bispecific antibody-armed nanoparticles for...
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Bispecific antibody-armed nanoparticles for future theranostic applications
John T.-A. Hsu
National Health Research Institutes
Acknowledgements
Yun-Ming Wang (王雲銘)
Shou-Cheng Wu (吳首成)
Yu-Jen Chen (陳俞任)
Hsiang-Ching Wang (王翔靖)
Min-Yuan Chou (周民元)
John Tsu-An Hsu (徐祖安)
Teng-Yuan Chang (張騰元)
Shyng-Shiou Yuan (袁行修)
Chiao-Yun Chen (陳巧雲)
Ming-Feng Hou (侯明鋒)
高雄醫學大學附設中和紀念醫院Kaoshiung Medical University
Chung-Ho Memorial Hospital
http://e021.life.nctu.edu.tw/~ymwang/first.html
Yun-Ming Wang (王雲銘)Research Expertise:
• Bioinorganic Chemistry
• Paramagnetic metal complex and bio-activated metal complex for magnetic resonance imaging(MRI)
• Superparamagnetic iron oxide nanoparticle (SPIO) for MRI
• Probe for Optical Imaging
• Molecular Imaging
The manuscript was accepted by Theranostics. (2015/09/22)
Bispecific Antibody Conjugated Manganese-Based Magnetic Engineered Iron Oxide for Imaging of HER2/neu- and EGFR-
Expressing Tumors
History of NHRI
The founding of the National Health ResearchInstitutes (NHRI) was first proposed in 1988 bymembers of Academia Sinica.
In 1994 a preparatory committee was formed.
NHRI was officially established in January 1996.
NHRI relocated to its permanent campus in Zhunanin 2004.
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Intramural Research Units
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Cancer Research
Cellular and System Medicine
Population Health Sciences
Biotechnology & Pharmaceutical Research
Divisions/Center
Environmental Health & Occupational Medicine
Molecular & Genomic
Immunology Research
Infectious Disease & Vaccinology
Institutes
National EnvironmentalHealth Research
Neuropsychiatric Research
Biomedical Engineering & Nanomedicine
PUBLICATIONS from PIs at NHRI:Please refer to National Health Research Institutes Institutional Repositoryhttp://ir.nhri.org.tw/handle/3990099045/17?locale=en-US
NHRI
Taipei
50 Km
Taiwan: Ilha Formosa ~ a Beautiful Island
NHRI
Please visit us at www.nhri.org.tw
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Considerations of
Targets
Assay Developments
Drug Screening
Active HitsLead
optimizationPatent issues
Lead compounds
Patent issues
Pharmacology &
Animal Models
Pharmaco-kinetics
Development Candidate
候選發展藥物
Pre-formulation
Pharmaceutical
studies
DMPK (Matabolism
& PK)
Safety Pharmacology
/Toxicity
原料藥/製劑GMP生產
臨床試驗規劃
IND
Clinical Trial
NHRI Intramural capability for new drug discovery
Integrated Drug Discovery Team at DBPR
Biology
Chemistry
High Throughput Screening
Pharmacokinetics/Drug Metabolism
Animal Pharmacology
Cancer
Infectious Disease
Medicinal Chemistry
Molecular ModelingVirtual Screening
Joe
C. S
hih
, D
irec
tor
Metabolic Disease
TA Hsu
C.C. Kuo
Y Yueh
SJ Lee
MS Hung
HP HsiehJH Chern
WT JiaangKS ShiaJC Lee
K. Chou
Joe C. Shih
CT Chen
SY Wu
TK Yeh
Core Technologies of IBPR
IBPR/NHRI
PharmacokineticsDrug Metabolism
Highthroughputscreening
Chemistry
Biology
Animal pharmacology
Pharm. Development
Project ManagementPreclinical/Clinical Development
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My Experiences• BS, Ch.E. NTU 1989
• Army Service, Second Lieutenant
• PhD, Ch.E., Johns Hopkins University 1991-1995
– Expression of IgG with chaperone and foldase
• Post-Doc at Human Genome Sciences, Inc., 1996 - 1997
– Learned many fields ranging from HTS cDNA sequencing, protein expression and purification, protein chemistry
• Merck & Co., 1997 - 1998
– Biopharmaceutics, Live virus vaccine production and formulation
• Nation Health Research Institutes (1998-2015; now 1,400 employees)
– Established drug discovery and development group
• ~ 10 patents, > 100 papers
– Management Service (2013 - 2015) – in charge of General Affairs, Human Resources, Finance/Accounting, Computing and Information, Procurement, etc..
