First immunotherapeutic CAR-T cells against the immune checkpoint protein HLA-G.

BACKGROUND: CAR-T cells immunotherapy is a breakthrough in the treatment of hematological malignancies such as acute lymphoblastic leukemia (ALL) and B-cell malignancies. However, CAR-T therapies face major hurdles such as the lack of tumor-specific antigen (TSA), and immunosuppressive tumor microenvironment sometimes caused by the tumorous expression of immune checkpoints (ICPs) such as HLA-G. Indeed, HLA-G is remarkable because it is both a potent ICP and a TSA. HLA-G tumor expression causes immune escape by impairing innate and adaptive immune responses and by inducing a suppressive microenvironment. Yet, to date, no immunotherapy targets it. METHODS: We have developed two anti-HLA-G third-generation CARs based on new anti-HLA-G monoclonal antibodies. RESULTS: Anti-HLA-G CAR-T cells were specific for immunosuppressive HLA-G isoforms. HLA-G-activated CAR-T cells polarized toward T helper 1, and became cytotoxic against HLA-G+ tumor cells. In vivo, anti-HLA-G CAR-T cells were able to control and eliminate HLA-G+ tumor cells. The interaction of tumor-HLA-G with interleukin (IL)T2-expressing T cells is known to result in effector T cell functional inhibition, but anti-HLA-G CAR-T cells were insensitive to this inhibition and still exerted their function even when expressing ILT2. Lastly, we show that anti-HLA-G CAR-T cells differentiated into long-term memory effector cells, and seemed not to lose function even after repeated stimulation by HLA-G-expressing tumor cells. CONCLUSION: We report for the first time that HLA-G, which is both a TSA and an ICP, constitutes a valid target for CAR-T cell therapy to specifically target and eliminate both tumor cells and HLA-G+ suppressive cells.

HLA-G Neo-Expression on Tumors.

HLA-G is known to modulate the immune system activity in tissues where physiological immune-tolerance is necessary (i.e., maternal-fetal interface, thymus, and cornea). However, the frequent neo-expression of HLA-G in many cancer types has been previously and extensively described and is correlated with a bad prognosis. Despite being an MHC class I molecule, HLA-G is highly present in tumor context and shows unique characteristics of tissue restriction of a Tumor Associated Antigen (TAA), and potent immunosuppressive activity of an Immune CheckPoint (ICP). Consequently, HLA-G appears to be an excellent molecular target for immunotherapy. Although the relevance of HLA-G in cancer incidence and development has been proven in numerous tumors, its neo-expression pattern is still difficult to determine. Indeed, the estimation of HLA-G's actual expression in tumor tissue is limited, particularly concerning the presence and percentage of the new non-canonical isoforms, for which detection antibodies are scarce or inexistent. Here, we summarize the current knowledge about HLA-G neo-expression and implication in various tumor types, pointing out the need for the development of new tools to analyze in-depth the HLA-G neo-expression patterns, opening the way for the generation of new monoclonal antibodies and cell-based immunotherapies.