PhD Scientific Days 2020

Clinical Medicine II. Posters

Culture Conditions Determine the Long Term Persistence and Phenotype of HER2 Specific CAR T Cells

Előadó neve

Csaplár, Marianna, MSc

Előadó munkahelye

UD Faculty of Medicine Department of Biophysics and Cell Biology

Előadó telefonszáma

+36308707398

Előadó e-mail címe

csaplarmarianna@gmail.com

Az előadás címe

Culture Conditions Determine the Long Term Persistence and Phenotype of HER2 Specific CAR T Cells

Szerző(k) neve és munkahelye

Marianna Csaplár1
1 UD Faculty of Medicine Department of Biophysics and Cell Biology, Debrecen

Szekció

Clinical Medicine II. Posters

Language of the presentation

English

Section, first choice

Clinical Medicine

Section, second choice

Health Sciences

Összefoglaló szövege

Introduction
Genetically modifying T cells to express synthetic chimeric antigen receptors (CARs) is one of the most promising strategies for the immunotheraphy of cancer. CARs constructed from a monoclonal antibody and effector domains of the T cell receptor complex allow T cells to recognise and kill tumor cells in a HLA unrestricted manner. The proliferative ability and the phenotype define the qualities of the produced CAR T cell population, which are largely determined by the method of T cell activation and expansion.
Aims
With a view to optimize the activation and expansion of CAR T cells produced in our lab, we aimed to explore and compare the persistence and phenotype of second generation HER2 specific CAR T cells using a conventional and a novel medium, as well as a conventional and a novel stimulation method.
Method
We used HER2 targeting CAR constructs containing the CD3z effector, and one of the CD28 or 41BB costimulatory domains. The constructs were retrovirally transduced into human T cells. Mock-transduced cells were used as negative controls. We cultured T cells from two donors in RPMI or Medium X, combined with either of two stimulation methods: anti CD3 + anti CD28 or anti CD3 + Reagent Y. For assessing in vitro persistence, a re-challenge assay was implemented, where CAR T cells were removed from the HER2-FC target antigen after an optimized killing time period (3 days), counted in flow cytometry and added to new targets to measure proliferation again, continued until anergy was reached. The phenotype of the CAR T cells was determined by immunostaining using antiCD4–A488 and antiCD8-A647 to measure cytotoxic/helper ratio, and antiCCR7-FITC with antiCD45RA-APC to determine naive/effector differentiation by flow cytometry.
Results
Among the applied methods the antiCD3 + antiCD28 stimulation combined with Medium X proved to be the most effective in terms of in vitro endurance. Anti-CD3 + Reagent Y significantly raised the ratio of CD8+ cytotoxic CAR T cells combined with both media, but lowered proliferation rates. The percentage of the naive phenotype showed no significant difference in either of the applied methods.
Conclusion
Among the tested methods for CAR T cell activation and expansion Medium X yielded promising results, thus we are going to test it further, adding more donors to the experiment.

Additional Information

György Vereb
vereb@med.unideb.hu
Árpád Szöőr
akuka@med.unideb.hu
EFOP-3.6.3-VEKOP-16-2017-00009

Bemutatás módja

Poszter

Kind

Szabad

Status

elfogadva

Accepted presentation method

poszter

Előadás fájl jóváhagyás

nem rendelkezett róla

Előadó

4667

Start

12:53

End

12:56

Authors (legacy)

Marianna Csaplár1
1 UD Faculty of Medicine Department of Biophysics and Cell Biology, Debrecen