AR-positive Castration-resistant Prostate Cancer Research Articles

Patient-derived castration-resistant prostate cancer model revealed CTBP2 upregulation mediated by OCT1 and androgen receptor

BackgroundProstate cancer is dependent on androgen receptor (AR) signaling, and androgen deprivation therapy (ADT) has proven effective in targeting prostate cancer. However, castration-resistant prostate cancer (CRPC) eventually emerges. AR signaling inhibitors (ARSI) have been also used, but resistance to these agents develops due to genetic AR alterations and epigenetic dysregulation.MethodsIn this study, we investigated the role of OCT1, a member of the OCT family, in an AR-positive CRPC patient-derived xenograft established from a patient with resistance to ARSI and chemotherapy. We conducted a genome-wide analysis chromatin immunoprecipitation followed by sequencing and bioinformatic analyses using public database.ResultsGenome-wide analysis of OCT1 target genes in PDX 201.1 A revealed distinct OCT1 binding sites compared to treatment-naïve cells. Bioinformatic analyses revealed that OCT1-regulated genes were associated with cell migration and immune system regulation. In particular, C-terminal Binding Protein 2 (CTBP2), an OCT1/AR target gene, was correlated with poor prognosis and immunosuppressive effects in the tumor microenvironment. Metascape revealed that CTBP2 knockdown affects genes related to the immune response to bacteria. Furthermore, TISIDB analysis suggested the relationship between CTBP2 expression and immune cell infiltration in prostate cancer, suggesting that it may contribute to immune evasion in CRPC.ConclusionsOur findings shed light on the genome-wide network of OCT1 and AR in AR-positive CRPC and highlight the potential role of CTBP2 in immune response and tumor progression. Targeting CTBP2 may represent a promising therapeutic approach for aggressive AR-positive CRPC. Further validation will be required to explore novel therapeutic strategies for CRPC management.

Abstract 2898: Development of the lipid kinase PIKfyve PROTAC degrader against neuroendocrine prostate cancer

Abstract Therapies targeting the androgen receptor (AR) as the main driver of prostate cancer (PCa) can lead to various mechanisms of resistance and promote progression to castration-resistant PCa (CRPC), which has a median survival of only 13-23 months. Amongst recurrent CRPC, 17%-30% of patients develop neuroendocrine prostate cancer (NEPC), which is a PCa subtype characterized by a unique histology. NEPC exhibits a loss of AR signaling during neuroendocrine transdifferentiation which results in resistance to AR-targeted therapies and gain of cell characteristics resembling poorly differentiated neuroendocrine tumors. Despite advances in the understanding of NEPC development, treatment options remain limited, with platinum-based chemotherapy as the first-line treatment for both de novo and treatment-induced NEPC. However, response to first-line chemotherapy in NEPC is short, with a median survival of only seven months. The poor prognosis of NEPC is attributed in part to late diagnosis and a lack of effective therapeutic agents. Our previous work demonstrated that NEPC and AR-negative PCa exhibit higher dependency on the lipid kinase PIKfyve than AR-positive CRPC. Thus, development of PIKfyve inhibitory therapies targeting the emergent vulnerabilities of NEPC or AR-negative forms of PCa is a promising approach. Using a proteolysis targeting chimera (PROTAC) technology, we designed a class of PIKfyve specific degrader. The in vitro degradation efficiency were determined by immunoblotting, and DC50s (degradation concentration at 50%) were calculated for top candidates. Pharmacokinetic and pharmacodynamic were evaluated for in vivo study. In summary, we have developed a class of PIKfyve specific degrader using the E3 ligase von Hippel-Lindau (VHL) as a ligand via a diverse set of linkers. Mechanistic studies revealed that it induced PIKfyve degradation in a VHL- and proteasome-dependent manner. Citation Format: Yuanyuan Qiao, Chungen Li, Yang Zheng, Xia Jiang, Sarah Nicole Yee, Caleb Cheng, Yi Bao, Yuping Zhang, Yuzhuo Wang, Zhen Wang, Ke Ding, Arul Chinnaiyan. Development of the lipid kinase PIKfyve PROTAC degrader against neuroendocrine prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2898.

