Adhesion-GPCRs in cancer progression and metastasis

Abstract

Adhesion-GPCRs, a novel family of G protein-coupled receptors (GPCRs), are characterized by an extended extracellular region linked to a seven-pass transmembrane moiety via GPCR proteolytic site (GPS)-containing stalk region known as GAIN domain. The name adhesion refers to the presence of functional domains in the extracellular region that commonly mediate cell-cell and cell-matrix interactions in various contexts. Recently, many genome-scale analyses of genetic alterations across diverse cancer types have revealed significant alterations (copy number and mutational) in adhesion-GPCRs, yet no comprehensive examination of their roles in cancer biology exists. Through a systematic screening for all adhesion-GPCRs by RT-qPCR in murine mammary carcinoma cell lines with varying metastatic abilities as well as tumor samples of different grades, I have identified several candidate genes with possible roles in breast cancer progression and metastasis. Based on these analyses and cross-referencing with the published gene expression data on human breast cancer cell lines and patient samples, I chose two candidate genes, CELSR2 and GPR126, for more detailed investigation. To elucidate their functions in cancer biology, I investigated the effects of their perturbations using RNAi (loss-of-function) methods both in vitro and in vivo. The results from my work reveal that loss of CELSR2 affects neither tumor growth nor lung metastasis in a xenograft mouse model of breast cancer, despite enhancing invadopodial activity in vitro. I also show that highly metastatic breast cancer and melanoma cells have elevated levels of GPR126, and confirm the significance of this result by revealing (a) reduction in pulmonary metastasis without affecting primary tumor growth in a spontaneous metastasis model of breast cancer, and (b) reduction in lung metastasis in three different experimental metastasis models of breast cancer and melanoma, upon shRNA-mediated knockdown of GPR126. After probing the different steps in the metastatic cascade to investigate how GPR126 promotes metastasis, I demonstrate that GPR126 specifically affects extravasation, most likely through its engagement with type IV collagen in the sub-endothelial basement membrane. Thus, the work described in this thesis contributes to our overall understanding of the perplexing problem of cancer metastasis via identification of novel regulators of distinct steps along the ominous path of malignant cells from primary sites to distant organs.