, Michael


585-273-4793

[email protected]

Emo, Jason


585-273-1408

[email protected]

Enelow, Richard

Dartmouth College

802-356-9653

[email protected]

Ginsberg, Arielle

BD Biosciences

347-205-5493

[email protected]

Ginty, Fiona

GE Global Research

518-366-6465

[email protected]

Gondre-Lewis, Timothy

NIAID/NIH

240-627-3566

[email protected]

Green, Kathy

Dartmouth College

603-650-5056

[email protected]

Green, William

Dartmouth College

603-650-8607

[email protected]

Harton, Jonathan


518-262-4445

[email protected]





141.

Howell, Tyger


716-845-8231

[email protected]

Huang, Weishan

Cornell University

607-253-4014

[email protected]

Huang, Yina

Dartmouth Medical College

603-650-7545

[email protected]

Jiang, Chenyang


860-771-9028

[email protected]

Katkere, Bhuvana

Pennsylvania State University

518-339-8133

[email protected]

Kenderes, Kevin


607-341-1511

[email protected]

King, Christine


315-464-5465

[email protected]

Ku, Amy


661-618-0258

[email protected]

Lawrence, David


518-486-9154

[email protected]

Lord, Edith


585-749-8604

[email protected]

Lozito, Shannon


802-258-0064

[email protected]

Lynes, Michael


860-486-4350

[email protected]

MacNamara, Kate


518-262-0921

[email protected]

Maharaj, Shivana


716-380-0223

[email protected]

Mantis, Nicholas


518-473-7487

[email protected]

Metzger, Dennis


518-262-6750

[email protected]









142.

Minchenberg, Scott


516-524-5812

[email protected]

Moon, Tayler


315-796-7728

[email protected]

Moshkani, Safie


518-264-2588

[email protected]

Muhitch, Jason


716-845-4930

[email protected]

Murthy, Aditi


508-813-3363

[email protected]

Nagar, Abhinit


518-262-4447

[email protected]

Norris, Carol


860-486-3648

[email protected]

O’Connor, William


518-262-6548

[email protected]

Oyesola, Oyebola

Cornell Univesity

607-379-7745

[email protected]

Palker, Thomas

NIAID/NIH

301-828-7192

[email protected]

Palmer, Claire

Dartmouth College

802-730-2807

[email protected]

Papillion, Amber


337-580-1924

[email protected]

Pelletier, Marianne

Astra Zeneca

518-872-1582

[email protected]

Peppenelli, Megan


315-464-7682

[email protected]

Poon, Amanda

Wadsworth Center

347-868-8796

[email protected]

Post, Christina


781-974-9431

[email protected]






143.

Powers, Colin


443-889-6314

[email protected]

Price, B. Duane

NIAID/NIH

240-669-5074

[email protected]

Ravi, Swetha


313-788-2486

[email protected]

Richards, Angelene

Wadsworth Center

518-420-8281

[email protected]

Robek, Michael

Albany Medical Center

518-264-2580

[email protected]

Rudd, Brian

Cornell University

607-253-4418

[email protected]

Praneet Kaur Sandhu


716-418-4355

[email protected]

Schell, Stephanie


443-243-9751

[email protected]

Schneider, Karin


315-559-8953

[email protected]

Sharda, Anand


716-845-7180

[email protected]

Singel, Kelly


716-845-3138

[email protected]

Snyder, James

NIAID/NIH

240-669-5060

[email protected]

Soucy, Alicia


518-262-6220

[email protected]

Steiner, Donald


518-262-6220

[email protected]

Szczepanek, Steven


860-486-8101

[email protected]

Thanavala, Yasmin


716-845-8536

[email protected]






144.

Thees, Amy


732-232-4739

[email protected]

Thwe, Phyu


414-241-3367

[email protected]

Twum, Danielle


716-845-3352

[email protected]

Utley, Adam


336-847-4725

[email protected]

Veazey, Janelle


585-519-8472

[email protected]

Vella, Jennifer

Dartmouth College

603-650-7546

[email protected]

Walrath, Travis


518-262-6341

[email protected]

Webb, Lauren

Cornell University

617-763-7499

[email protected]

Wilmore, Joel

University of Pennsylvania

315-882-2959

[email protected]

Winslow, Gary


315-464-7658

[email protected]

Wohlfert, Beth


716-829-3969

[email protected]

Yang, Qi


518-264-2582

[email protected]

Yates, Jennifer

Wadsworth Center

518-339-2556

[email protected]

Yokoyama, Wayne

Washington University at St. Louis

314-229-8797

[email protected]




19th annual upstate new york immunology conference · keynote speakers are dr. wayne m. yokoyama...

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