CARTIV - a novel platform for regulation of gene expression under the control of inflammation-induced promoters

Engineered immune cells, such as chimeric antigen receptor (CAR) T cells, are facing the danger of an overt life-threatening immune response due to the ON-target OFF-tumor cytotoxicity and Cytokine Release Syndrome. Even though several treatments have been approved for different B-Cell malignancies and hundreds of ongoing clinical trials, to date none are approved for solid tumors. We therefore developed a synthetic promoter platform for regulation of gene expression under the control of inflammation and Hypoxia-induced signals that are associated with the tumor microenvironment (TME). We termed this methodology as chimeric-antigen-receptor-tumor-induced-vector (CARTIV). As a proof of concept, we studied synthetic promoters based on promoter-responsive elements (PREs) of IFNγ, TNFα, TGF beta, IL6 and hypoxia; triple PRE-based CARTIV promoters manifested a synergistic activity in cell-lines and potent activation in human primary T-cells. We also designed, screened and validated a promoter library that resulted in novel promoters, some of which possess non-trivial properties such as extremely low background expression levels while maintaining a high induction fold change. Using CARTIV platform can improve safety of CAR T-cells or other engineered immune-cells, providing TME-focused activity and opening a therapeutic window for many tumor-associated antigens that are also expressed by non-tumor healthy tissues.