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PARP Inhibition Suppresses GR–MYCN–CDK5– RB1–E2F1

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PARP Inhibition Suppresses GR–MYCN–CDK5– RB1–E2F1 Published OnlineFirst August 22, 2019; DOI: 10.1158/1078-0432.CCR-19-0317 Translational Cancer Mechanisms and Therapy Clinical Cancer Research PARP Inhibition Suppresses GR–MYCN–CDK5– RB1–E2F1 Signaling and Neuroendocrine Differentiation in Castration-Resistant Prostate Cancer Bo Liu1, Likun Li1, Guang Yang1, Chuandong Geng1, Yong Luo1, Wenhui Wu2, Ganiraju C. Manyam2, Dimitrios Korentzelos1, Sanghee Park1, Zhe Tang1, Cheng Wu1, Zhenyang Dong1, Michael Sigouros3, Andrea Sboner4,5,6, Himisha Beltran7, Yu Chen3,8,9, Paul G. Corn1, Michael T. Tetzlaff10, Patricia Troncoso10, Bradley Broom2, and Timothy C. Thompson1 Abstract Purpose: In this study, we addressed the underlying ing pathway that drives NED in prostatic adenocarcinoma mechanisms for the association between enzalutamide (ENZ) PDX and cell line models. Mechanistically, long-term treatment and neuroendocrine prostate cancer (NEPC), and ENZ treatment transcriptionally upregulates signaling of the critical involvement of MYCN, and loss of RB1 function in the GR–MYCN axis, leading to CDK5R1 and CDK5R2 upre- neuroendocrine differentiation (NED) of prostatic epithelial gulation, Rb1 phosphorylation, and N-Myc–mediated cells, and the development of NEPC. We further sought to and E2F1-mediated NED gene expression. Importantly, determine whether PARP inhibition could suppress NEPC, olaparib (OLA) or talazoparib (TALA) suppressed these and to identify molecular determinants of this therapeutic activities, and the combination of OLA and dinaciclib activity. (DINA), an inhibitor of CDK2 and CDK5, which also Experimental Design: We used a novel prostate cancer inhibits Rb1 phosphorylation, suppressed NED and signif- patient–derived xenograft (PDX) treatment model, prostatic icantly improved therapeutic efficiency in NEPC cells in vitro adenocarcinoma and NEPC cell lines, an NEPC organoid line, and in NEPC tumors in vivo. and NEPC xenograft models to address the mechanistic basis Conclusions: The results of our study indicate an impor- of ENZ-induced NED, and to analyze suppression of NED and tant role of GR–MYCN–CDK5R1/2–RB1–NED signaling in NEPC growth by PARP inhibition. ENZ-induced and PARP inhibitor–suppressed NEPC. We Results: We identified an ENZ treatment–associated glu- also demonstrated efficacy for OLAþDINA combination cocorticoid receptor (GR)–MYCN–CDK5–RB1–E2F1 signal- therapy in NEPC xenograft models. Introduction abiraterone (ABI) and enzalutamide (ENZ), which maximally Hormonal therapy remains the principal treatment for meta- suppress androgen signaling, increase survival by 4 and static prostate cancer, including disease that continues to 4.8 months for patients with metastatic CRPC, respectively (1, 2). progress following initial androgen deprivation to a lethal phe- Despite the moderate benefit of these agents, their success in notype known as castration-resistant prostate cancer (CRPC). treating CRPC has been hampered by emergence of drug resis- Novel second-generation androgen signaling inhibitors, such as tance. Notably, glucocorticoid receptor (GR), a steroid receptor 1Department of Genitourinary Medical Oncology, The University of Texas MD B. Liu and L. Li contributed equally to this article. Anderson Cancer Center, Houston, Texas. 2Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Current address for Bo Liu: Department of Oncology, Tongji Hospital, Tongji Houston, Texas. 3Department of Medicine, Weill Cornell Medical College, New Medical College, Huazhong University of Science and Technology, Wuhan, York, New York. 4Englander Institute for Precision Medicine, Weill Cornell Hubei, China. Medical College and New York Presbyterian Hospital, New York, New York. 5Department of Pathology and Laboratory Medicine, Weill Cornell Medical T.C. Thompson is a lead contact. College, New York, New York. 6Institute for Computational Biomedicine, Weill Corresponding Author: Timothy C. Thompson, The University of Texas MD Cornell Medical College, New York, New York. 7Department of Medical Oncol- Anderson Cancer Center, 1515 Holcombe Boulevard, Unit 18-3, Houston, TX ogy, Dana Farber Cancer Institute, Harvard Medical School, Boston, Massachu- 77030. Phone: 713-792-9955; Fax: 713-792-9956; E-mail: setts. 