FUNCTIONAL CHARACTERIZATION OF THE 14Q24 RENAL CANCER SUSCEPTIBILITY LOCUS: BIOLOGY OF SUSCEPTIBILITY AND RESPONSE TO TREATMENT

The pursuit of the functional underpinnings of cancer susceptibility alleles identified by genome-wide association studies (GWAS) affords an important opportunity to gain insights into the biology of susceptibility to sporadic cancers. The sentinel renal cell cancer (RCC) susceptibility signal 14q24, rs4903064, confers an allele-specific effect on expression of the DPF3 gene, a member of the SWI/SNF complex. We demonstrate this in a massively parallel reporter assay of RCC GWAS loci and validate the allele-specific activity by individual luciferase assay. The RCC risk-associated rs4903064-C allele creates a HIF1a binding site, increasing DPF3 expression in normal kidney and RCC. The non-risk rs4903064-T allele creates a IRX2/IRX5 binding site leading to transcriptional repression. RNA-seq analyses of four stable, dox-inducible renal cell lines which can overexpress DPF3 identified over 500 deregulated genes. ATAC-seq analysis of renal cell lines over expressing DPF3 identified chromatin remodeling for 86 regions; 12% of the genes deregulated by DPF3 over expression were located up to ± 2 MB away. DPF3 over expression increased the growth rate for renal cell lines while siRNA interference of two DPF3-deregulated genes, CEMIP and IL23R, reduced the effect on growth of DPF3. We confirm the role of CEMIP in apoptosis in dox-inducible cell lines and observed DPF3 over expressing cells have a lower fraction of apoptotic cells, with reduction of cleavage of Caspase-3 and PARP, as well as BAX localization to the mitochondrial membrane. Since IL23R interacts with signaling of STAT3, an RCC oncogene, we report increased levels of STAT3 phosphorylation and observed evidence of increased activation of STAT3 signaling by expression analysis. Co-cultures of RCC tumor cells and stimulated PBMCs demonstrate that DPF3 overexpression in tumor cells leads to elevated PBMC-cytotoxicity due to an increased proportion of T CD8+ cells, suggesting DPF3 could influence immunotherapy response through the SWI/SNF complex genes. In summary, our characterization of the 14q24 RCC susceptibility locus suggests dysregulation of DPF3 expression can perturb the SWI/SNF complex leading to reduction in apoptosis and activation of the STAT3 pathway. It is remarkable that the underlying mechanism for susceptibility risk might also be associated with response to immune checkpoint inhibitor treatment, thus opening the avenue for evaluating DPF3 expression as marker for immunotherapy in RCC studies.