Abstract A020: Calcitonin receptor-like receptor is expressed in blood vessels in clear cell renal cell carcinoma and upregulated in endothelial cells co-cultured with tumor cells

Abstract

Introduction. Clear cell renal cell carcinoma (ccRCC) is a highly vascularized and clinically aggressive kidney cancer, with high rates of metastasis and recurrence after cytoreductive nephrectomy. ccRCC can be treated with anti-angiogenic drugs, however drug resistance usually develops within one year and 5-year survival for metastatic cases is approximately 10%. Calcitonin receptor-like receptor (CLR) is a G-protein-coupled receptor (GPCR) expressed in endothelial cells (EC) in core- (intracellular) and terminally-glycosylated (cell surface-associated) forms. CLR and its three agonists play a role in angiogenesis. In ccRCC, CLR is upregulated in tumor vessels when compared to vessels in adjacent tissues. The aims of this study were to: (1) investigate which types of vessels (blood or lymphatic) in ccRCC tissues express CLR and (2) determine whether ccRCC tumor cells affect expression of this GPCR in EC. Methods. CLR expression was analyzed in sections of ccRCC tissues (primary tumors) from three patients by using immunohistochemistry and our rabbit anti-human CLR antibody LN1436, with controls performed using the antigen (21 amino-acid peptide). Distribution of CLR immunoreactivity was compared to pan-EC (cluster of differentiation 31, CD31) and lymphatic EC-specific (podoplanin, PDPN) marker expression patterns, revealed by using immunohistochemistry on serial sections with mouse monoclonal antibodies JC/70A and D2-40 respectively. Two ccRCC cell lines (786-O and KTCTL-140) were co-cultured with human dermal blood vessel EC (HDBEC) in vitro for 48 hours. Cell types were separated by using magnetic-activated cell sorting (MACS) with anti-CD31 antibody-conjugated microbeads. CLR expression in ccRCC and HDBEC before, during and after co-culture was analyzed by using immunoblotting and LN1436 antibody. Results. In ccRCC tissue sections, CLR immunoreactivity was detected predominantly in blood and not lymphatic vessels. Core- and terminally-glycosylated forms of CLR were expressed in HDBEC, and the expression levels of both were upregulated after co-culture with each of the two studied ccRCC cell lines. Conclusions. These data advance knowledge about CLR expression in ccRCC and suggest that tumor cells may be responsible for its upregulation in EC within the tumor microenvironment. Our findings warrant further investigation of the potential role of the expressed in tumor blood vessels GPCR CLR as a prognostic marker and/or a target for therapy in ccRCC.

Citation Format: Matthew A. Morfitt, John Greenman, Anthony Maraveyas, Adrian L. Harris, Leonid L. Nikitenko. Calcitonin receptor-like receptor is expressed in blood vessels in clear cell renal cell carcinoma and upregulated in endothelial cells co-cultured with tumor cells [abstract]. In: Proceedings of the AACR Special Conference: Advances in Kidney Cancer Research; 2023 Jun 24-27; Austin, Texas. Philadelphia (PA): AACR; Cancer Res 2023;83(16 Suppl):Abstract nr A020.