Jeremy Lin, an NBA player (Hornet)
Experiences at Merck
• To improve Varivax
– Varivax is a live virus vaccine for chickenpox
(Chickenpox = 水痘 = Ветряная оспа)
– To improve the storage conditions for Varivax
• Membrane stability, virus stability, cryobiology, …
Experiences at Merck
• To improve Varivax
– Varivax is a live virus vaccine for chickenpox
(Chickenpox = 水痘 = Ветряная оспа)
– To improve the storage conditions for Varivax
• Membrane stability, virus stability, cryobiology, …
– To transform Varivax into Zostavax for shingles
• Zostavax was approved in 2005
(Shingle (herpes zoster) = 皮蛇 = опоясывающий лишай)
Shingles/Zostavax
Company Merck & Co. Inc.
DescriptionLive attenuated zoster virus vaccine
Standard Indication Varicella zoster virus (VZV)
Indication Details
Prevent shingles (herpes zoster) in adults 60 years or older
Vaccinate against shingles (herpes zoster) in adults ages 50-59
Experiences at Merck
• To improve Varivax– Varivax is a live virus vaccine for chickenpox
(Chickenpox = 水痘 = Ветряная оспа)– To improve the storage conditions for Varivax
• Membrane stability, virus stability, cryobiology, …
– To transform Varivax into Zostavax for shingles• Zostavax was approved in 2005(Shingle (herpes zoster) = 皮蛇 = опоясывающий лишай)
– To develop a tratavalent vaccine for• Chickenpox, Measles, Mumps, and Rubella• ProQuod: combining Varivax with MMR, approved in 2005
Chickenpox/ProQuod
Company Merck & Co. Inc.
DescriptionLive vaccine against measles, mumps, rubella and varicella viruses
Indication Details Prevent measles, mumps, rubella and varicella infection
Generation of Virus-Like Particles (VLP) of Enterovirus 71 (EV71)
Two recombinant baculoviruseswere constructed to encode P1and 3CD of enterovirus 71 (EV71),respectively. The expressed 3CDsuccessfully cleaved P1 in vitro andin vivo.
Biotechnology Letters, June 2003, Volume 25, Issue 12, pp 919-925In collaboration with Prof. YC Hu
P1 capsid protein
EV71
P1 & 3CD coding region
P1
P1 & 3CD recombinant baculovirus
Virus Production
Purification
P1 molecule VP1, VP2, VP3proteins
3C
RotavirusesInfluenza viruses
The Virus Images
Polio viruses
Recombinant Proteins Core
Value
Speed
Quality
• Protein engineering/expression and purification
• To Support drug discovery programs
Mission of IBPR
To establish, conduct and support research on new medicines and biotechnology that will lead to better human health and quality of life.
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Top 10 Drugs by Worldwide Sales 2003
Treats: Cholesterol$ 10.3 billion
Lipitor Zyprexa
Treats: Antipsychotic$ 4.8 billion
Zocor
Treats: Cholesterol$ 6.1 billion
Norvasc
Treats: Hypertension$ 4.5 billion
Prevacid
Treats: Stomach acid$ 4.0 billion
Procrit/Eprex
Treats: Anemia$ 4.0 billion
Nexium
Treats: Stomach acid$ 3.8 billion
PIavix
Treats: Cardiovascular Disease$ 3.7 billion
Zoloft
Treats: Antidepressant$ 3.4 billion
Latus Solostar
Treats: Asthma$ 3.7 billion
As traZenec aAbbott & Takeda GlaxoKlineSmith PfizerBristol Myers
Johnson & Johnson PfizerEli LillyMerck
Pfizer
Top 10 Drugs by Worldwide Sales 2014
Treats: Atypical antipsychotic$ 7.2 billion
Aripiprazole Esomeprazole
Treats: Proton pump inhibitor$ 6.3 billion
Adalimumaba
Treats: Rheumatoid arthritis$ 6.3 billion
Rosuvastatin
Treats: Cholesterol$ 5.6 billion
Fluticasonepropionate and salmeterol
Treats: asthma, allergic rhinitis nasal polyps$ 5.0 billion
Etanerceptb
Treats: Rheumatoid arthritis$ 5.0 billion
Sofosbuvir
Treats: HCV$ 4.4 billion
Infliximaba
Treats: Crohn's disease; rheumatoid arthritis$ 4.3 billion
Pegfilgrastime
Treats: Neutropenia$ 3.6 billion
Latus Solostar
Treats: diabetes$ 3.8 billion
Sovaldi
Advair Diskus
insulin glargine
Top 10 Drugs by Worldwide Sales 2014
Treats: Atypical antipsychotic$ 7.2 billion
Aripiprazole Esomeprazole
Treats: Proton pump inhibitor$ 6.3 billion
Adalimumaba
Treats: Rheumatoid arthritis$ 6.3 billion
Rosuvastatin
Treats: Cholesterol$ 5.6 billion
Fluticasonepropionate and salmeterol
Treats: asthma, allergic rhinitis nasal polyps$ 5.0 billion
Etanerceptb
Treats: Rheumatoid arthritis$ 5.0 billion
Sofosbuvir
Treats: HCV$ 4.4 billion
Infliximaba
Treats: Crohn's disease; rheumatoid arthritis$ 4.3 billion
Pegfilgrastime
Treats: Neutropenia$ 3.6 billion
Latus Solostar
Treats: diabetes$ 3.8 billion
Sovaldi
Advair Diskus
insulin glargine
Anti-TNF Protein Drugs
Bispecific antibody-armed nanoparticles for future theranostic applications
Antibody structure and types of antibodies
Nat Biotechnol. 2005 May;23(5):556-7.
