presented by z . jafari msc in medical biotechnology of ... · • pet imaging uses...

Presented by Z . Jafari

MSc in Medical Biotechnology of QUMS

Supervisor : Dr. Ahmad pour

05/23/2017

1

Overview Introductions

Methods and techniques

Discussion

References2

Contents Introduction of cellular imaging modalities……..……………………………………..…………………….....…...6

The principle of cell Tracking with MRI…......……9

Cancer tracking and MRI Direct cell labeling techniques.............................11

Indirect cell labeling techniques……………………..12

Limitation of indirect cell labeling techniques....16

Encapsulation cell labeling techniques..…………..18

Statistic …………………………………………….…………..19

Articles………………………………………………...…...….21

References ……………..………………………….………..54

3

Cancer tracking

contentsIntroduction…….…………………………………………….……………….27

The principal of Pet CT imaging…….………………….....…………..28

Radiotracer of Pet CT……………………………….……………….…....29

Mechanism of radiotracer like FDG..……………......................30

Which cancer we can use FDG? ……………………………….….….32

30-deoxy-30-18F-fluorothymidine (FLT)……….……….………..33

FMAU…………………………………………………….……………….…....34

PET radiotracers have been developed to image hypoxia…....35

Cancer tracking with PET CT Imaging Apoptosis…………………………………….…………...………….……...37

The second class of imaging agents targets caspases…………..………...38

A third class of imaging agents……………………………….….……..39

References ……………………………………………………………….….…56

4Cancer tracking

Introduction………..………………………………………………………………….43

Cancer tracking and immunology Cytotoxic T lymphocytes and cell labeling method…..…………….…...45

And bioluminescence and optical Tracking T cells by optical fluorescence and bioluminescence imaging

Fleurcence imaging direct labeling methods…………………………………………………………….46

Indirect labeling methods…………………………………………………….……47

Articles……………………………………………………………………………………51

references…………………………………………………………………………….…57

5

Contents:

6

cellular Imaging modalities

introduction

Cancer tracking

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5297829/figure/ijms-18-00198-f002/

7

discussion

in vivo

8

The principle of cell Tracking with MRI

The indirect cell labeling techniques

introduction

Cancer tracking

9

The direct cell labeling techniques

Cells(for cancer issues dendritic cells , T cells

cancer immunotherapy and cancer properties MSCs ) are incubated with an MRI contrast

agent in vitro and prior to transplantation , with a

transfection agent such as PLL or lipofectamine

Cancer tracking

10


Endogen exogen

Entrance of CA PinocytosisPhagocytosisWith sonicationelectroporation

CA ( MNP , paramagnetic , Gd , SPIONs superparamagnetic iron oxidate np)to macrophage

The way of labeling Targeting the cell surface receptors with antibody conjugated MRI CA ortargeting phagocytic nature of endogenous cell population

Like endogen

limitation Yes for the difficulty of isolating the neural stem cell because they are sub ventricular zone

It can’t distinguish between live and death cells 1:the differences T1and T2 of CA in live and death cells 2:effect of PH which usually with death cells

11


Cancer tracking


12


Cells transfected with transfection agents or transduced with viral vectors to express an MRI reporter gene.

A Reporter gene is a gene that can fused to a gene of interest or cloned after that expression of it can create a peptide, protein Nano structure receptor or enzymatic activation to generate an MRI contrast

Cancer tracking

13

Cancer tracking


14


Advantage of the

indirect technique

s

Use genetically engineering

Engineered cells proliferate to

generate daughter cells

MRI signal is not diluted

Live labeled cells distinguished

from dead cells

The reporter gene is expressed only in live cells and switched off in

dead cells

Cancer tracking

15

Examples of MRI

reporter gene

systems

Iron _binding receptor transferrin (that require T2/T2* MRI contrast agent )

The iron storage protein ferritin (that require T2/T2* MRI contrast agent )

The_ βgalactosidase which hydrolysis of β _D _ galactosidases(the require T1 MRI contrast agent like Gd )

Reporter genes which use CEST contrast generation mechanism such as :LRP(lysine rich protein)or HSV1


Cancer tracking

16

-use of imaging agents with unfavorable pharmacokinetic profile lead to a delayed MRI signal and then result in false negative low sensitivity of reporter genes by biological function lead to low detection_developed reporter gen in bacterial origins non human origins _usually reporter genes are immunogenic

Limitation of indirect cell labeling techniques:

Cancer tracking

17

The encapsulation cell labeling Technique

_Use of biomaterials : such as alginate capsules to protect therapeutic cells from immune cells and permit the diffusion of small molecular weight such as water and nutrients_ polycations such as :PLL(poly _L _ lysine)to control pore sizes-Therapeutic cells and MRI agents are encapsulated together in vitro prior to transplantation

