Castration-resistant Prostate Cancer Growth Research Articles

The LAT1 inhibitor JPH203 suppresses the growth of castration-resistant prostate cancer through a CD24-mediated mechanism.

L-type amino acid transporter 1 (LAT1) is specifically expressed in many malignancies, contributes to the transport of essential amino acids, such as leucine, and regulates the mammalian target of rapamycin (mTOR) signaling pathway. We investigated the expression profile and functional role of LAT1 in prostate cancer using JPH203, a specific inhibitor of LAT1. LAT1 was highly expressed in castration-resistant prostate cancer (CRPC) cells, including C4-2 and PC-3 cells, but its expression level was low in castration-sensitive LNCaP cells. JPH203 significantly inhibited [14C] leucine uptake in CRPC cells but had no effect in LNCaP cells. JPH203 inhibited the proliferation, migration, and invasion of CRPC cells but not of LNCaP cells. In C4-2 cells, Cluster of differentiation (CD) 24 was identified by RNA sequencing as a novel downstream target of JPH203. CD24 was downregulated in a JPH203 concentration-dependent manner and suppressed activation of the Wnt/β-catenin signaling pathway. Furthermore, an invivo study showed that JPH203 inhibited the proliferation of C4-2 cells in a castration environment. The results of this study indicate that JPH203 may exert its antitumor effect in CRPC cells via mTOR and CD24.

Microwave hyperthermia enhances radiosensitization by decreasing DNA repair efficiency and inducing oxidative stress in PC3 prostatic adenocarcinoma cells

Purpose Radiotherapy (RT) is the primary treatment for prostate cancer (PCa); however, the emergence of castration-resistant prostate cancer (CRPC) often leads to treatment failure and cancer-related deaths. In this study, we aimed to explore the use of microwave hyperthermia (MW-HT) to sensitize PCa to RT and investigate the underlying molecular mechanisms. Methods We developed a dedicated MW-HT heating setup, created an in vitro and in vivo MW-HT + RT treatment model for CRPC. We evaluated PC3 cell proliferation using CCK-8, colony experiments, DAPI staining, comet assay and ROS detection method. We also monitored nude mouse models of PCa during treatment, measured tumor weight, and calculated the tumor inhibition rate. Western blotting was used to detect DNA damage repair protein expression in PC3 cells and transplanted tumors. Results Compared to control, PC3 cell survival and clone formation rates decreased in RT + MW-HT group, demonstrating significant increase in apoptosis, ROS levels, and DNA damage. Lower tumor volumes and weights were observed in treatment groups. Ki-67 expression level was reduced in all treatment groups, with significant decrease in RT + MW-HT groups. The most significant apoptosis induction was confirmed in RT + MW-HT group by TUNEL staining. Protein expression levels of DNA-PKcs, ATM, ATR, and P53/P21 signaling pathways significantly decreased in RT + MW-HT groups. Conclusion MW-HT + RT treatment significantly inhibited DNA damage repair by downregulating DNA-PKcs, ATM, ATR, and P53/P21 signaling pathways, leading to increased ROS levels, aggravate DNA damage, apoptosis, and necrosis in PC3 cells, a well-established model of CRPC.

Identification of an anaplastic subtype of prostate cancer amenable to therapies targeting SP1 or translation elongation.

Histopathological heterogeneity is a hallmark of prostate cancer (PCa). Using spatial and parallel single-nucleus transcriptomics, we report an androgen receptor (AR)-positive but neuroendocrine-null primary PCa subtype with morphologic and molecular characteristics of small cell carcinoma. Such small cell-like PCa (SCLPC) is clinically aggressive with low AR, but high stemness and proliferation, activity. Molecular characterization prioritizes protein translation, represented by up-regulation of many ribosomal protein genes, and SP1, a transcriptional factor that drives SCLPC phenotype and overexpresses in castration-resistant PCa (CRPC), as two potential therapeutic targets in AR-indifferent CRPC. An SP1-specific inhibitor, plicamycin, effectively suppresses CRPC growth in vivo. Homoharringtonine, a Food And Drug Administration-approved translation elongation inhibitor, impedes CRPC progression in preclinical models and patients with CRPC. We construct an SCLPC-specific signature capable of stratifying patients for drug selectivity. Our studies reveal the existence of SCLPC in admixed PCa pathology, which may mediate tumor relapse, and establish SP1 and translation elongation as actionable therapeutic targets for CRPC.

Mediator kinase inhibition reverses castration resistance of advanced prostate cancer.

