Abstract B97: Somatic copy number alterations by whole-exome sequencing reveals YWHAZ and PTK2 as potential therapeutic targets in castration-resistant prostate cancer
2013-02-01

Abstract Aim: Castration resistant prostate cancer (CRPC) is the most aggressive form of prostate cancer (PCa) with a poor prognosis, and remains a significant therapeutic challenge. The key to the development of novel therapeutic targets for CRPC is to decipher the molecular alterations underlying this lethal disease. The aim of our study was to identify therapeutic targets for CRPC by assessing somatic copy number alterations (SCNA) by whole exome sequencing on five CRPC/normal paired formalin fixed paraffin embedded (FFPE) samples using the SOLiD4 next generation sequencing platform. Methods: Genomic DNA was extracted from 5 CRPC/normal paired FFPE samples and sequenced using the SOLiD4 next generation sequencing platform. The sequencing output was mapped, sorted, filtered and annotated using human genome databases. Among a set of prominent amplified/deleted genes in PCa, the 8q12.2-24.22 region encoding PTK2 and YWHAZ was found to be amplified. The data was validated using fluorescence in-situ hybridization (FISH) assays with a PCa progression cohort containing 137 localized PCa samples, 105 primary tumors, 71 corresponding lymph node metastases and 39 CRPC samples. Additionally PTK2 and YWHAZ amplification status, mRNA expression (LightCycler) and protein expression (Western Blot) were determined in selected PCa cell lines. Within PC-3 cells, inhibition of PTK2 by the specific inhibitor TAE226 (Novartis), as well as knock down of YWHAZ were analyzed. The effect of PTK2 inhibition and YWHAZ knock down on cell proliferation (MTT assay) and cell-invasion (Matrigel invasion chambers) were determined. Results: Whole exome sequencing analysis identified regions of deletion and amplification, which included well-known genes such as PTEN, CMYC and AR. Furthermore, we identified PTK2 and YWHAZ on chromosome 8 to be amplified in the sequenced PCa samples. The FISH analysis of the region showed an increasing amplification frequency of PTK2 and YWHAZ with progression of the disease. PTK2 was amplified in 1% of the localized PCa and in 35% of the CRPC samples. YWHAZ was amplified in 4% of the localized PCa and in 48% of the CRPC samples. The FISH analysis revealed a high level amplification for PTK2/YWHAZ, in the PC-3 cells, which also exhibited a high PTK2/YWHAZ protein expression. PTK2 inhibition by TAE226 and YWHAZ knock down had a significant effect on cell proliferation and migration in the PC-3 cells in vitro. PC-3 cells treated with TAE226 or YWHAZ knocked down cells had a significantly reduced cell proliferation and cell migration ability compared to untreated cells. Conclusion: Our findings suggest that the inhibition of PTK2 and YWHAZ could delay the progression of the disease in CRPC patients harbouring amplification of the latter genes. The validated whole exome sequencing data shows that FFPE tissue could be a promising alternative for SCNA screening using next generation sequencing technologies. Citation Format: Roopika Menon, Kerstin Rüenauver, Mario Deng, Friedrich Kunze, Diana Boehm, Wenzel Vogel, Veit Scheble, Falko Fend, Glen Kristiansen, Nicolas Wernert, Nicole Oberbeckmann, Saskia Biskup, Mark Rubin, Zaki Shaikhibrahim, Sven Perner. Somatic copy number alterations by whole-exome sequencing reveals YWHAZ and PTK2 as potential therapeutic targets in castration-resistant prostate cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr B97.