Abstract 7222: Antiandrogen equipped with HDAC inhibitors activates autophagy-mediated growth inhibition in castrate-resistant prostate cancer

Abstract Targeting androgen receptor (AR) by pharmacologic intervention is one of the practical approaches for the treatment of castration-resistant prostate cancer (CRPC). Hence, this study aimed to develop novel molecules that employ multiple mechanisms to inhibit AR expression and curtail the growth of CRPC. The target antiandrogen-equipped histone deacetylase inhibitors were synthesized, adapting the convergent chemistry we used to synthesize the first-generation compounds. The final compounds were characterized using 1H-NMR, 13C-NMR, and high-resolution mass spectrometry. We synthesized and analyzed several compounds, in which KK62 emerged as the lead compound. KK62 significantly inhibits a panel of CRPC cell lines in nanomolar concentrations without causing toxicity to healthy prostate epithelial cells. The molecular analysis suggested that inhibition of AR and AR-splice variants was evident, followed by downregulation of PSA expression in AR-positive CRPC cell lines. Further analysis confirmed that KK62 overcomes dihydrotestosterone (DHT) induced AR signaling and enzalutamide-resistant CRPC cells by downregulating AR and AR-splice variants in CRPC cells. Subsequent analysis confirmed that KK62 induces autophagy signaling by upregulating autophagosome (LC3B) and lysosome (LAMP1) markers expression and facilitating the autophagosome and lysosome fusion, which resulted in the induction of apoptosis in all AR-positive CRPC cell lines. The pro-apoptotic markers, such as BAX and cleaved caspase, were upregulated, whereas the downregulation of pro-survival markers was evident in KK62-treated CRPC cells. Our ongoing castrated and non-castrated xenotransplanted CRPC tumors and Patient-derived xenograft in vivo studies would validate our in vitro findings and confirm that KK62 is a potent compound that may improve therapeutic advances in treating CRPC. Citation Format: Balaji Chandrasekaran, Kiran Kumar Yalla, Ashish Tyagi, Vaibhav Shukla, Neha Tyagi, Bhawna Tyagi, Mohit Vashishta, Adegboyega K Oyelere, Chendil Damodaran. Antiandrogen equipped with HDAC inhibitors activates autophagy-mediated growth inhibition in castrate-resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7222.

Targeting the fibroblast growth factor pathway in molecular subtypes of castration-resistant prostate cancer.

Androgen receptor (AR) pathway inhibition remains the cornerstone for prostate cancer therapies. However, castration-resistant prostate cancer (CRPC) tumors can resist AR signaling inhibitors through AR amplification and AR splice variants in AR-positive CRPC (ARPC), and conversion to AR-null phenotypes, such as double-negative prostate cancer (DNPC) and small cell or neuroendocrine prostate cancer (SCNPC). We have shown previously that DNPC can bypass AR-dependence through fibroblast growth factor receptor (FGFR) signaling. However, the role of the FGFR pathway in other CRPC phenotypes has not been elucidated. RNA-Seq analysis was conducted on patient metastases, LuCaP patient-derived xenograft (PDX) models, and CRPC cell lines. Cell lines (C4-2B, VCaP, and 22Rv1) and ex vivo LuCaP PDX tumor cells were treated with enzalutamide (ENZA) and FGFR inhibitors (FGFRi) alone or in combination and sensitivity was determined using cell viability assays. In vivo efficacy of FGFRi in ARPC, DNPC, and SCNPC were evaluated using PDX models. RNA-Seq analysis of FGFR signaling in metastatic specimens, LuCaP PDX models, and CRPC cell lines revealed significant FGF pathway activation in AR-low PC (ARLPC), DNPC, and SCNPC tumors. In vitro/ex vivo analysis of erdafitinib and CH5183284 demonstrated robust and moderate growth suppression of ARPC, respectively. In vivo studies using four ARPC PDX models showed that combination ENZA and CH5183284 significantly suppressed tumor growth. Additional in vivo studies using four ARPC PDX models revealed that erdafitinib monotherapy was as effective as ENZA in suppressing tumor growth, and there was limited combination benefit. Furthermore, two of three DNPC models and two of four SCNPC models responded to CH5183284 monotherapy, suggesting FGFRi responses were model dependent. RNA-Seq and gene set enrichment analysis of end-of-study ARPC tumors treated with FGFRi displayed decreased expression of E2F and MYC target genes and suppressed G2M checkpoint genes, whereas end-of-study SCNPC tumors had heterogeneous transcriptional responses. Although FGFRi treatments suppressed tumor growth across CRPC phenotypes, our analyses did not identify a single pathway or biomarker that would identify tumor response to FGFRi. This is very likely due to the array of FGFR1-4 expression and tumor phenotypes present in CRPC. Nevertheless, our data nominate the FGFR pathway as a clinically actionable target that promotes tumor growth in diverse phenotypes of treatment-refractory metastatic CRPC.

