Enzalutamide (Enz) is commonly utilized as the initial treatment strategy for advanced prostate cancer (PCa). However, a notable subset of patients may experience resistance to Enz, resulting in reduced effectiveness. Utilizing Gene Expression Omnibus (GEO) databases, we identified CBX2 as a crucial factor in mediating resistance to Enz, primarily due to its inhibitory effect on the P53 signaling pathway. Silencing of CBX2 using small interfering RNA (siRNA) led to elevated levels of P53 expression in LNCaP cells. This indicates that CBX2 may have a critical effect on PCa Enz resistance and could serve as a promising therapeutic target for individuals with Enz resistance.

BACKGROUND: Circular RNAs (circRNAs) have been shown to be involved in tumorigenesis and progression. However, the role of circGLIS3 (hsa_circ_0002874) in prostate cancer (PCa) has yet not been reported.
METHODS: Candidate circRNA were determined through comprehensive analysis of public datasets, PCa cell lines, and tissues data. A series of cellular functional assays, including CCK-8, colony formation, wound healing, and transwell assays were performed. Subsequently, RNA sequencing, RNA immunoprecipitation, methylated RNA immunoprecipitation, microRNA pulldown, luciferase reporter assay, and western blot were used to explore the underlying molecular mechanisms. Moreover, the xenograft tumor mouse model was established to elucidate the function of circGLIS3.
RESULTS: CircGLIS3, derived from exon 2 of the parental GLIS3 gene, was identified as a novel oncogenic circRNA in PCa that was closely associated with the biochemical recurrence. Its expression levels were upregulated in PCa tissues and cell lines as well as enzalutamide high-resistant cells. The cellular functional assays revealed that circGLIS3 promoted PCa cell proliferation, migration, and invasion. METTL3-mediated N6-methyladenosine (m6A) modification maintained its upregulation by enhancing its stability. Mechanically, CircGLIS3 sponged miR-661 to upregulate MDM2, thus regulating the p53 signaling pathway to promote cell proliferation, migration, and invasion. Furthermore, in vitro and in vivo experiments, the knockdown of circGLIS3 improved the response of PCa cells to ARSI therapies such as enzalutamide.
CONCLUSIONS: METTL3-mediated m6A modification of circGLIS3 regulates the p53 signaling pathway via the miR-661/MDM2 axis, thereby facilitating PCa progression. Meanwhile, this study unveils a promising potential target for ARSI therapy for PCa.

Sharply increased reactive oxygen species (ROS) are thought to induce oxidative stress, damage cell structure and cause cell death; however, its role in prostate cancer remains unclear. Enzalutamide is a widely used anti-prostate cancer drug that antagonizes androgen binding with its receptor. Further exploration of the mechanism and potential application strategies of enzalutamide is crucial for the treatment of prostate cancer. Here, we confirmed PEX10 can be induced by ROS activators while reduce ROS level in prostate cancer cells, which weakened the anti-tumor effect of ROS activators. The androgen receptor (AR) can promote the expression of PEX10 by acting as an enhancer in cooperation with FOXA1. The anti-tumor drug enzalutamide inhibits PEX10 by inhibiting the function of AR, and synergize with ROS activators ML210 or RSL3 to produce a stronger anti-tumor effect, thereby sensitizing cells to ROS activators. This study reveals a previously unrecognized function of enzalutamide and AR by regulating PEX10 and suggests a new strategy of enzalutamide application in prostate cancer treatment.

OBJECTIVE: Androgen receptor inhibitors (ARIs) have become an effective treatment for advanced prostate cancer (PC). However, it is unknown which ARI is the most helpful and safe for men with advanced PC. Our aim is to help physicians make clinical decisions and provide medication guidelines for patients with advanced PC to avoid potential risks when using ARIs for treatment.
METHODS: We systematically searched the following databases: PubMed, Embase and Cochrane Library, with a literature publication deadline of February 2023. The primary efficacy outcomes were 18-month overall survival (OS), treatment-emergent adverse events (TEAEs), hypertension and fatigue. The network meta-analysis (NMA) was performed by Stata 15.1, and Revman 5.3 was used to assess the included studies\' risk of bias.
RESULTS: The analysis included 26 trials with 26 263 people. The surface under the cumulative ranking curve (SUCRA) concluded that enzalutamide (86.8%) showed the best effect in prolonging the OS of patients. Flutamide led to the highest risk of TEAEs (29.9%) and AEs leading to discontinuation (12.8%). Apalutamide (13.4%) led to the highest risk of grade ≥3 TEAEs. Enzalutamide had the highest risk of hypertension (0.2%), grade ≥3 hypertension (4.5%) and fatigue (5.2%).
CONCLUSIONS: This NMA indicates there is no one ARI to reach both the most effective and safe therapy aims for treating advanced PC and that there is a compromise between the efficacy and safety of ARIs in the treatment of advanced PC. Physicians should weigh the risks to safety against the anticipated benefits when prescribing these drugs to patients with PC.