8Human Oncology and Pathogenesis Program, Memorial Sloan Kettering [email protected] Cancer Center, New York, New York. 9Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, New York. 10Department of Pathology, The Clin Cancer Res 2019;XX:XX–XX University of Texas MD Anderson Cancer Center, Houston, Texas. doi: 10.1158/1078-0432.CCR-19-0317 Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). Ó2019 American Association for Cancer Research. www.aacrjournals.org OF1 Downloaded from clincancerres.aacrjournals.org on October 2, 2021. © 2019 American Association for Cancer Research. Published OnlineFirst August 22, 2019; DOI: 10.1158/1078-0432.CCR-19-0317 Liu et al. recombination model (15–17). Thus, inactivation of RB1 and Translational Relevance p53 appear to be critical for lineage plasticity—related Ad prostate Emerging evidence has revealed an association between cancer–NEPC transition. However, the mechanisms involved in hormonal treatment and neuroendocrine prostate cancer this process remain elusive. (NEPC), and the critical involvement of MYCN, and loss of PARP inhibitors are promising therapeutic agents that show RB1 function in the development of NEPC. However, the synthetic lethality against many types of cancer with BRCA1 mechanisms that underlie enzalutamide (ENZ) treatment– or BRCA2 deficiencies (18, 19). Targeting PARP together with associated neuroendocrine differentiation (NED) and the specific DNA damage response (DDR) pathways exposed development of NEPC remain to be addressed. The GR– previously unrecognized therapeutic vulnerabilities in CRPC MYCN–CDK5–RB1–E2F1 signaling pathway identified in our models (20–22). However, whether PARP inhibitors can study defines a transcriptional regulatory mechanism that specifically suppress or alter the course of NED is not known. underlies the transition from ENZ resistance to NED, in the Overall, emerging evidence has revealed an association development of NEPC. This mechanism links ENZ resistance between hormonal treatment and NEPC, and the critical involve- in prostatic adenocarcinoma to NED through GR-induced ment of MYCN and RB1 in NED of prostatic epithelial cells and MYCN, and reveals CDK5-driven inactivation of intact Rb1 the development of NEPC. However, their exact roles and the as a driver of E2F1-regulated NED gene expression. Our results mechanisms that underlie ENZ treatment—associated NED and further demonstrate that PARP inhibition suppresses the GR– the development of NEPC remain to be addressed. Our results MYCN–CDK5–RB1–E2F1 signaling pathway, and direct herein introduce a GR–MYCN–CDK5–RB1–E2F1–NED signaling MYCN and E2F1 induction of NED, and that the combination pathway, and reveal the underlying molecular mechanisms for of PARP inhibition and Rb-associated CDK inhibition may be suppression of ENZ-induced NED by PARP inhibition. a viable strategy toward effective clinical treatment of NEPC. Materials and Methods Cell lines, patient-derived xenografts, and reagents family member, was found to be overexpressed in AR signaling LNCaP C4-2b cells were maintained in RPMI1640 cell culture inhibitor resistant tumors (3). medium supplemented with 10% FBS. C4-2b-ENZR cells were Primary localized prostate cancer almost exclusively exhibits generated by long-term ENZ treatment with incremental increase adenocarcinoma (Ad) morphology, whereas neuroendocrine of ENZ concentrations in each passage (with 0.5 mmol/L intervals prostate cancer (NEPC) more commonly arises after hormonal up to 3 and 1 mmol/L intervals afterward up to 10 mmol/L). therapy (4), and is more prevalent in CRPC (5, 6). A recent The resulting C4-2b-ENZR cells were maintained in RPMI1640 prospective study demonstrated that prostate cancer from medium supplement with 10% FBS plus 10 mmol/L ENZ. NCI- patients treated with AR signaling inhibitors transitioned from H660 cells were purchased from ATCC, and maintained in Ad prostate cancer into NEPC at a higher rate than those who were suggested culture medium. MSKCC PCa4 organoids were main- not treated with these agents (7). Although platinum-based tained in complete human media as described previously (23). chemotherapy is active in NEPC, responses are short-lived and MDA prostate cancer patient-derived xenograft (PDX) 133-4 and novel therapeutic approaches are needed. MDA prostate cancer PDX 144-13C were developed in the "Pros- The phenotypic conversion from Ad prostate cancer to NEPC tate Cancer Patient Derived Xenografts Program" in the depart- has been associated with specific genetic lesions including over- ment of Genitourinary Medical Oncology, MD Anderson Cancer expression and genomic amplification of MYCN (encoding N- Center and the David H. Koch Center for Applied Research of Myc oncoprotein; refs. 5, 8). MYCN is overexpressed in the Genitourinary Cancers following published methods (24). majority of NEPC but also in up to 20% of CRPC without NEPC LNCaP C4-2b was validated by short tandem repeat DNA finger- morphology (5, 9). Furthermore, a recent study demonstrated print with the AmpFlSTR Identifier PCR Amplification Kit (Themo that MYCN could drive the initiation and maintenance of NEPC Fisher Scientific) in MD Anderson's Characterized
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