Synthesis and evaluation of 7-(3-aminopropyloxy)-substituted flavone analogue as a topoisomerase IIα catalytic inhibitor and its sensitizing effect to enzalutamide in castration-resistant prostate cancer cells

Prostate cancer patients primarily receive androgen receptor (AR)-targeted drugs as a primary treatment option because prostate cancer is associated with highly activated AR signaling. AR amplification made prostate cancer cells viable under treatment of AR-targeted therapy, leading to castration resistance. AR amplification was more common in enzalutamide-resistant patients. As a strategy to overcome castration resistance and to improve the efficacy of enzalutamide, second-generation nonsteroidal antiandrogen drugs for castration-resistant prostate cancer (CRPC) including topoisomerase II (topo II) poisons such as etoposide and mitoxantrone, have been administered in combination with enzalutamide. In the present study, it was confirmed that amplification of topo IIα, but not I and IIβ, was directly and proportionally associated with poor clinical outcome of Prostate cancer. Among a novel series of newly designed and synthesized 7-(3-aminopropyloxy)-substituted flavone analogues, compound 6, the most potent derivative, was further characterized and identified as a topo IIα catalytic inhibitor that intercalates into DNA and binds to the DNA minor groove with better efficacy and less genotoxicity than etoposide, a topo II poison. Compound 6 showed remarkable efficacy in inhibiting AR-negative CRPC cell growth and sensitizing activity to enzalutamide in AR-positive CRPC cells, thus confirming the potential of topo IIα catalytic inhibitor to overcome resistance to androgen deprivation therapy.

Novel Dual BET and PLK1 Inhibitor WNY0824 Exerts Potent Antitumor Effects in CRPC by Inhibiting Transcription Factor Function and Inducing Mitotic Abnormality.

Castration-resistant prostate cancer (CRPC) is a lethal disease with few treatment alternatives once patients become resistant to second-generation antiandrogens. In CRPC, BET proteins are key regulators of AR- and MYC-mediated transcription, while the PLK1 inhibitor potentially downregulates AR and MYC besides influencing the cell cycle. Therefore, synchronous inhibition of BET and PLK1 would be a promising approach for CRPC therapy. This study developed a dual BET and PLK1 inhibitor WNY0824 with nanomolar and equipotent inhibition of BRD4 and PLK1. In vitro, WNY0824 exhibited excellent antiproliferation activity on AR-positive CRPC cells and induced apoptosis. These activities are attributable to its disruption of the AR-transcriptional program and the inhibition of the ETS pathway. Furthermore, WNY0824 downregulated MYC and induced mitotic abnormality. In vivo, oral WNY0824 administration suppressed tumor growth in the CRPC xenograft model of enzalutamide resistance. These findings suggest that WNY0824 is a selective dual BET and PLK1 inhibitor with potent anti-CRPC oncogenic activity and provides insights into the development of other novel dual BET- and PLK1-inhibiting drugs.

