Dados do Resumo

Título

Triple-negative breast cancer treatment using Anti-CAIX CAR-T cells releasing anti-PD-L1 monoclonal antibodies in NSG mice

Introdução

Triple-negative breast carcinoma (TNBC) is an aggressive tumor usually resistant to conventional treatments, resulting in an unfavorable prognosis. Advances in cancer therapy, such as using T cells modified to express chimeric antigen receptors (CAR-T cells), have shown success against hematological tumors and must be improved to be useful against solid tumors. The carbonic anhydrase IX (CAIX), an enzyme present in several hypoxic tumors, and PD-L1, an immune checkpoint molecule, are often expressed in TNBC, being both promising targets for developing CAR-T cell therapies.

Objetivo

This project aims to evaluate the anti-tumor effects of CAIX-targeted CAR T cells containing CD28 or 4-1BB as costimulatory domains and capable of decreasing T cell exhaustion by secretion of anti-PD-L1 antibodies using a single treatment in a TNBC model established from Hs578T cell line in NSG mice.

Métodos

The effects of all anti-CAIX CAR T cells were tested against human triple-negative cells Hs578T in vitro and in an in vivo orthotopic model established using the same cells in NSG mice. Lentiviral vectors were generated via transient transfection, concentrated, titrated, and transduced into T cells purified from healthy donor blood mononuclear fractions at a CD4:CD8 2:1. The resulting CAR T cells had transduction levels assessed. Then, around 1x10(7) viable and transduced CAR T cells were injected into each animal through the tail vein. Treatment effects with CAR T cells were evaluated by monitoring tumor growth, final tumor weight, occurrence of macroscopic metastases, abdominal wall infiltration, tumor-infiltrating T cells, and overall survival. The Anti-CAIX CAR-T groups used in this assay comparatively had the 4-1BB and/or CD28 co-stimulatory domains and were capable of secreting anti-PD-L1 antibodies, and as a control group, we used Anti-BCMA CAR-T cells. Approved by CEUA 088/21/ CIPE A.C. Camargo Cancer Center; CAAE 71055617.3.0000.5594/ UFABC; CIBio/UFABC CQB 0304/10 001/2017. FAPESP n. 2023/03631-3 and n.2018/17656-0.

Resultados

The CD28 construct secreting anti-PD-L1 antibodies decreased tumor area and volume up to 6-8-fold in the first 18 days after treatment, resulting in tumors with lower weight. The same construction was associated with better overall survival than the 4-1BB construction, the combination of these two constructions 50%/50% and the control group. No macroscopic metastases were observed in any of the groups. The presence of tumor-infiltrating cells was not durable due to the absence of human T or B cells secreting cytokines necessary to keep CAR T cells alive in the long term in NSG mice, resulting in a bias that leads to a loss of objective response in this model with time.

Conclusões

A promising objective response and improved overall survival were observed, especially with Anti-CAIX CD28 secreting anti-PD-L1 antibodies in this model of human TNBC tumor in mice. This encourages CAR-T Anti-CAIX therapy inducing immune checkpoint blockade to be tested in other models of TNBC, as well as in additional types of solid tumors that are difficult to treat and have the expression of this target molecule, proposing alternatives for complex tumors with scarce treatment options.

Financiador do resumo

FAPESP n. 2023/03631-3 and n.2018/17656-0; CAPES

Palavras Chave

triple-negative breast cancer; CAIX; CAR-T Cell