Cancer tracking

18

The encapsulation cell labeling Technique


Statistic in cancer tracking between 2000-2017 years

19

2000-2004 2004-2008 2008-2012 2012-2016 2016-2017

تعداد مقاالت 44300 66900 106000 59700 34300

44300

66900

106000

59700

34300

0

20000

40000

60000

80000

100000

120000

140000

Ar

tic

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years

Cancer tracking

Cancer tracking with mri

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2000-2004 2004-2008 2008-2012 2012-2016 2016-2017

Series1 5270 13500 18100 17900 9590

5270

13500

18100 17900

9590

0

5000

10000

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20000

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Art

icle

s

years

cancer tracking with mri

Cancer tracking

21

Discussion

in vivo

https://www.ncbi.nlm.nih.gov/pubmed/25625023

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invitro

Discussion


23

discussion

invitro


24

discussion

in vivo


25

Cancer tracking with pet CT

Cancer tracking

26


27

PET- CT(positron emission computed tomography)

• Pet CT is a type of nuclear medicine imaging• Nuclear medicine is a branch of medical imaging

that uses small amounts of radioactive material to detect severity or treat diseases include many type of cancers heart diseases , neurological disorder endocrine and gastrointestinal • Because nuclear medicine procedure able pinpoint

molecular activity within the body

Cancer tracking

https://image.slidesharecdn.com/medicalimaging-130804074029-phpapp01/95/medical-imaging-overview-21-638.jpg?cb=1375602411

introduction

28

the principals of pet CT imaging

• PET imaging uses radiopharmaceuticals (or radiotracers like DG( deoxy glucose )and … labeled with positron emitting radioisotopes such as 11C, 13N, 15O, and 18F,which are produced in a cyclotron which are produced in a radioisotope generator.

• the positron annihilates and generates 2 annihilation photons (each with an energy of 511 k eV),which travel in opposite directions.

• PET scanners are equipped with coincidence electronics to detect these pairs of photons as they hit opposing detectors

Cancer tracking

30

Radiotracer of PET CT

Cancer tracking


31

the mechanism of radiotracers like FDG

• Radiotracers in PET/CT allows to quantify the metabolic activity of a tumor (like glycolysis) and become a reference tool in oncology for the tissue staging ,radiotherapy planning and monitoring response in many cancers.

• In 18 F- fluorodeoxyglucose (18F-FDG) is transported into cells in parallel with glucose and is phosphorylated with hexokinase to F_deoxyglycose-6-phosphate because lack of a hydroxyl group at the 2 position , it’s prevented from being a substrate of enzymes farther down the glycolytic pathway

• FDG_6_phosphate trapped within the cancer cells• Because of isotope like F we can detect them with PET CT

Cancer tracking

31

FDG uptake mechanism 32

https://www.researchgate.net/profile/Artem_Lebedev/publication/259010016/figure/fig2/AS:267481113690123@1440783875152/Figure-10-10-Proposed-mechanism-for-the-cellular-accumulation-of-18FFDG.png

32

which cancer we can use FDG?

FDG-PET include lymphoma, head and neck cancer,lung, colorectal cancer, breast cancer, esophageal cancer, melanoma, cervical cancer, thyroid cancer, and pancreatic cancer are “hot”

prostate cancer, neuroendocrine cancer,and well differentiated hepatocellular carcinoma, are often “cold” which limits the utility of FDG-PET

Cancer tracking

33

30-deoxy-30-18F-fluorothymidine (FLT)

• It uses for imaging cell proliferation

• Initial efforts focused on synthesizing analogs of thymidine, because thymidine is used byproliferating cells for DNA synthesis during the S-phaseof the cell cycle but, unlike other nucleosides, is not incorporated into RNA

• After injection, FLT enters the cell by nucleoside transporters and is trapped in the cytosol through phosphorylation

• Because FLT lacks a hydroxyl group at the 3′position, itis not incorporated into DNA , and its accumulationserves as a measure of cellular proliferation

Cancer tracking

34

The thymidine analog (1-[20-deoxy-20-18Ffluoro-beta-D_arabinofuranosyl]thymine) (FMAU)

• proliferation imaging probes• for studying bone metastases and genitourinary

malignancies

Cancer tracking

35

PET radiotracers have been developed to image hypoxia

2-nitroimidazoles, such as:

• 18F_fluoromisonidazole (FMISO)(research)

• 18F-fluoroetanidazole (FETA)

In solid tumors, hypoxia is associated with restrained proliferation, differentiation, apoptosis, and necrosis

nucleoside conjugates, such as

• 18F-fluoroazomycin arabinoside (FAZA)

• 64Cu-diacetyl-bis(N4-methylthiosemicarbazone) (Cu-ATSM)