Gene therapy for castration-resistant prostate cancer cells using JC polyomavirus-like particles packaged with a PSA promoter driven-suicide gene.

Prostate cancer is the second most common cancer in men globally. Prostate cancer patients at advanced stages are usually treated with androgen deprivation therapy (ADT). However, with disease progression, it often becomes the incurable castration-resistant prostate cancer (CRPC). JC polyomavirus (JCPyV) is a human DNA virus. Its virus-like particles (VLPs) exhibit similar tropism to native virions and they are capable of delivering exogenous genes to the target cells for expression. JCPyV has been detected in prostate cells; therefore, prostate cancer cells may be susceptible to JCPyV infection and JCPyV VLPs may be used as a vector for gene therapy against prostate cancer. Here we constructed a plasmid (pPSAtk) that allows expression of the thymidine kinase suicide gene only in androgen receptor (AR) positive prostate cancer cells using the prostate-specific antigen (PSA) promoter, and used JCPyV VLPs as a vector to carry pPSAtk (PSAtk-VLPs) for transcriptional targeting in prostate cancer cells. In this study, we found that PSAtk-VLPs could only kill AR-positive CRPC 22Rv1 cells in vitro and inhibit the growth of tumor nodules in the xenograft mouse model. Our results reveal that PSAtk-VLPs could potentially be used as a new option for treating CRPC patients in the future.

Abstract 3837: Bumped kinase inhibitor 1553 selectively inhibits androgen receptor positive prostate cancer

Abstract BACKGROUND: Prostate cancer (PCa) is the second most common cancer in men and one of the leading causes of cancer death. Resistance to current PCa therapies, including androgen deprivation, occurs in almost all patients leading to development of castration resistant prostate cancer (CRPC). Resistance is associated with expression of splice variants of the androgen receptor (AR-Vs) that are constitutively active. The intrinsically disordered N-terminus of the AR/AR-Vs makes targeting the AR-Vs difficult. Thus, therapies that indirectly target the AR by inhibiting factors important in regulating AR levels and activity may be the most successful against inhibiting the constitutively active AR variants. Kinases have been shown to be important in regulating the AR, thus kinase inhibitors have the potential to inhibit androgen receptor signaling and function. OBJECTIVE: To demonstrate that our lead kinase inhibitor targets and inhibits AR positive CRPC. RESULTS: Bumped kinase inhibitors designed at the University of Washington are based on a pyrazolopyrimidine backbone, with R1 and R2 groups that confer a “bump” thereby preventing the BKIs from binding to most human kinase ATP binding pockets. While the majority of BKIs do not have activity against CRPC, a subset have activity against AR+ PCa lines, including CRPC lines expressing AR-Vs, but with no activity against AR negative PCa lines or a broad range of other human cell lines. We next examined the effects of our lead BKI compound (1553) on AR expression, phosphorylation, and function. BKI-1553 blocked transactivation activity of AR as judged by the reporter assays using an ARE-luciferase construct. Expression of canonical AR target genes was downregulated in LNCaP cells treated with BKI-1553. Westerns demonstrated a decrease in total AR 4 and 24 hours after treatment with BKI-1553. Levels of phospho-Serine81on the AR, which is critical for AR activity, were decreased even further than total AR by 4 hours after treatment. Thus, AR signaling appears to constitute BKI-sensitive components. We then decided to examine the effect of BKI-1553 on in vivo tumor growth. Toxicity studies demonstrated that it was non-toxic in mice at oral doses that achieved therapeutic levels. In vivo treatment with BKI-1553 significantly decreased growth of a castrate-sensitive patient-derived PCa xenograft and a castration-resistant PCa cell line (LuCaP35, p < 0.01 and LNCaP95, p < 0.005, respectively). SUMMARY: Our lead bumped kinase inhibitor, BKI-1553, selectively inhibits the growth of prostate cancer cells that express full-length AR and/or AR-Vs. BKI-1553 reduced not only cell proliferation, but decreased levels of AR and AR variants, decreased pSer81 levels, and decreased activation of AR-regulated promoters. Since the majority of CRPCs are AR driven, our targeted BKIs could be a novel therapy for cancers that are resistant to currently available therapies. Citation Format: Takuma Uo, Shihua Sun, Kathleen Haugk, Kathryn Soriano Epilepsia, Mamatha Damodarasamy, Matthew Hulverson, Wesley VanVoorhis, Kayode K. Ojo, RamaSubbaRao Vidadala, Dustin Maly, Stephen Plymate, Cynthia C. Sprenger. Bumped kinase inhibitor 1553 selectively inhibits androgen receptor positive prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3837.