Mediator kinases CDK19 and CDK8, pleiotropic regulators of transcriptional reprogramming, are differentially regulated by androgen signaling, but both kinases are upregulated in castration-resistant prostate cancer (CRPC). Genetic or pharmacological inhibition of CDK8 and CDK19 reverses the castration-resistant phenotype and restores the sensitivity of CRPC xenografts to androgen deprivation in vivo. Prolonged CDK8/19 inhibitor treatment combined with castration not only suppressed the growth of CRPC xenografts but also induced tumor regression and cures. Transcriptomic analysis revealed that Mediator kinase inhibition amplified and modulated the effects of castration on gene expression, disrupting CRPC adaptation to androgen deprivation. Mediator kinase inactivation in tumor cells also affected stromal gene expression, indicating that Mediator kinase activity in CRPC molded the tumor microenvironment. The combination of castration and Mediator kinase inhibition downregulated the MYC pathway, and Mediator kinase inhibition suppressed a MYC-driven CRPC tumor model even without castration. CDK8/19 inhibitors showed efficacy in patient-derived xenograft models of CRPC, and a gene signature of Mediator kinase activity correlated with tumor progression and overall survival in clinical samples of metastatic CRPC. These results indicate that Mediator kinases mediated androgen-independent in vivo growth of CRPC, supporting the development of CDK8/19 inhibitors for the treatment of this presently incurable disease.

Combined therapy targeting AR and EZH2 curbs castration-resistant prostate cancer enhancing anti-tumor T-cell response.

Aim: Castration-resistant prostate cancer (CRPC) eventually becomes resistant to androgen receptor pathway inhibitors like enzalutamide. Immunotherapy also fails in CRPC. We propose a new approach to simultaneously revert enzalutamide resistance and rewire anti-tumor immunity. Methods: We investigated in vitro and in subcutaneous and spontaneous mouse models the effects of combining enzalutamide and GSK-126, a drug inhibiting the epigenetic modulator EZH2. Results: Enzalutamide and GSK-126 synergized to reduce CRPC growth, also restraining tumor neuroendocrine differentiation. The anti-tumor activity was lost in immunodeficient mice. Indeed, the combination treatment awoke cytotoxic activity and IFN-γ production of tumor-specific CD8+ T lymphocytes. Conclusion: These results promote the combination of enzalutamide and GSK-126 in CRPC, also offering new avenues for immunotherapy in prostate cancer.

Abstract 973: Plasma androgen receptor (AR) copy number gain at progression in patients randomized in the STAMPEDE phase 3 abiraterone acetate, prednisolone (AAP) and enzalutamide (ENZ) trial: An ancillary biomarker study

Abstract Addition of ENZ or AAP to androgen deprivation therapy (ADT) as first-line treatment for advanced prostate cancer improves survival compared to ADT alone. Genomic alterations in AR associate with resistance to ENZ/AAP when started for castration-resistant prostate cancer (CRPC). We aimed to test whether early use of AAP/ENZ altered the incidence of AR alterations at progression. Between July 2014 and March 2016, 1976 patients with metastatic or high-risk locally advanced prostate cancer were randomized to ADT or ADT+ENZ+AAP in the STAMPEDE trial (NCT00268476). Consenting patients were given the option to donate blood for the ancillary biomarker study at trial-defined progression and prior to start of the next line of treatment. We focused on patients who progressed within 4 years from the end of recruitment (early progressors), as these patients represent the greatest unmet need. We obtained plasma DNA at emergence of CRPC from 115 early progressors (ADT n=63, ADT+ENZ+AAP n=52). We performed liquid biopsy-focused, custom, high-coverage targeted enrichment sequencing (PCF-SELECT, Orlando et al, NAR Cancer 2022). We used a bespoke computational pipeline, leveraging allelic imbalance and fragmentome analysis, to detect circulating tumor DNA (ctDNA) in plasma samples and copy number changes. Of 108 samples that passed quality control, 94 were positive for tumor (ADT n=52, ADT+AAP+ENZ n=42). The detection rate and ctDNA fractions were similar between the two treatment groups [ADT: 86.7% (52/60), mean ctDNA fraction 0.18; ADT+ENZ+AAP: 87.5% (42/48), mean ctDNA fraction 0.21; p=0.34]. The proportion of plasma with detectable ctDNA and AR gain was significantly higher with ADT+AAP+ENZ vs ADT alone (15/42, 35.7% vs 8/52, 15.4%, p=0.03). We then implemented the same approach on plasma collected from patients progressing on ENZ started for metastatic CRPC in a prospective Phase 3b trial (PRESIDE, NCT02288247): we confirmed a high prevalence of AR gain in plasma samples with detectable ctDNA (47.7%, 95% CI 38.8-54.9) in the CRPC setting. Significant copy number aberrations in relevant prostate cancer-related genes/pathways (deletions of TP53, RB1, PTEN or DNA repair genes, or gain of PI3K/AKT1) were identified in 9 out of 42 evaluable samples in the ADT group and 18 out of 38 evaluable samples in the ADT+ENZ+AAP group (21.4% vs 47.4%, p=0.014). Treatment with ADT+ENZ+AAP increases detection of plasma AR gain at emergences of CRPC in early progressors compared to ADT alone. Detection of plasma AR gain increases at progression on ADT+ENZ regardless of the treatment setting. The impact of early use of ENZ/AAP on the detection of copy number aberrations, especially homozygous deletions, in other prostate cancer-related genes in plasma requires further investigation. Citation Format: Gianmarco Leone, Francesco Orlando, Suparna Thakali, Sharanpreet Lall, Daniel Wetterskog, Anna Wingate, Noel W. Clarke, Nicholas D. James, Francesca Demichelis, Gerhardt Attard. Plasma androgen receptor (AR) copy number gain at progression in patients randomized in the STAMPEDE phase 3 abiraterone acetate, prednisolone (AAP) and enzalutamide (ENZ) trial: An ancillary biomarker study [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 973.

