PROJECT SUMMARY/ABSTRACT T cells are central players in immune responses to infections and cancer. Once activated by antigens, T cells are able to directly kill infected cells and cancer cells, or direct other components of the immune system to attack targets. The awesome destructive power of T cells needs to be tightly controlled in order to avoid collateral damages to normal tissues and to prevent autoimmune diseases. An important mechanism of control is to inhibit T cell activities through B7 family immune checkpoint ligands. To date nine such ligands have been identified. They bind to receptors on T cells to inhibit T cell proliferation and activation. Interestingly, many types of tumor cells co-opt these ligands to evade T cell attack. Recently, antibodies that block the interactions between these ligands and their receptors have been developed to enhance immune response to cancer. These so-called immune checkpoint blockade drugs have been successful in controlling tumors in some patients, but the overall response rates are still low. One of the reasons may be that most of the drugs target the receptor of only one of the immune checkpoint ligands while cancer cells may express many of the different types of ligands. Currently, targeting additional receptors are difficult because the receptors for five of the ligands have yet to be identified. A major challenge for identifying their receptors is that they bind their ligands very weakly and currently available methods do not have enough sensitivity. To overcome this, we propose to develop a novel and highly sensitive cell-based approach to identify receptors for the five orphan ligands. The feasibility of the approach will be tested using model ligands and receptors that are known bind to each other. We will use the cell-based approach to screen transmembrane protein collections and the identified receptor candidates will be tested for their abilities to bind the orphan ligands. Receptors identified using our approach should help understand how B7 family immune checkpoint ligands control T cell responses and may serve as new targets for immune checkpoint blockade therapies against cancer.