The expression of AURKA is androgen regulated in castration-resistant prostate cancer

Although second generation endocrine therapies have significantly improved survival, castration-resistant prostate cancer (CRPC) cells are eventually able to escape available hormonal treatments due to reactivation of androgen receptor (AR) signaling. Identification of novel, non-classical and druggable AR-target genes may provide new approaches to treat CRPC. Our previous analyses suggested that Aurora kinase A (AURKA) is regulated by androgens in prostate cancer cells that express high levels of AR. Here, we provide further evidence that AURKA is significantly overexpressed in AR-positive CRPC samples carrying amplification of AR gene and/or expressing AR in high levels. We also demonstrate androgen-induced AR binding in the intronic region of AURKA. The expression of AURKA is increased upon androgen stimulation in LNCaP-ARhi cells that express high levels of AR. The growth of the cells was also significantly inhibited by an AURKA specific inhibitor, alisertib (MLN8237). Together, these findings suggest that the expression of AURKA is regulated by androgen in prostate cancer cells that highly express AR, emphasizing its potential as a therapeutic target in patients with CRPC.

Abstract 3844: Targeting EZH2 in neuroendocrine prostate cancer

Abstract Background and aim of the study EZH2 is the catalytic subunit of the Polycomb repressive complex 2 (PRC2) and it is involved in controlling gene expression through trimethylation of histone H3 lysine 27 (H3K27me3), a key mechanism that regulates chromatin structure and gene silencing. EZH2 is overexpressed in a wide range of cancers including prostate. Recent findings show that EZH2 can form a complex with the androgen receptor (AR), enhancing its transcriptional activity. However the role of EZH2 in AR low/negative prostate cancer has not been previously investigated. Long-term exposure to AR targeted therapies can cause an adaptive response mechanism in a subset of advanced prostate cancers. Often this adaptation is associated with loss of AR expression, clinical aggressiveness, and pathologic features of small cell or neuroendocrine carcinoma. This AR independent resistant phenotype is broadly termed neuroendocrine prostate cancer (NEPC). Initial studies from our lab have highlighted a landscape of genomic alterations that characterize NEPC. In this study we explore the hypothesis that the epigenetic modifier, EZH2, can lead the adaptive response towards an androgen-independent phenotype and towards NEPC. Results Using Next Generation RNA-sequencing, we profiled tumors from a cohort of patients including 20 NEPC, 68 Prostate Adenocarcinoma (PCa), and 19 castration resistant prostate cancer (CRPC) and we discovered that EZH2 is highly expressed in NEPC (characterized by low or absent AR and morphologic changes). These findings were also confirmed at protein level by immunohistochemistry. Based on the driving role of EZH2 in other tumor types and significant overexpression in NEPC tumors, we evaluated the effects of the EZH2 inhibitor, GSK343, in NEPC cells (NCI-H660) and PCa cells (LNCaP and DU145) in 3D Matrigel cultures. GSK343 effectively inhibited H3K27me3 and resulted in a significant reduction of NCI-H660 viability, measured with an ATP-based assay, whereas LNCaP and DU145 cells were minimally affected after 7 or 14 days of treatment. Using a custom designed Nanostring assay, we also demonstrated a significant increase in the expression of AR signaling genes (e.g. PSA, PSMA) and decrease in NEPC-associated genes (e.g. chromogranin A, AURKA, ENO2) when NEPC cells were treated with GSK343. These data suggest a modulation of the neuroendocrine phenotype via EZH2. We therefore extended these drug studies to patient-derived organoid models including both CRPC and NEPC organoids and we observed similar results with preferential sensitivity of the AR-negative NEPC organoids compared to AR-positive CRPC organoids and similar reversion of downstream AR/NEPC gene expression. Conclusions Altogether, these data suggest that EZH2-mediated epigenetic changes contribute to the AR-independent NEPC resistant phenotype, and this effect can be reverted or delayed by targeting EZH2 activity. EZH2 represents a promising drug target and a potential modulator of the NEPC phenotype. Citation Format: Loredana Puca, Dong Gao, Myriam Kossai, Clarisse Marotz, Juan Miguel Mosquera, Theresa Y. MacDonald, Kyung Park, Rema Rao, Andrea Sboner, Yu Chen, Mark A. Rubin, Himisha Beltran. Targeting EZH2 in neuroendocrine prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3844. doi:10.1158/1538-7445.AM2015-3844