Cancer tracking

36 PET radiotracers have been developed to image hypoxia

All of these agents undergo intracellular trapping at a rate

inversely proportional to intracellular oxygen concentration

Cancer tracking

Imaging Apoptosis

the detection of apoptosis can potentially be used to provide anearly indication of the success of therapy(in radiotherapy)

phosphatidylserine residues that normally reside on the intracellularmembrane surface but that are translocated to the extracellular surface during apoptosis

99mTc-annexin V is able to image apoptosis in vivo and topredict patient outcome after chemotherapy or radiationtherapy.

a radiolabeled protein with affinity to phosphatidylserine

37

Cancer tracking

The second class of imaging agents targets caspases

18F-ICMT-11 (a caspase-3- specific small molecule PET tracer based on the caspase inhibitor statin)

18F-CP18 (a pentapeptide-based PET tracer that is a substrate of caspase-3)

38

Cancer tracking

A third class of imaging agents

It detect plasma membrane depolarization

the most notable of these imaging agents is the PET tracer 18F-ML-10

39

Cancer tracking

cancer tracking with PET CT

40

2000-2004 2004-2008 2008-2012 2012-2016 2016-2017

Series1 2230 6230 12300 16300 3850

2230

6230

12300

16300

3850

-5000

0

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cancer tracking with PET CT

Cancer tracking

41

discussion

In vivo


Immunology , bioluminescence and

optical fluorescence imaging

43

Cancer tracking with immunology

The imaging modalities applied for T cell tracking in both preclinical and clinical studies include:

optical fluorescence/bioluminescenceimaging , computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET)

Cancer tracking

44

Cytotoxic T lymphocytes and cell labeling methods

However, in immunotherapy, cytotoxic T cells can be manipulated to recognize tumor-specific antigens When infused into a patient, the engineered T cells actively

attack and destroy the tumors displaying these antigens

There are two major principles in labeling the T cells for in vivo cell tracking:

direct and indirect labeling

Cancer tracking

45

Cytotoxic T lymphocytes and cell labeling methods

Direct labeling

generally requires isolation and ex vivoexpansion of the T cells from the subject, followed bylabeling with a proper imaging probe in vitro and injection of these cells into the subject

indirect labeling involves genetic engineering of the T cells by transfecting

them with a reporter gene that encodes an enzyme or transporter, which can utilize the designated imaging probe as a substrate and allow for visualization and tracking of these T cells over time

Cancer tracking

46

Tracking T cells by optical fluorescence and bioluminescence imaging

In optical fluorescence imaging, T cells are labeled by fluorophores, fluorescent proteins, or quantum dots

The fluorophores are usually near infrared (NIR) fluorescent dyes, such as indocyanine green

labeled cells could be detected by flow cytometry

the labeled CTLs and the tumor cells could be detected by confocal intravital microscopy

The optical fluorescence imaging with fluorophores usually adopts a direct labeling strategy, while imaging with fluorescent proteinsinvolves an indirect method

Quantum dots (QDs) are a class of semiconductor nanocrystals (2-6 nm in size) that have broad excitation spectra

Direct labeling methods

47


Indirect labeling methods

Bioluminescence optical imaging is in principle distinct from the fluorescence imaging and involves an indirect labeling strategy.

Indirect labeling by fluorescent proteins. Fluorescent proteins, such as green fluorescent protein (GFP)

and red fluorescent protein (RFP), are also used to tag the T cells

A DNA construct carrying a coding sequence for GFP or RFPis introduced into the cell

It has become a powerful means to probe the mechanisms for biological processes in vitro and in preclinical studies.

Cancer tracking

48


Indirect labeling methods

Indirect labeling by bioluminescent agents:

a luciferase enzyme is expressed in the cell; when its substrate luciferin is introduced, luciferase can catalyze the oxidation of luciferin in the presence of ATP and oxygen

The reaction emits photons and only living cells can produce signals

produced a transgenic bioluminescence mouse model from which they isolated the T cells that constantly expressed luciferase

The BLI results clearly showed these adoptively transferred T cells homed to the antigen-positive tumors.

