Abstract 1240: Targeting epigenetic regulation in clear cell renal cell carcinoma

Abstract

Abstract Clear cell renal cell carcinoma (ccRCC), the most common subtype of renal cell cancer, is highly resistant to traditional radiation and chemotherapy. Recently, targeted therapies have extended progression-free survival, but responses are variable and no significant overall survival benefit has been achieved; thus, treatment options are scarce for patients bearing this disease. The majority of ccRCC cases (∼90%) are characterized by bi-allelic loss of the von Hippel Lindau (VHL) gene, followed by mutations in the epigenetic regulators PBRM1 (23%), SETD2 (8.4%), BAP1 (7.1%), and KDM5C (4.7%). The latter four genes implicate an integral role for chromatin remodeling in ccRCC, and thus signify a new realm of exploration and therapeutic targeting for this disease. Given the limited number of commercially available ccRCC cell lines, and reports that they may not accurately reflect primary tumors at the molecular level, our lab developed a novel method to generate primary patient-derived ccRCC (VHLmut) cells and matched normal renal proximal tubular epithelial (VHLwt) cells from human surgical specimens with high efficiency. This involved fluorescence-activated cell sorting of primary tumor single cell suspensions using the cell surface marker carbonic anhydrase IX (CA9), a HIF target which is upregulated upon VHL loss, to establish CA9+ VHLmut and CA9− VHLwt cells. Transcriptional profiling of VHLmut and VHLwt pairs found gene signatures that corresponded with patient matched primary ccRCC tumors and adjacent normal tissues in The Cancer Genome Atlas, demonstrating that these cells represent novel models for interrogation of ccRCC biology and testing of therapeutic strategies. Here, in the current study, these newly established cells were utilized to screen a focused library of 30 well-characterized drug-like compounds targeting specific components of chromatin remodeling complexes to determine the effect on cell growth in culture. At Day 0, 786-0 (−VHL) cells and VHLmut cells corresponding to four patient primary ccRCC tumors were plated at 2,500 cells/well in 96-well black clear-bottom plates (n = 5). Cells were then treated with the library of chromatin-targeting drugs on Day 1 and monitored until control-treated cells were 90-95% confluent. At endpoint, cells were fixed and stained with DRAQ5, a DNA intercalating dye that stains DNA stoichiometrically, for visualization on the LI-COR Odyssey CLx Imaging System. Across all five samples, there was a significant reduction in growth observed among cells treated with drugs targeting: i) BET bromodomains (BRD2, BRD3, BRD4 and BRDT), ii) EZH1/2, and iii) JMJD3, UTX and JARID1B. Ongoing work will determine mechanisms of action and whether these targets are also affected in matching VHLwt cells in order to identify compounds that may translate well in the clinic to treat patients with ccRCC. Citation Format: Anthony J. Apostoli, Nazleen Lobo, Panagiotis Prinos, Dalia Barsyte-Lovejoy, Cheryl Arrow Smith, Laurie Ailles. Targeting epigenetic regulation in clear cell renal cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1240.