Dados do Resumo

Título

ANTI-CARBONIC ANHYDRASE IX CAR T CELLS RELEASING PROGRAMMED CELL DEATH LIGAND 1 ANTIBODIES IN THE TREATMENT OF GLIOBLASTOMA

Introdução

Glioblastoma (GBM) is the most prevalent of all brain tumors. Within GBM, hypoxia is a frequent characteristic that leads to the overexpression of several proteins, including carbonic anhydrase IX (CAIX). Solid tumors have a microenvironment that promotes T cell exhaustion, which favors tumor development. In the case of GBM, an overexpression of programmed cell death ligand-1 (PD-L1), a molecule related to exhaustion induction, is very often. A new therapeutic strategy has been highlighted in the fight against cancer: T cells expressing chimeric antigen receptors (CAR T cells) designed to target a tumor antigen. The project was approved by CEP 3312/22 and CEUA 088/21 by the Institutional Ethics Committee of the A. C. Camargo Cancer Center.

Objetivo

This project aims to evaluate the in vitro efficacy of anti-CAIX CAR T cells releasing anti-PD-L1 antibodies in the tumor microenvironment against GBM cell lines in vitro.

Métodos

GBM cells U251 and A172 were used for this study. Basal levels of CAIX and PD-L1 expression were assessed by flow cytometry. The lentiviruses will be produced by transient transfection, concentrated, titrated and transduced into CD4:CD8 2:1 T cells purified from the mononuclear blood fraction of healthy donors. The resulting CAR T cells were expanded, and their transduction levels were assessed in the short and long term. Anti-CAIX CAR T cells containing different CD28 or 4-1BB co-stimulatory domains will be evaluated in vitro in a GBM model, determining the secretion of IL-2, IFNγ, and IgG secretion levels.

Resultados

U251 is positive for CAIX and PD-L1, while A172 has lower levels of CAIX and higher PD-L1. A high percentage of T cells were efficiently transduced with CAR-Coding lentiviruses. All anti-CAIX CAR T cells were able to induce cytotoxicity in U251 (CAIX+/PD-L1+), with anti-CAIX CAR T cells secreting anti-PD-L1 being slightly more cytotoxic to these cells than CAR T cells targeting only CAIX. In addition, about 35% of all Anti-CAIX CAR T cells were activated in contact with tumor cells, as demonstrated by IL-2 expression in about 35% of T cells, and INFγ expressed in 20-40% CAR T cells from different groups.

Conclusões

U251 is positive for CAIX and PD-L1, while A172 has lower levels of CAIX and higher PD-L1. A high percentage of T cells were efficiently transduced with CAR-Coding lentiviruses. All anti-CAIX CAR T cells were able to induce cytotoxicity in U251 (CAIX+/PD-L1+), with anti-CAIX CAR T cells secreting anti-PD-L1 being slightly more cytotoxic to these cells than CAR T cells targeting only CAIX. In addition, about 35% of all Anti-CAIX CAR T cells were activated in contact with tumor cells, as demonstrated by IL-2 expression in about 35% of T cells, and INFγ expressed in 20-40% CAR T cells from different groups.

Financiador do resumo

FAPESP 2023/03631-3 and 2018/17656-0; CAPES 88887.970277/2024-00

Palavras Chave

CAR-T; Glioblastoma; CAIX