Discovery of small molecule inhibitors of immune checkpoint proteins

Abstract

CTLA-4 and PD-1 are immune checkpoints that inhibit T cell activity to maintain immune homeostasis. These checkpoints have also emerged as highly validated cancer targets to stimulate immune responses against cancer. Despite the potential advantages of small molecule inhibitors have over antibodies, the discovery of small molecules that target the CTLA-4 or PD-1 checkpoints has greatly lagged behind the development of monoclonal antibodies. To discover small molecule inhibitors of the CTLA-4 and PD-1 signaling pathways, we have utilized a fragment-based approach. Fragment screens were conducted against CTLA-4, PD-1, and PD-L1 proteins to identify lead compounds for further optimization. While CTLA-4 and PD-1 fragment screens resulted in low hit rates and suboptimal start points, PD-L1 had a much higher hit rate and is likely druggable by small molecules. Fragments were identified that bind to PD-L1 at the PD-1 binding site and displace PD-1 using an NMR-based displacement assay. Co-crystal structures of these fragments bound to PD-L1 were obtained to facilitate structure-based design to develop more potent analogs. The fragment screening results and preliminary SAR discussed in this thesis serve as important starting points for the further development of novel small molecule-based checkpoint inhibitors.