Abstract A74: Proteasome subunit RPN13 as a candidate target for treatment of ovarian carcinoma.
Authors: Rosie T. Jiang, Ravi K. Anchoori, Anna V. Yemelyanova, Chien-fu Hung
Journal: Clinical Cancer Research (2016)

Abstract The aberrant metabolism of ovarian cancer cells produces an excess of misfolded proteins and particular sensitivity to proteasome inhibition. Blockade of proteasome 19S regulatory particle (RP) deubiquitinase and/or 20S subunit proteolytic function results in a rapid buildup of denatured protein aggregates and triggers a p53-independent cell death. However, treatment of ovarian cancer with the licensed proteasome inhibitor bortezomib did not produce sufficient benefit to warrant further development. This setback led us to develop second generation inhibitors representing a distinct class of molecule and new mechanism of proteasome inhibition. We have developed novel bis-benzylidine piperidones, RA183 and RA190, which bind covalently to the RPN13 subunit of the 19S RP. RPN13 acts as an ubiquitin receptor and promotes the deubiquitinase activity of its binding partner UCH37. RA183 and RA190 cause rapid accumulation of very high molecular weight poly-ubiquitinated proteins, higher and more rapid than seen with bortezomib treatment, suggesting they inhibit RPN13 function, and thus deubiquitination by the 19S regulatory particle and 20S catalytic subunit activity. The accumulation of polyubiquitinated proteins triggered by RA183 or RA190 causes endoplasmic reticulum (ER) stress and an unresolved unfolded protein response that rapidly triggers apoptosis. RA190 and RA183 are active against multiple high-grade ovarian carcinoma cell lines in vitro and ovarian cancer models in mice. In nude mice carrying intraperitoneal ES2-luciferase xenografts, RA190 treatment (10mg/kg) significantly inhibited tumor growth. RPN13 is amplified in a subset of high grade ovarian cancers (10-20%), but its amplification did not correlate with a change in the sensitivity of ovarian carcinoma cell lines to RPN13 inhibitor. Sensitivity to RA190 also did not correlate with the level of pre-treatment stress or protein poly-ubiquitination. Knockdown of RPN13 expression in mouse ovarian cell line, ID8, increased its sensitivity to RA190. However attempts to generate knockout of RPN13 by CRISPR in NIH:OVCAR-3 failed and appeared to reflect toxicity. RPN13 mRNA was detected by chromogenic RNA in situ hybridization in all high grade serous carcinoma (HGSC) cases tested and matched precursor serous tubal intraepithelial carcinomas(n=16), and was overexpressed in all relative to adjacent normal fallopian tube epithelium. We conclude that RPN13 has promise as a target for treatment of ovarian carcinomas and possibly HGSC precursor STIC lesions, and that further development of RPN13 inhibitors warranted. Citation Format: Rosie T. Jiang, Ravi K. Anchoori, Anna V. Yemelyanova, Chien-fu Hung. Proteasome subunit RPN13 as a candidate target for treatment of ovarian carcinoma. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr A74.