UC San Diego

Programmed cell death protein 1 (PD-1) and signal regulatory protein α (SIRPα) are receptors that transduce inhibitory signals to negatively regulate effector functions of immune cells, which were both utilized by cancers to evade and inhibit the immune response. Through protein engineering, integrated sensing and activating proteins (iSNAPs) were developed to replace the cytoplasmic domains of PD-1 and SIRPα receptors. With Fyn as the activating module, engineered Fyn-iSNAP replaced the cytoplasmic tail of PD-1, which was intended to reprogram PD-1/PD-L1 inhibitory signaling into activating Fyn signaling upon ligand binding. However, characterization of Fyn-iSNAP in HeLa cells demonstrated that this engineered protein failed to show desired functions. In the second chapter, in order to rewire SIRPα/CD47 inhibitory pathway, Shp2-iSNAPs and Syk-iSNAPs were developed and used to replace the original SIRPα cytoplasmic domain in our previous studies (data not published yet). In order to increase gene delivery efficiency, the sizes of the engineered SIRPα-iSNAPs were reduced. The expression of the truncated proteins in mouse macrophages led to enhanced phagocytosis of opsonized red blood cells (RBCs). Also, human promyelocytic leukemia cell derived macrophages (PLDMs) were engineered for the first time using the SIRPα-Shp2-iSNAPs and its truncated version, which demonstrated enhanced engulfment against opsonized RBCs as well as tumor cells. Therefore, these engineered proteins with activating module potentially offer a new approach for future cancer immunotherapy.