Abstract 6048: Targeting AR positive prostate cancer cells by the novel P300/CBP oral available PROTAC degrader

Abstract Background The Androgen Receptor (AR) serves as the core lineage-specific transcription factor for prostatic epithelial cells. Targeting AR is currently the most promising therapeutic strategy. However, the inevitable development of acquired resistance to anti-androgen treatments underscores the urgent need to devise new therapeutic strategies. Recent studies have revealed that in prostate cancer cells, AR is hijacked by oncogenic transcription factors, such as FOXA1 and HOXB13, which promiscuously bind to distinct enhancer sites, driving the progression of prostate cancer. This process leads to the formation of a substantially rewired AR cancer specific enhanceosome, requiring a variety of coregulators to exhibit oncogenic driver characteristics, which can be potential therapeutic targets for prostate cancer. Histone acetyltransferases (HATs) P300 and CBP are paralogs which play dominant roles in orchestrating high-order chromatin structures and transcription factor complexes by acetylating histone proteins at multiple residuals. Although accumulating evidence suggests that P300/CBP function as AR coregulator and targeting P300/CBP can markedly suppress AR-positive prostate cancer cell growth, including growth of castration-resistant prostate cancer (CRPC), most of the P300/CBP inhibitors have performed poorly in clinical trials. Methods We developed a novel P300/CBP dual oral available PROTAC degrader, namely CBPD-409. The on-target effects of CBPD-409 were assessed by quantitative proteomics (TMT-mass spectrometry) in prostate cancer cell lines. The RNA-seq analysis was conducted to investigate CBPD-409 effects on global transcriptome in prostate cancer cells. The cytotoxicity of CBPD-409 in multiple prostate cancer cell lines was examined by cell-titer-glo and incucyte assays. Conclusion In multiple cell lines, CBPD-409 exhibits remarked potency to degrade both P300 and CBP proteins in time and does dependent manner, without evident off-target effects. By employing CBPD-409, we demonstrate that degrading P300/CBP preferentially suppress AR+ prostate cancer cell growth. The global transcriptomic profiling in CBPD-409 treated prostate cancer cells revealed that loss of P300/CBP results in remarkedly downregulation of AR, Myc, FOXA1 and ERG signaling. Assessment of CBPD-409 cytotoxicity in a panel prostate cancer cell lines suggests that AR positive prostate cancer cells display highest sensitivity to CBPD-409. Citation Format: Jie luo, Abhijit Prolia, Yuanyuan Qiao, Jean Tian, Rahul Mannan, Somnath Mahapatra, Zhixiang Chen, Rithvik Seri, Shaomeng Wang, Arul Chinnaiyan. Targeting AR positive prostate cancer cells by the novel P300/CBP oral available PROTAC degrader [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 6048.

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.

