Grainger, Jordan

Abstract Background: With few targeted therapies available for triple-negative breast cancer (TNBC), there is a critical need to identify biomarkers that can predict chemotherapy-response. Copy number amplifications (CNAmps) have been implicated in tumorigenesis, progression, and treatment response. Previously, we performed a prospective neoadjuvant chemotherapy study (BEAUTY) and identified a 5 Mb CNAmp in TNBC patients associated with pathological complete response rates to taxane- and anthracycline-based chemotherapy. Here we further interrogated this amplicon and its role in chemotherapy response. Methods: Using siRNA screening of genes identified in the 5 Mb CNAmp, followed by cytotoxicity assays, we identified RDH10 significantly associated with chemo-sensitivity in TNBC cell lines. Functional studies were performed using RDH10 knockdown or overexpression followed by cytotoxicity, cell proliferation, luciferase reporter assays, HPLC or western blot. Results: RDH10 is a rate-limiting step involved in the synthesis of all-trans retinoic acid (ATRA) - the active metabolite of retinoid metabolism that is frequently aberrant in cancer. Previous studies demonstrated ATRA to be a potent inhibitor of PIN1, a key regulator of several oncogenic signaling pathways often amplified in TNBC. We discovered that genetic manipulation of RDH10 affected ATRA intracellular concentrations, contributing to PIN1 degradation and taxane-response. Furthermore, co-treatment of TNBC cells with ATRA significantly increased taxane-sensitivity. Conclusion: RDH10 amplification increases endogenous levels of ATRA and degrades PIN1 in TNBC. ATRA-mediated degradation of PIN1 sensitizes TNBC to chemotherapy, suggesting that RDH10 may be a novel biomarker of taxane response and the addition of ATRA to standard chemotherapy may improve chemo-sensitivity.

Abstract Background: With few targeted therapies available for triple-negative breast cancer (TNBC), there is a critical need to identify biomarkers that can predict chemotherapy-response. Copy number amplifications (CNAmps) have been implicated in tumorigenesis, progression, and treatment response. Previously, we performed a prospective neoadjuvant chemotherapy study (BEAUTY) and identified a 5 Mb CNAmp in TNBC patients associated with pathological complete response rates to taxane- and anthracycline-based chemotherapy. Here we further interrogated this amplicon and its role in chemotherapy response. Methods: Using siRNA screening of genes identified in the 5 Mb CNAmp, followed by cytotoxicity assays, we identified RDH10 significantly associated with chemo-sensitivity in TNBC cell lines. Functional studies were performed using RDH10 knockdown or overexpression followed by cytotoxicity, cell proliferation, luciferase reporter assays, HPLC or western blot.Results: RDH10 is a rate-limiting step involved in the synthesis of all-trans retinoic acid (ATRA) - the active metabolite of retinoid metabolism that is frequently aberrant in cancer. Previous studies demonstrated ATRA to be a potent inhibitor of PIN1, a key regulator of several oncogenic signaling pathways often amplified in TNBC. We discovered that genetic manipulation of RDH10 affected ATRA intracellular concentrations, contributing to PIN1 degradation and taxane-response. Furthermore, co-treatment of TNBC cells with ATRA significantly increased taxane-sensitivity.Conclusion: RDH10 amplification increases endogenous levels of ATRA and degrades PIN1 in TNBC. ATRA-mediated degradation of PIN1 sensitizes TNBC to chemotherapy, suggesting that RDH10 may be a novel biomarker of taxane response and the addition of ATRA to standard chemotherapy may improve chemo-sensitivity.

Abstract Triple negative breast cancer (TNBC) accounts for 15-20% of breast cancer cases in the United States. With currently no targeted therapies for TNBC, identification of novel biomarkers to improve chemo-response is vital. Recently, copy number amplifications (CNAs) have been implicated in treatment response in several diseases including breast cancer. Our clinical studies have associated a 5 Mb CNA in TNBC patients with chemo-response, which may elucidate novel biomarkers and treatment strategies for TNBC. Analysis of RNAseq and Whole Exome Sequencing data from 42 TNBC patient tumors enrolled in the BEAUTY Study, identified a 5 Mb CNA on chromosome 8q which associated with pathological complete response following taxane and anthracycline chemotherapy. By performing siRNA knockdown cytotoxicity studies in TNBC cell lines for genes within this amplified region, we identified RDH10 as a potential novel biomarker for increasing chemo-sensitivity. RDH10 is the rate limiting step in the synthesis of all-trans retinoic acid (ATRA). Previous studies have demonstrated ATRA to be a potent inhibitor of PIN1, a key regulator of several oncogenic signaling pathways, amplified in TNBC. We discovered overexpression of RDH10 to increase endogenous levels of ATRA, contributing to the degradation of PIN1 and taxane sensitivity. A rescue experiment with PIN1 was able to restore the chemo-resistance phenotype. Similarly, we found treating TNBC cells with ATRA phenocopied RDH10's effect on PIN1 and improved chemo-sensitivity. Furthermore, analyses of chemo-responsive PDX tumors containing the 5 Mb CNA, overexpressed RDH10 and exhibited lower levels of PIN1 when compared to non-responders. In conclusion, RDH10 amplification increases endogenous levels of ATRA and degrades PIN1 in TNBC. ATRA-mediated degradation of PIN1 sensitizes TNBC to chemotherapy, suggesting RDH10 as a novel biomarker for chemo-response and ATRA as a novel treatment to improve chemo-sensitivity. Citation Format: Jordan Mark Grainger, Jia Yu, Krishna R. Kalari, Peter Vedell, Kevin Thompson, Matthew P. Goetz, Vera J. Suman, Judy C. Boughey, Liewei Wang. Study of copy number amplification and chemo-response in triple negative breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1928.