Novel mechanism for eliminating tumor collagens to boost efficacy by attIL12-TIL therapy in PDX sarcoma models

Tumor-targeted T-cell therapies of various types have been booming, but T-cell therapy is limited by its inability to penetrate the collagen barrier that “stuffed” within tumors and surrounding tumors/tumor cells. Compared to normal human tissues, sarcoma tissues exhibited significantly higher density of collagens and extracellular matrix (ECM) determined via bulk RNAseq and Masson's trichrome staining. The destruction of tumor collagen is significant because collagen does not only suppresse T cells but also contributes to the formation of the extracellular matrix. In this study, for the first time, we used three pairs of autologous sarcoma patient-derived xenograft (PDX) and autologous tumor infiltrating T cells (TILs) models to discover that CCKAR directly boosts collagen production by tumor cells in vitro and in vivo. attIL12-modified TILs disabled collagen production from CCKAR-high autologous tumor cells in vitro and in vivo. This attIL12-TILs induced disruption of tumor cell derived collagen production required a simultaneous interaction between two signaling pathways: 1) the CSV on autologous tumor cells, which is targeted by attIL12, and 2) HLA-TCR on attIL12-TILs; when either interaction was abrogated, collagen production and CCKAR expression were not shut down. Mechanistically, the dual signaling activation between attIL12-TILs and autologous tumor cells synergistically induced super high IFNgamma production in tumors, which in combination with CCKAR downregulation reduced collagen expression through suppression of both TGFbeta-stimulated SMAD activation and CCKAR-AKT signaling. Diminishing collagen expression from tumor cells significantly increased T-cell infiltration and improved tumor growth inhibition in PDX sarcomas. This study revealed for the first time that CCKAR, which is highly expressed in immunotherapy resistant sarcomas, may serve as a novel target for reducing tumor collagens. attIL12 modification mediated collagen disrupting reversed the lost tumor penetration function of TILs post ex vivo expansion, resulting in sarcoma tumor regression that TILs alone fail to achieve. Thus, this attIL12-TIL therapy holds great clinical potential for boosting T-cell infiltration in high-grade, collagen-rich tumors.