Androgen Suppresses the Proliferation of Androgen Receptor-Positive Castration-Resistant Prostate Cancer Cells via Inhibition of Cdk2, CyclinA, and Skp2

The majority of prostate cancer (PCa) patient receiving androgen ablation therapy eventually develop castration-resistant prostate cancer (CRPC). We previously reported that androgen treatment suppresses Skp2 and c-Myc through androgen receptor (AR) and induced G1 cell cycle arrest in androgen-independent LNCaP 104-R2 cells, a late stage CRPC cell line model. However, the mechanism of androgenic regulation of Skp2 in CRPC cells was not fully understood. In this study, we investigated the androgenic regulation of Skp2 in two AR-positive CRPC cell line models, the LNCaP 104-R1 and PC-3AR Cells. The former one is an early stage androgen-independent LNCaP cells, while the later one is PC-3 cells re-expressing either wild type AR or mutant LNCaP AR. Proliferation of LNCaP 104-R1 and PC-3AR cells is not dependent on but is suppressed by androgen. We observed in this study that androgen treatment reduced protein expression of Cdk2, Cdk7, Cyclin A, cyclin H, Skp2, c-Myc, and E2F-1; lessened phosphorylation of Thr14, Tyr15, and Thr160 on Cdk2; decreased activity of Cdk2; induced protein level of p27Kip1; and caused G1 cell cycle arrest in LNCaP 104-R1 cells and PC-3AR cells. Overexpression of Skp2 protein in LNCaP 104-R1 or PC-3AR cells partially blocked accumulation of p27Kip1 and increased Cdk2 activity under androgen treatment, which partially blocked the androgenic suppressive effects on proliferation and cell cycle. Analyzing on-line gene array data of 214 normal and PCa samples indicated that gene expression of Skp2, Cdk2, and cyclin A positively correlates to each other, while Cdk7 negatively correlates to these genes. These observations suggested that androgen suppresses the proliferation of CRPC cells partially through inhibition of Cyclin A, Cdk2, and Skp2.

Abstract 2966: The role of histone demethylase GASC1 in promoting prostate cancer progression.