Abstract 7116: Targeting the CBP/p300 axis in lethal prostate cancer impacts DNA repair

Abstract Prostate cancer (PCa) is the 2nd leading cause of cancer related deaths in US men. PCa is an androgen dependent disease driven by the androgen receptor (AR). Currently, androgen-deprivation therapy (ADT) is the standard of care first-line therapy for metastatic PCa. Resistance to ADT uniformly leads to lethal disease, termed castration-resistant prostate cancer (CRPC). Thus, there is an unmet clinical need to identify and develop novel strategies to treat CRPC. CBP/p300 are potent co-activators for AR. High expression of CBP/p300 is associated with locally advanced disease and castration resistant function of AR, resulting in poor patient outcomes and further highlighting the need to discern the role of CBP/p300 to potentially develop therapeutic targets for precision medicine. Previous studies have relied on non-specific compounds and genetic silencing to target CBP/p300. In this study, CBP/p300 mediated bromodomain activity is targeted by CCS1477 (inobrodib), a first-in-class bromodomain inhibitor developed by Cell Centric. CCS1477 treatment demonstrated effective inhibition in growth and clonogenicity assays. Inhibition of the CBP/p300 bromodomain with CCS1477 resulted in significant downregulation of AR-FL, AR-V7, and its targets’ mRNA expression in addition to inhibition of associated factors such as c-MYC and its downstream targets in PCa cell lines as well as patient derived xenograft (PDX) models. This study shows that CBP and p300 are highly expressed and correlate closely with AR gene expression and AR activity score in primary PCa and CRPC patient samples. The clinical significance of CBP/p300 expression in PCa has been investigated via elucidation of CBP/p300 transcriptional reprogramming and its role in DNA damage response (DDR) pathways. Specifically, findings revealed that CBP/p300 bromodomain suppression sensitizes to AR-dependent DNA-repair. Transcriptional mapping identified CBP/p300 as regulators of cell proliferation and DNA repair processes, which were functionally confirmed across several PCa model systems. To assess relevance, exogenous challenge with radiation revealed that CBP/p300 are required for AR-mediated DNA repair, and CBP/p300 expression is linked to DNA repair capacity in the clinical setting. Molecular analyses revealed that CBP/p300 facilitate double-strand break (DSB) repair efficiency via homologous recombination (HR) mediated DDR. Congruently, CBP/p300 strongly correlated with HR gene expression in PCa patient tissue. These collective findings reveal that CBP/p300 govern repair of DNA DSBs by regulating HR, thus modulating genome integrity and promoting CRPC growth. These studies identify CBP/p300 as a driver of PCa tumorigenesis through coordinated control of critical transcriptional events and lay the groundwork to optimize therapeutic strategies for advanced PCa via CBP/p300 inhibition, potentially in combination with AR-directed therapies. Citation Format: Sumaira Sardar, Lakshmi Ravindranath, Christopher McNair, Saswati N. Chand, Wei Yuan, Denisa Bogdan, Jonathan Welti, Adam Sharp, Matthew Schiewer, Lisa Butler, Johann de Bono, Kris Frese, Nigel Brooks, Neil Pegg, Karen Knudsen, Ayesha A. Shafi. Targeting the CBP/p300 axis in lethal prostate cancer impacts DNA repair [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 7116.

Abstract 495: Targeting undruggable transcription factor HOXB13 in metastatic prostate cancer by CRISPR/Cas13d-based nanoparticle therapy

Abstract Background: The oncogenic transcription factor HOXB13, which is highly expressed in castration-resistant prostate cancer (CRPC), has been shown to promote CRPC growth and metastasis. However, HOXB13 is considered untargetable by traditional small-molecule-based drug design. Gene therapy is critical alternative strategy with potential to directly target such traditionally undruggable genes. Methods: Selective cell in organ targeting (SCORT) nanoparticles for precise delivery of nanoparticles to metastatic cancer cells in an organ were constructed and characterized by the NanoAssemblr Spark, Zetasizer Nano ZS, and transmission electron microscopy (TEM); Cas13d mRNA with pseudouridine modification was synthesized using in vitro transcription; Hemi-spleen injection of CRPC cells was performed to build CRPC liver metastatic models; RNA-seq was employed to evaluate off-target effects of Cas13d targeting. Results: By incorporating a prostate cancer-specific E3 aptamer to functionalized lipid-like nanoparticles (FTT5 LNP-E3), we have successfully constructed SCORT nanoparticles that enable preferential delivery of mRNAs to metastatic CRPC cells in the liver, as opposed to various normal cells in the surrounding liver tissue. We demonstrated that Cas13d-pre-gHOXB13 mediates highly effective and specific HOXB13 mRNA knockdown in CRPC cells. Significantly, systemic treatment of SCORT nanoparticles carrying Cas13d-pre-gHOXB13 mRNA decreased HOXB13 expression in the metastatic tumors, inhibited metastasis, and prolonged survival of mice bearing androgen receptor (AR)-positive (AR+) or negative (AR-) tumors. Notably, long term (6 and a half weeks) administration of SCORT nanoparticle-Cas13d-pre-gHOXB13 did not significant alter body weight, hepatic and renal function, chemokines and cytokines, and other factors, effectively highlighting its safety. Conclusions: This study is the first to demonstrate that undruggable oncogenic transcription factors can be targeted using nanoparticle-delivered gene therapy based on CRISPR/Cas13 RNA-targeting. The SCORT-CRISPR/Cas13d system is a highly flexible technology that would impact the larger field of translational science by allowing the design of Cas13d-based, cancer cell-specific delivered nanoparticles targeting other previously-undruggable oncogenic transcription factors in metastatic prostate cancer and other solid tumors. Citation Format: Zhifen Cui, Furong Huang, Kun Fang, Jingyue Yan, Yufan Zhou, Jeffrey Everitt, Yue Zhao, William Hankey, Zhong Chen, Hongyan Wang, Victor X. Jin, Yizhou Dong, Qianben Wang. Targeting undruggable transcription factor HOXB13 in metastatic prostate cancer by CRISPR/Cas13d-based nanoparticle therapy [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 495.