Cancer tracking

cancer tracking with immunology

49

2000-2004 2004-2008 2008-2012 2012-2016 2016-2017

Series1 5670 10600 15000 17700 6270

5670

10600

15000

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6270

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cancer tracking with immunology

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Bioluminescence and optical fluorescence imaging

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Discussion

In vivo

52

Discussion

In vivo In vitro

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4781579/

55

53

discussion

in vivo

https://www.ncbi.nlm.nih.gov/pubmed/25157278 43

referencesMri references

1.Ethel J. Ngen and Dmitri Artemov. Advances in Monitoring Cell-Based Therapies with Magnetic Resonance Imaging: Future Perspectives.received: 26 October 2016; Accepted: 10 January 2017; Published: 19 January 2017

2. Yusri D Heryanto1, Arifudin Achmad1,2,3, Ayako Taketomi-Takahashi1, Yoshito Tsushima Department of Diagnostic Radiology and Nuclear Medicine, Gunma University Graduate School of Medicine,Maebashi, Gunma, Japan; Human Research Cultivation Center, Gunma University, Kiryu, Gunma, Japan; Department of Radiology, Faculty of Medicine, Gadjah Mada University, Yogyakarta, Indonesia . In vivo molecular imaging of cancer stem cells Received August 27, 2014; Accepted September 5, 2014; Epub December 15, 2014; Published January 1, 2015

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Mri references

3. Daniel Spira, MD, Fritz Schick, MD, PhD, Ru¨ diger Bantleon, PhD, Hartwig Wolburg, PhD and Benjamin Wiesinger, MD, Gerd G , Department of Diagnostic and Interventional Radiology, Universita¨tsklinikum Heidelberg, Im Neuenheimer Feld 110, 69120 Heidelberg Department of Diagnostic and Interventional Radiology, Eberhard- Karls-University Tu¨bingen, Hoppe- Seyler -Str. 3, 72076 Tu¨bingen, Germany Institute of Pathology and Neuropathology, Eberhard- Karls-University . Labeling Human Melanoma Cells With SPIO: In Vitro Observations. Submitted: 29/11/2014. Revised: 16/11/2015. Accepted: 20/11/2015.

4. Christian Weis, Fabian Blank,Adrian West,Gregory lack,RobertWoodward,MatthewR.J.Carroll,Astrid Mainka,Ren_e artmann,Andreas Brandl,Heiko Bruns,Elizabeth Hallam,Jeremy Shaw,John Murphy,Wey Yang Katerina E. Aifantis,Rose Amal,Mike House,Tim St. Pierre,and Ben Fabry . Labeling of cancer cells with magnetic nanoparticles for magnetic resonance imaging. Published online 28 June 2013 in Wiley Online library(wileyonlinelibrary.com).

5. Paolo E. Porporato Pierre Danhier, Géraldine De Preter, Julie Magat, Quentin Godechal Pierre Sonveaux, Bénédicte F. Jordan, Olivier Feron and Bernard Gallez. Multimodal cell tracking of a spontaneous metastasis model: comparison between MRI , electron paramagnetic resonance and Bioluminescence received: 2 January 2013,Revised: 3 May 2013,Accepted: 3 June 2013,Published

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Pet CT/MRI References

6. Kim JS1, Kim YH, Kim JH, Kang KW, Tae EL, Youn H, Kim D, Kim SK, Kwon JT, Cho MH, Lee YS, Jeong JM, Chung JK, Lee DS. Cancer. Development and in vivo imaging of a PET/MRI Nano probe with enhanced NIR fluorescence by dye encapsulation. 2014 Jun 19

7. Haubner R1, Maschauer S2, Prante O2PET radiopharmaceuticals for imaging integrin expression: tracers in clinical studies and recent developments. Epub 2014 Jun 11

8. Danhier P1, De Preter G, Magat J, Godechal Q, Porporato PE, Jordan BF, Feron O, Sonveaux P, Gallez B. Multimodal cell tracking of a spontaneous metastasis model: comparison between MRI, electron paramagnetic resonance and bioluminescence. 2014 Mar-Apr

9. Farwell MD1, Pryma DA, Mankoff DA. PET/CT imaging in cancer: current applications and future directions. Cancer. 2014 Nov

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Immunology and bioluminescence and optical imaging references

10. Zhiyi Liu and Zheng Li. Molecular Imaging in Tracking Tumor-Specific Cytotoxic T Lymphocytes (CTLs) . Theranostics 2014, Vol. 4, Issue 10

11.Judith M. Runnels, Alicia L. Carlson, Costas Pitsillides, Brian Thompson, Juwell Wu, Joel A. Spencer, John M. J. Kohler,Abdel Kareem Azab, Anne Sophie Moreau, Scott J. Rodig, Andrew L. Kung, Kenneth C. Anderson,Irene M. Ghobrial, and Charles P.Lin,Optical techniques for tracking multiple myeloma engraftment, growth, and response to therapy published online 2011 Jan 11. doi:10.1117/1.3520571

12. Aaron J. Clark Shayan Fakurnejad, Quanhong Ma, Rintaro Hashizume, Bioluminescence Imaging of an Immunocompetent Animal Model for Glioblastoma,jovevideo.com, 1/15/2016

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Thanks for your attention

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presented by z . jafari msc in medical biotechnology of ... · • pet imaging uses...

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