武装肿瘤坏死因子-α的TCR-T和CAR-T细胞在无毒性的情况下为实体瘤提供优越的抗癌活性

Background: TCR-T and CAR-T cells directed against tumor-specific antigens are associated with robust initial clinical responses which are often not durable against solid tumors, demonstrating an urgent need for more potent cell therapy products. We created TNF-alpha (TNF-a) armed variants of TCR-T and CAR-T cells to test their effectiveness and toxicity against a variety of solid tumors. Methods: Lentiviral vectors encoding the TCR/CAR of interest with or without a supplemental copy of TNF-a were generated for NY-ESO-1 TCR, HPV-16 E7 TCR, and GD2 CAR. Mock, conventional TCR/CAR, and TNF-a armed TCR/CAR T-cells were co-cultured with nRFP-transduced tumor cells expressing the corresponding tumor antigen and appropriate HLA-A haplotype for the TCR-T cells (A375/M257-A2 melanoma cells for NY-ESO-1, 4050/CaSki cervical carcinoma cells for HPV-16 E7, and 143B/G292 osteosarcoma cells for GD2). Cells were subjected to repetitive stimulation assays to assess tumor killing efficiency in the setting of chronic antigen exposure. In parallel, in vivo tumor challenges were performed using humanized NSG mouse xenografts treated with mock, conventional TCR/CAR, or TNF-a-armed TCR/CAR T-cells. Mice were subjected to end of study necropsies to assess end-organ toxicity. T-cell phenotypes in circulation and intratumorally were assessed via flow cytometry, while serum cytokines were assessed weekly via multiplex sandwich ELISA. Tumors were also subjected to spatial transcriptomic analysis via CosMx. Results: In vitro, TNF-a-armed TCR/CAR T-cells displayed significantly increased tumor cell killing and significantly increased secretion of TNF-alpha both initially and over time, exclusively in a tumor antigen-dependent manner. In vivo, TNF-a-armed TCR/CAR T-cells displayed superior tumor control and mouse survival compared to the corresponding conventional TCR/CAR. Importantly, this superior tumor control occurred in the absence of any systemic increases in TNF-a or IL-6, systemic toxicity (temperature, weight, body condition score), as well as without any differences in end-organ inflammatory damage/lymphocytic infiltration. Tumors treated with TNF-a-armed TCR/CAR T-cells displayed significantly increased T-cell infiltration, as well as significant reductions in intratumoral MDSCs, CD4 Tregs and Th2 cells and associated cytokines, while tumor cells displayed significant increases in TNF-a signaling, and significant decreases in IL-10 immunosuppressive pathways arising from the Tregs. Conclusions: TNF-a-armed TCR/CAR-T cells possess superior antitumor efficacy against a wide variety of solid tumors, in the absence of any systemic toxicity, due to the antigen-dependent nature of the increased TNF secretion. This is due to both increased local TNF-a secretion and antitumor effector activity, as well as decreased Treg activity within the tumor.