YAP1 Regulates the YAP1/AR/PSA Axis through Autophagy in Castration-Resistant Prostate Cancer and Mediates T-Cell Immune and Inflammatory Cytokine Infiltration.

The emergence of castration-resistant prostate cancer (CRPC) following androgen deprivation therapy (ADT) is associated with increased malignancy and limited treatment options. This study aims to investigate potential connections between immune cell infiltration and inflammatory cytokines with the YAP1/AR/PSA axis by exploring their interactions with autophagy. Our research reveals heightened levels of Yes-associated protein 1 (YAP1) expression in CRPC tissues compared with tissues from androgen-dependent prostate cancer (ADPC) and benign prostate hyperplasia (BPH). Additionally, a correlation was observed between YAP1 and PSA expressions in CRPC tissues, suggesting that YAP1 may exert a regulatory influence on PSA expression within CRPC. Enhanced YAP1 expression in C4-2 cells resulted in the upregulation of androgen receptor (AR) nuclear translocation and intracellular prostate-specific antigen (PSA) levels. Conversely, the suppression of YAP1 led to a decrease in PSA expression, suggesting that YAP1 may positively regulate the PSA in castration-resistant prostate cancer (CRPC) by facilitating AR nuclear import. The modulation of the autophagy activity exerts a significant impact on the expression levels of YAP1, the AR, and the PSA. Moreover, recent advancements in immunity and inflammation studies present promising avenues for potential therapies targeting prostate cancer (PC).

Chromatin activation with H3K36me2 and compartment shift in metastatic castration-resistant prostate cancer

Epigenetic modifiers are upregulated during the process of prostate cancer, acquiring resistance to castration therapy and becoming lethal metastatic castration-resistant prostate cancer (CRPC). However, the relationship between regulation of histone modifications and chromatin structure in CRPC has yet not fully been validated. Here, we reanalyzed publicly available clinical transcriptome and clinical outcome data and identified NSD2, a histone methyltransferase that catalyzes H3K36me2, as an epigenetic modifier that was upregulated in CRPC and whose increased expression in prostate cancer correlated with higher recurrence rate. We performed ChIP-seq, RNA-seq, and Hi-C to conduct comprehensive epigenomic and transcriptomic analyses to identify epigenetic reprogramming in CRPC. In regions where H3K36me2 was increased, H3K27me3 was decreased, and the compartment was shifted from inactive to active. In these regions, 68 aberrantly activated genes were identified as candidate downstream genes of NSD2 in CRPC. Among these genes, we identified KIF18A as critical for CRPC growth. Under NSD2 upregulation in CRPC, epigenetic alteration with H3K36me2-gain and H3K27me3-loss occurs accompanying with an inactive-to-active compartment shift, suggesting that histone modification and chromatin structure cooperatively change prostate carcinogenesis.