Nat Rev Cancer. 2015 Jun;15(6):361-70.
Monoclonal antibody-based cancer therapeutic strategies
Sci Rep. 2014 Oct 24;4:6760
Traditional antibodies and antibody fragments vs nanobodies
Next-generation antibodies
1. Antibody–drug conjugates (ADCs)2. Engineered antibodies3. Multispecific antibodies
Nat Rev Drug Discov. 2014 Jun;13(6):413-4.
MAbs. 2014 Jan-Feb;6(1):34-45.
Bispecific antibody development timeline
2011 Kontermann RE. Bispecific Antibodies: Developments and Current Perspectives.
Structure of Catumaxomab (RemovabTM), the first bispecific antibody to achieve approval in EU
Nat Biotechnol. 2010 Sep;28(9):917-24.
Catumaxomab was approved in the european Union for the intraperitoneal treatment of patients with malignant ascites. (2009)
Mechanism of action of the novel bispecific antibody Blinatumomab (BlincytoTM)
Clin Cancer Investig J 2014;3:577-8
US FDA Approves Blinatumomab for B-Cell ALL (2014)
Antibody Fragment Configurations
https://fusionantibodies.com/services/antibody-engineering/antibody-fragments/
http://www.nist.gov/director/vcat/upload/June-2015-VCAT_Koenig_post.pdf
https://www.antibodysociety.org/news/approved_mabs.php
Schematic diagram of tetravalent bispecific antibody (Bis-Ab)
Background
• A highly specific contrast agent system comprising an iron oxide nanoparticle core coated with poly(ethylene glycol) was previously developed (MnMEIO).
• The tetravalent bispecific antibody may facilitate detection of HER2/neu- and EGFR-expressing tumors.
The manuscript was accepted by Theranostics. (2015/09/22)
Bispecific Antibody Conjugated Manganese-Based Magnetic Engineered Iron Oxide for Imaging of HER2/neu- and EGFR-
Expressing Tumors
TEM images and schematic representation of MnMEIONPs conjugated with or without mPEG, CyTE777 and Bis-Ab
Binding specificity of Ab-armed NPs to HER2/neu- and EGFR-expressing tumor cells
KD (EGFRv3): ~10 nMKD (HER2/neu): ~3 nM
Histological analyses of Colo-205 and SKBR-3 tumor tissue specimens acquired 96 hrs after injection of Ab-armed NPs
Sections were stained with Prussian blue.
Prussian blue staining of SKBR-3, A431, and Colo-205 cells treated with Ab-armed NPs
In vitro optical imaging of Ab-armed NPs treated SKBR-3,
A431, and Colo-205 cells
Blocking study of Bis Ab-armed NPs with tumor cells at 37 °C
The bivalent binding modes of a Bis-Ab
Curr Opin Chem Biol. 2013 Jun;17(3):400-5.
The in vitro T2-weighted images of SKBR-3, A431, and Colo-205 tumor cells after treatment with Ab-armed NPs
Pharmacokinetics of MnMEIO-CyTE777-(Bis)-mPEG and control NPs (MnMEIO-CyTE777-mPEG NPs) in tumor-
bearing mice
In vivo optical images of tumor-bearing mice after intravenous injection of NPs w/wo Bis-Ab
NPs (10 mg/kg)
T2-weighted MR images (7.0 T) of tumor-bearing mice before (Pre), and 2 hrs and 24 hrs after injection of Ab-
armed NPs
Conclusions
• Using a bispecific Ab for nanoparticle conjugation not onlysimplifies conjugation process but also overcome the diverseexpression patterns of individual receptors.
• Multifunctional nanoparticles (MnMEIO-CyTE777-(Bis)-mPEGNPs) can be applied to optical imaging, dual targeting effectand MRI contrast agent for early diagnoses of HER2/neu- orEGFR-overexpressing cancers.
• Bispecific antibody-armed nanoparticles can be applied tofuture theranostic applications.
Acknowldgements
National Health Research Institutes
Спасибоspasibo
Kan-Xie