VHL-mutant renal cell carcinomas contain cancer cells with mesenchymal phenotypes.

4568 Background: To study “cancer stem cells” it is imperative to account for all stromal cell populations within the tumour. The existence of “cancer stem cells” in clear cell renal cell carcinoma (ccRCC) has not been examined in ex vivo patient samples. Methods: We established a multiplex flow cytometry (FC) antibody panel in ccRCC, which reliably identified stromal lineages including CD45+ immune, CD31+/CD144+ endothelial and fibroblast-marker-positive subpopulations, thus allowing isolation of "lineage-negative" tumor cells. To verify the identity of tumour-derived populations as either cancer cells or normal stromal cells, we took advantage of the fact that mutations in VHL occur early during ccRCC tumorigenesis and are found in two-thirds of patients. Results: We sequenced 18 patient tumor samples, 12 of which had VHL exome mutations. Targeted re-sequencing of FC sorted subpopulations from these patients’ samples revealed that while CD45+ immune cells and CD31+/CD144+ endothelial cells were genetica...