Tanshinone analog inhibits castration-resistant prostate cancer cell growth by inhibiting glycolysis in an AR-dependent manner

Androgen receptor (AR) is one of the key targets for the treatment of Castration-resistant prostate cancer (CRPC). Current endocrine therapy can greatly improve CRPC patients. However, with the change of pathogenic mechanism, acquired resistance often leads to the failure of treatment. Studies have shown that tanshinone IIA (TS-IIA) and its derivatives have significant antitumor activity, and have certain AR targeting effects, but the mechanism is unknown. In this study, the TS-IIA analog TB3 was found to significantly inhibit the growth of CRPC in vitro and in vivo. Molecular docking, cellular thermal shift assay and CHX experiments confirmed that AR was the target of TB3 and promoted the degradation of AR. Furthermore, TB3 can significantly inhibit glycolysis metabolism by targeting the AR/PKM2 axis..The addition of pyruvic acid could significantly alleviate the inhibitory effect of TB3 on CRPC cells. Besides, the knockdown of AR or PKM2 also could reverse the effect of TB3 on CRPC cells. Taken together, our study suggests that TS-IIA derivative TB3 inhibits glycolysis to prevent the CRPC process by targeting the AR/PKM2 axis.

Stat5 induces androgen receptor (AR) gene transcription in prostate cancer and offers a druggable pathway to target AR signaling.

Androgen receptor (AR) drives prostate cancer (PC) growth and progression, and targeting AR signaling is the mainstay of pharmacological therapies for PC. Resistance develops relatively fast as a result of refueled AR activity. A major gap in the field is the lack of understanding of targetable mechanisms that induce persistent AR expression in castrate-resistant PC (CRPC). This study uncovers an unexpected function of active Stat5 signaling, a known promoter of PC growth and clinical progression, as a potent inducer of AR gene transcription. Stat5 suppression inhibited AR gene transcription in preclinical PC models and reduced the levels of wild-type, mutated, and truncated AR proteins. Pharmacological Stat5 inhibition by a specific small-molecule Stat5 inhibitor down-regulated Stat5-inducible genes as well as AR and AR-regulated genes and suppressed PC growth. This work introduces the concept of Stat5 as an inducer of AR gene transcription in PC. Pharmacological Stat5 inhibitors may represent a new strategy for suppressing AR and CRPC growth.

Exploring androgen receptor signaling pathway in prostate cancer: A path to new discoveries.

Androgen deprivation therapy has achieved significant success in treating prostate cancer through strategies centered on the androgen receptor. However, the emergence of castration-resistant prostate cancer highlights this therapy limitation, underscoring the need to elucidate the mechanisms of treatment resistance. This review aimed to focus on multifaceted resistance mechanisms, including androgen receptor overexpression, splice variants, missense mutations, the involvement of the glucocorticoid receptor, and alterations in coregulators and transcription factors, revealing their roles in castration-resistant prostate cancer progression. These mechanisms promote cell survival and proliferation, depending on the androgen receptor signaling pathway, leading to resistance to conventional therapies. Amplification and mutations in the androgen receptor gene facilitate selective adaptation in treatment-resistant cells, consequently diminishing therapeutic efficacy. Furthermore, the activation of glucocorticoid receptors and aberrant regulation of specific coregulators and transcription factors contribute to the activation of androgen receptor-independent signaling pathways, promoting cell survival and proliferation. These findings hold promise for identifying new targets for treating castration-resistant prostate cancer and developing personalized treatment strategies. The development of future therapies will hinge on precisely targeting the androgen receptor signaling pathway, necessitating a deeper understanding of the molecular targets unique to castration-resistant prostate cancer.

Abstract B009: Targeting the AR co-activator CBP/p300 axis in castration resistant prostate cancer Impacts DNA damage repair function