Abstract Prostate cancer affects 1 in 6 American men, and is the second leading cause of cancer-related deaths in males. The defiance of castration-resistant prostate cancer (CRPC) to currently available treatments represents the most challenging aspect of this disease. Although resistant to androgen deprivation treatment, most CRPCs continue to express the androgen receptor (AR) and remain dependent on AR signaling for growth and proliferation. Previously, we cloned a novel gene, GASC1 (gene amplified in squamous cell carcinoma 1), from 9p24 amplified cancer cells. Recently, the GASC1 protein has been identified as a key histone demethylase that controls chromatin-mediated regulation of gene expression. Importantly, the GASC1 protein interacts with the AR and serves as a co-activator of AR-induced transcription in prostate cells. In this study, we demonstrated that various aggressive tumors, including CRPCs, have upregulated GASC1. We used an shRNA approach to determine whether GASC1 knockdown would affect the proliferation and transformation of CRPC cell lines. We found that knocking down GASC1 inhibits growth and colony formation of CRPC cells (C4-2B and CWR22Rv1) in vitro. Previously, it was reported that the GASC1 protein interacts with the AR and functions as a co-activator of AR-induced transcription in LNCaP cells. The GASC1 protein contains N-terminal Jumonji catalytic domains and C-terminal Tudor and PHD domains; however, which domains are responsible for the association of GASC1 with the AR have not yet been identified. To address this question, we generated stable LNCaP cell lines expressing V5-tagged N-terminal Jumonji catalytic domains and C-Terminal PHD/Tudor domains, and performed Co-IP analyses. We found that the C-terminal region is essential for GASC1 association with the AR in prostate cancer cells. To explore the contribution of GASC1 to endogenous AR-regulated gene expression in AR-positive CRPC cells, we performed quantitative RT-PCR assays to measure the expression of both classical AR target genes (including PSA/KLK3, KLK2, FKBP5, and NDRG1) and CRPC-specific genes (including UBE2C, CDK1, CDC20, and CCNA2) in C4-2B cells with or without GASC1 knockdown. We found that knocking down GASC1 significantly reduced transcript levels of a set of classical and CRPC-specific AR target genes in either the presence or absence of the AR ligand. Our data demonstrates that GASC1 is a target of substantial relevance to aggressive prostate cancers, including CRPCs. As such, targeting GASC1 is a promising strategy to control CRPC or prevent its emergence. Citation Format: Lihong Zhang, Jinling Hou, Andreana Holowatyj, Zeng-Quan Yang. The role of histone demethylase GASC1 in promoting prostate cancer progression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2966. doi:10.1158/1538-7445.AM2013-2966

CCI-779 Inhibits Cell-Cycle G2–M Progression and Invasion of Castration-Resistant Prostate Cancer via Attenuation of UBE2C Transcription and mRNA Stability

The cell-cycle G(2)-M phase gene UBE2C is overexpressed in various solid tumors including castration-resistant prostate cancer (CRPC). Our recent studies found UBE2C to be a CRPC-specific androgen receptor (AR) target gene that is necessary for CRPC growth, providing a potential novel target for therapeutic intervention. In this study, we showed that the G(1)-S cell-cycle inhibitor-779 (CCI-779), an mTOR inhibitor, inhibited UBE2C mRNA and protein expression in AR-positive CRPC cell models abl and C4-2B. Treatment with CCI-779 significantly decreased abl cell proliferation in vitro and in vivo through inhibition of cell-cycle progression of both G(2)-M and G(1)-S phases. In addition, exposure of abl and C4-2B cells to CCI-779 also decreased UBE2C-dependent cell invasion. The molecular mechanisms for CCI-779 inhibition of UBE2C gene expression involved a decreased binding of AR coactivators SRC1, SRC3, p300, and MED1 to the UBE2C enhancers, leading to a reduction in RNA polymerase II loading to the UBE2C promoter, and attenuation of UBE2C mRNA stability. Our data suggest that, in addition to its ability to block cell-cycle G(1) to S-phase transition, CCI-779 causes a cell-cycle G(2)-M accumulation and an inhibition of cell invasion through a novel UBE2C-dependent mechanism, which contributes to antitumor activities of CCI-779 in UBE2C overexpressed AR-positive CRPC.