The human glucagon receptor (GCGR) belongs to the class B G protein-coupled receptor (GPCR) family and plays a key role in glucose homeostasis and the pathophysiology of type 2 diabetes. Here we report the 3.0 Å crystal structure of full-length GCGR containing both extracellular domain (ECD) and transmembrane domain (TMD) in an inactive conformation. The two domains are connected by a 12-residue segment termed the ‘stalk’, which adopts a β-strand conformation, instead of forming an α-helix as observed in the previously solved structure of GCGR-TMD. The first extracellular loop (ECL1) exhibits a β-hairpin conformation and interacts with the stalk to form a compact β-sheet structure. Hydrogen/deuterium exchange, disulfide cross-linking and molecular dynamics studies suggest that the stalk and ECL1 play critical roles in modulating peptide ligand binding and receptor activation. These insights into the full-length GCGR structure deepen our understanding about the signaling mechanisms of class B GPCRs.
BackgroundAdoptive transfer of tumor infiltrating or circulating lymphocytes transduced with tumor antigen receptors has been examined in various clinical trials to treat human cancers. The tumor antigens targeted by transferred lymphocytes affects the efficacy of this therapeutic approach. Because cancer stem cells (CSCs) play an important role in tumor growth and metastasis, we hypothesized that adoptive transfer of T cells targeting a CSC antigen could result in dramatic anti-tumor effects.ResultsAn EpCAM-specific chimeric antigen receptor (CAR) was constructed to transduce human peripheral blood lymphocytes (PBLs) and thereby enable them to target the CSC marker EpCAM. To investigate the therapeutic capabilities of PBLs expressing EpCAM-specific CARs, we used two different tumor models, PC3, the human prostate cancer cell line, which has low expression levels of EpCAM, and PC3M, a highly metastatic clone of PC3 that has high expression levels of EpCAM. We demonstrate that CAR-expressing PBLs can kill PC3M tumor cells in vitro and in vivo. Despite the low expression of EpCAM on PC3 cells, CAR-expressing PBLs significantly inhibited tumor growth and prolonged mouse survival in a PC3 metastasis model, probably by targeting the highly proliferative and metastatic population of cancer cells.ConclusionsOur data demonstrate that PBLs expressing with EpCAM-specific CARs have significant anti-tumor activity against prostate cancer. Therefore, the adoptive transfer of T cells targeting EpCAM could have great potential as a cancer treatment.
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