Abstract Prostate cancer (PCa) is the second leading cause of cancer related deaths in US men. PCa is an androgen dependent disease and the transcription factor, androgen receptor (AR), is crucial for the overall outcome. Currently, androgen-deprivation therapy (ADT) is the standard of care first-line therapy for metastatic PCa. Resistance to ADT almost uniformly leads to lethal disease, termed castration-resistant prostate cancer (CRPC). Thus, there is a significant unmet clinical need to identify and develop novel strategies to treat CRPC. CBP/p300 are potent co-activators for AR. High expression of CBP/p300 are associated with locally advanced disease and castration resistant function of AR; thus, resulting in poor patient outcome and further highlighting the need to discern the role of CBP/p300 to potentially develop novel therapeutic targets for precision medicine. Previous studies have relied on non-specific compounds and genetic silencing to target CBP/p300 and its associated transcriptional machinery. In this study, CBP/p300 mediated bromodomain activity is targeted by CCS1477 (Inobrodib), a first-in-class bromodomain inhibitor developed by Cell Centric. CCS1477 was shown to demonstrate effective inhibition in growth and clonogenicity assays. Inhibition of the CBP/p300 bromodomain with CCS1477 resulted in significant downregulation of AR-FL, AR-V7, and its targets’ mRNA expression in addition to inhibition of associated factors such as c-MYC and its downstream targets in PCa cell lines as well as patient derived xenograft (PDX) models. This study shows that CBP and p300 are highly expressed and correlate closely with AR gene expression and AR activity score in primary PCa and CRPC patient samples. The clinical significance of CBP/p300 expression in PCa has been investigated via elucidation of CBP/p300 transcriptional reprogramming and role in DNA damage response (DDR) pathways. Specifically, findings revealed that CBP/p300 bromodomain suppression sensitizes to AR-dependent DNA-repair. Transcriptional mapping identified CBP/p300 as regulators of cell proliferation and DNA repair processes, which were functionally confirmed across several PCa model systems. To assess relevance, exogenous challenge with radiation revealed that CBP/p300 bromodomain is required for AR-mediated DNA repair, and CBP/p300 expression is linked to DNA repair capacity in the clinical setting. Molecular analyses revealed that CBP/p300 facilitate double-strand break (DSB) repair efficiency via homologous recombination (HR) mediated DDR. Congruently, CBP/p300 strongly correlated with HR gene expression in PCa patient tissue. These collective findings reveal that CBP/p300 govern repair of DNA DSBs by regulating HR, thus modulating genome integrity, and promoting CRPC growth. In conclusion, these studies identify CBP/p300 as a driver of PCa tumorigenesis through coordinated control of critical transcriptional events and lay the groundwork to optimize therapeutic strategies for advanced PCa via CBP/p300 inhibition, potentially in combination with AR-directed therapies. Citation Format: Sumaira Sardar. Targeting the AR co-activator CBP/p300 axis in castration resistant prostate cancer Impacts DNA damage repair function [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr B009.

Targeting glutamine dependence with DRP-104 inhibits proliferation and tumor growth of castration-resistant prostate cancer.

Prostate cancer (PCa) continues to be one of the leading causes of cancer deaths in men. While androgen deprivation therapy is initially effective, castration-resistant PCa (CRPC) often recurs and has limited treatment options. Our previous study identified glutamine metabolism to be critical for CRPC growth. The glutamine antagonist 6-diazo-5-oxo-l-norleucine (DON) blocks both carbon and nitrogen pathways but has dose-limiting toxicity. The prodrug DRP-104 is expected to be preferentially converted to DON in tumor cells to inhibit glutamine utilization with minimal toxicity. However, CRPC cells' susceptibility to DRP-104 remains unclear. Human PCa cell lines (LNCaP, LAPC4, C4-2/MDVR, PC-3, 22RV1, NCI-H660) were treated with DRP-104, and effects on proliferation and cell death were assessed. Unbiased metabolic profiling and isotope tracing evaluated the effects of DRP-104 on glutamine pathways. Efficacy of DRP-104 in vivo was evaluated in a mouse xenograft model of neuroendocrine PCa, NCI-H660. DRP-104 inhibited proliferation and induced apoptosis in CRPC cell lines. Metabolite profiling showed decreases in the tricarboxylic acidcycle and nucleotide synthesis metabolites. Glutamine isotope tracing confirmed the blockade of both carbon pathway and nitrogen pathways. DRP-104 treated CRPC cells were rescued by the addition of nucleosides. DRP-104 inhibited neuroendocrine PCa xenograft growth without detectable toxicity. The prodrug DRP-104 blocks glutamine carbon and nitrogen utilization, thereby inhibiting CRPC growth and inducing apoptosis. Targeting glutamine metabolism pathways with DRP-104 represents a promising therapeutic strategy for CRPC.

Inhibition of orthotopic castration-resistant prostate cancer growth and metastasis in mice by JC VLPs carrying a suicide gene driven by the PSA promoter.

Metastatic castration-resistant prostate cancer (mCRPC) is challenging to treat. Virus-like particles (VLPs), originating from JC polyomavirus (JCPyV) and carrying a suicide gene driven by the PSA promoter (PSAtk-VLPs), can inhibit tumor growth in animal models of human prostate cancer. However, the efficacy of suppression of orthotopic PCa growth and metastasis by PSAtk-VLPs remains undetermined. Here, we established an iRFP stable expression CRPC cell line suitable for deep-tissue observation using fluorescence molecular tomography (FMT). These cells were implanted into murine prostate tissue, and PSAtk-VLPs were systemically administered via the tail vein along with the prodrug ganciclovir (GCV), allowing for the real-time observation of orthotopic prostate tumor growth and CRPC tumor metastasis. Our findings demonstrated that systemic PSAtk-VLPs administration with GCV and subsequent FMT scanning facilitated real-time observation of the suppressed growth in mouse iRFP CRPC orthotopic tumors, which further revealed a notable metastasis rate reduction. Systemic PSAtk-VLPs and GCV administration effectively inhibited orthotopic prostate cancer growth and metastasis. These findings suggest the potential of JCPyV VLPs as a promising vector for mCRPC gene therapy. Conclusively, systemically administered JCPyV VLPs carrying a tissue-specific promoter, JCPyV VLPs can protect genes within the bloodstream to be specifically expressed in specific organs.

