IN VIVO CD19 CAR-T CELL KINETICS IN MULTIPLE ORGANS USING A POINT OF CARE APPROACH
Details
- Presentation type: MS - Master's student
- Track: Cellular Biology
- Keywords: CAR-T cell; adoptive cell transfer; point-of-care; acute lymphoblastic leukemia;
- 1 Programa de Imunologia e biologia tumoral, Instituto Nacional do Cancer (INCA), Rio de Janeiro
- 2 INSTITUTO NACIONAL DE CÂNCER (INCA)
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Log in Abstract
INTRODUCTION AND OBJECTIVES: Chimeric antigen receptor (CAR) – T cell therapy is a promising immunotherapy approach for cancer treatment with outstanding results since its approval in 2017. However, the high associated cost limits its broad application. Our group has recently developed a point-of-care (POC) approach, that enables the rapid delivery of CAR-T cells without previous activation and ex vivo expansion, which could greatly reduce the costs of production. This project aims to evaluate the in vivo kinetics of CAR-T cells in the POC and expansion systems. MATERIAL AND METHODS: NOD-scid IL2R gamma null (NSG) mice were engrafted with Nalm-6 GFP+ luciferase+ cell line and treated with CAR-T cells 24 h after electroporation (POC group), after 8 days expansion, with mock electroporated cells or with PBS. Tumor burden was determined by bioluminescence imaging. At pre-determined timepoints, mice were euthanized and samples from blood, bone marrow, brain, liver, and spleen were collected. Using flow cytometry, we determined the presence of CD4+, CD8+, CD4+CAR+ and CD8+CAR+ cells. RESULTS AND CONCLUSION: At day 21 post-infusion, POC CAR-T cells persistence was limited and restricted mostly to the liver. However, POC CAR-T cells significantly reduced tumor burden in the liver and bone marrow (BM). When analyzing multiple time points in a comparison between POC and expansion systems, we saw that CD8+CAR+ lymphocytes were higher in the blood and BM at 24 h and increased in the brain and spleen after 7 days, with no major apparent difference between expanded and POC cells. CD4 T cells were present in the liver and spleen after 7 days but only in very low levels in the other organs at all time points. Further experiments are needed to validate these results and we are currently working to integrate a NanoLuc luciferase in the CAR construct to easily track and quantify the CAR-T cells in vivo.
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Karina Lôbo Hajdu
Oi Lucas, obrigada pelo interesse e pela pergunta. A medição é feita por bioluminescência, então as células tumorais (Nalm-6) são modificadas para expressar luciferase e ai injetamos os animais por via intraperitoneal com luciferina, que é o substrato. A reação enzimática entre a luciferase e luciferina produz luz, que é capturada por um aparelho chamado IVIS. Essa medição pode sim ser feita em vários momentos. Nos nossos experimentos, a gente costuma começar a medir por volta de 7 dias após a injeção do tumor e continua medindo até o fim do experimento semanalmente. No caso deste experimento específico que eu apresentei, fizemos a medição em dois pontos: Após 14 e 21 dias, mas a figura representativa no slide é de 21 dias.
Lucas Delmonico
Obrigado.