Newly developed preclinical models reveal broad-spectrum CDK inhibitors as potent drugs for CRPC exhibiting primary resistance to enzalutamide.

Androgen-deprivation therapy is a standard treatment for advanced prostate cancer. However, most patients eventually acquire resistance and progress to castration-resistant prostate cancer (CRPC). In this study, we established new CRPC cell lines, AILNCaP14 and AILNCaP15, from LNCaP cells under androgen-deprived conditions. Unlike most pre-existing CRPC cell lines, both cell lines expressed higher levels of androgen receptor (AR) and prostate-specific antigen (PSA) than parental LNCaP cells. Moreover, these cells exhibited primary resistance to enzalutamide. Since AR signaling plays a significant role in the development of CRPC, PSA promoter sequences fused with GFP were introduced into AILNCaP14 cells to conduct GFP fluorescence-based chemical screening. We identified flavopiridol, a broad-spectrum CDK inhibitor, as a candidate drug that could repress AR transactivation of CRPC cells, presumably through the inhibition of phosphorylation of AR on the serine 81 residue (pARSer81 ). Importantly, this broad-spectrum CDK inhibitor inhibited the proliferation of AILNCaP14 cells both in vitro and in vivo. Moreover, a newly developed liver metastatic model using AILNCaP15 cells revealed that the compound attenuated tumor growth of CRPC harboring highly metastatic properties. Finally, we developed a patient-derived xenograft (PDX) model of CRPC and DCaP CR from a patient presenting therapeutic resistance to enzalutamide, abiraterone, and docetaxel. Flavopiridol successfully suppressed the tumor growth of CRPC in this PDX model. Since ARSer81 was found to be phosphorylated in clinical CRPC samples, our data suggested that broad-spectrum CDK inhibitors might be a potent candidate drug for the treatment of CRPC, including those exhibiting primary resistance to enzalutamide.

Polyphyllin I induces ferroptosis in castration-resistant prostate cancer cells through the ERK/DNMT1/ACSL4 axis.

Castration-resistant prostate cancer (CRPC) inevitably arises after androgen deprivation therapy (ADT). Therefore, there is an urgent need to search for novel treatment strategies for CRPC. Polyphyllin I (PPI), one of the steroidal saponins in paris polyphylla, has been shown to have an anticancer effect. This study investigated the role and mechanism of PPI in CRPC cell ferroptosis. Protein levels of GPX4, p-extracellular regulated protein kinases (ERK), ERK, DNMT1, and ACSL4 were measured by Western blot. DNMT1 and ACSL4 mRNA expression was analyzed byreverse transcription-quantitative polymerase chain reaction (RT-qPCR). Prostate cancer cells (DU145, PC3) were treated with PPI. Cell viability was assessed utilizing Cell Counting Kit-8 (CCK-8) assay. The role of PPI in regulating ferroptosis was determined by analyzing lipid reactive oxygen species (ROS), malonyl dialdehyde (MDA), iron (Fe2+ ), and glutathione (GSH) content. Chromatin immunoprecipitation (ChIP) assay verified the effect of DNMT1 on the ACSL4 promoter. The methylation level of ACSL4 promoter was assessed utilizing MSP. A nude mice xenograft was adopted to detect the effect of PPI in vivo. PPI inhibited CRPC cell proliferation, reduced levels of GSH and GPX4, and increased levels of MDA, Fe2+ , and ROS, while ERK inhibitor reversed the effect of PPI on ferroptosis. PPI repressed the methylation level of ACSL4 promoter by inhibiting DNMT1. DNMT1 knockdown promoted CRPC cell ferroptosis by regulating ACSL4. PPI induced ferroptosis and suppressed CRPC growth in nude mice. PPI can be used as a ferroptosis inducer to induce ferroptosis in CRPC cells via the ERK/DNMT1/ACSL4 axis, suggesting that PPI may be a new strategy for CRPC treatment.