BACKGROUND: Endocrine resistance driven by sustained activation of androgen receptor (AR) signaling pathway in advanced prostate cancer (PCa) is fatal. Characterization of mechanisms underlying aberrant AR pathway activation to search for potential therapeutic strategy is particularly important. Rac GTPase-activating protein 1 (RACGAP1) is one of the specific GTPase-activating proteins. As a novel tumor proto-oncogene, overexpression of RACGAP1 was related to the occurrence of various tumors.
METHODS: Bioinformatics methods were used to analyze the relationship of expression level between RACGAP1 and AR as well as AR pathway activation. qRT-PCR and western blotting assays were performed to assess the expression of AR/AR-V7 and RACGAP1 in PCa cells. Immunoprecipitation and immunofluorescence experiments were conducted to detect the interaction and co-localization between RACGAP1 and AR/AR-V7. Gain- and loss-of-function analyses were conducted to investigate the biological roles of RACGAP1 in PCa cells, using MTS and colony formation assays. In vivo experiments were conducted to evaluate the effect of RACGAP1 inhibition on the tumor growth.
RESULTS: RACGAP1 was a gene activated by AR, which was markedly upregulated in PCa patients with CRPC and enzalutamide resistance. AR transcriptionally activated RACGAP1 expression by binding to its promoter region. Reciprocally, nuclear RACGAP1 bound to the N-terminal domain (NTD) of both AR and AR-V7, blocking their interaction with the E3 ubiquitin ligase MDM2. Consequently, this prevented the degradation of AR/AR-V7 in a ubiquitin-proteasome-dependent pathway. Notably, the positive feedback loop between RACGAP1 and AR/AR-V7 contributed to endocrine therapy resistance of CRPC. Combination of enzalutamide and in vivo cholesterol-conjugated RIG-I siRNA drugs targeting RACGAP1 induced potent inhibition of xenograft tumor growth of PCa.
CONCLUSIONS: In summary, our results reveal that reciprocal regulation between RACGAP1 and AR/AR-V7 contributes to the endocrine resistance in PCa. These findings highlight the therapeutic potential of combined RACGAP1 inhibition and enzalutamide in treatment of advanced PCa.

Prostate carcinoma represents a predominant malignancy affecting the male population, with androgen deprivation therapy (ADT) serving as a critical therapeutic modality for advanced disease states, but it often leads to the development of resistance. Enzalutamide (Enz), a second-generation antiandrogen drug, initially offers substantial therapeutic benefit, but its efficacy wanes as drug resistance ensues. In this study, we found that synaptotagmin 4 (SYT4) is an upregulated gene in enzalutamide-resistant (EnzR) cell lines. The downregulation of SYT4, in combination with enzalutamide therapy, substantially enhances the antiproliferative effect on resistant prostate cancer cells beyond the capacity of enzalutamide monotherapy. SYT4 promotes vesicle efflux by binding to the synaptosome-associated protein 25 (SNAP25), thereby contributing to cell resistance against enzalutamide. The elevated expression of SYT4 is mediated by bromodomain-containing protein 4 (BRD4), and BRD4 inhibition effectively suppressed the expression of SYT4. Treatment with a therapeutic dose of enzalutamide combined with ASO-1, an antisense oligonucleotide drug targeting SYT4, shows promising results in reversing the resistance of prostate cancer to enzalutamide.

The androgen receptor signaling inhibitor (ARSI) enzalutamide (Enz) has shown critical efficacy in the treatment of advanced prostate cancer (PCa). However, the development of drug resistance is a significant factor contributing to mortality in PCa patients. We aimed to explore the key mechanisms of Enz-resistance. Through analysis of GEO databases, we identified SLC4A4 as a novel driver in Enz resistance. Long-term Enz treatment leads to the up-regulation of SLC4A4, which in turn mediates P53 lactylation via the NF-κB/STAT3/SLC4A4 axis, ultimately leading to the development of Enz resistance and progression of PCa. SLC4A4 knockdown overcomes Enz resistance both in vitro and in vivo. Hence, our results suggest that targeting SLC4A4 could be a promising therapeutic strategy for Enz resistance. STATEMENT OF SIGNIFICANCE: SLC4A4 is a novel driver of enzalutamide resistance.

Despite the initial efficacy of enzalutamide in castration-resistant prostate cancer (CRPC), inevitable resistance remains a significant challenge. Here, the synergistic induction of copper-dependent cell death (cuproptosis) in CRPC cells is reported by enzalutamide and copper ionophores (elesclomol/disulfiram). Mechanistically, enzalutamide treatment increases mitochondrial dependence in CRPC cells, rendering them susceptible to cuproptosis, as evidenced by specific reversal with the copper chelator tetrathiomolybdate. This susceptibility is characterized by hallmarks of cuproptosis, including lipoylated protein aggregation and iron-sulfur cluster protein instability. Interestingly, the mitochondrial matrix reductase, FDX1, specifically correlates with elesclomol sensitivity, suggesting a potential mechanistic divergence between the two copper ionophores. Notably, this synergistic effect extends beyond in vitro models, demonstrating efficacy in 22Rv1 xenografts, mouse Pten p53 knockout organoids. Importantly, enzalutamide significantly enhances copper ionophore-mediated cytotoxicity in enzalutamide-resistant cells. Collectively, these findings indicate that enzalutamide and copper ionophores synergistically induce cuproptosis, offering a promising therapeutic avenue for CRPC, potentially including enzalutamide-resistant cases.

Second-generation AR antagonists, such as enzalutamide, are the primary therapeutic agents for advanced prostate cancer. However, the development of both primary and secondary drug resistance leads to treatment failures and patient mortality. Bifunctional agents that simultaneously antagonize and degrade AR block the AR signaling pathway more completely and exhibit excellent antiproliferative activity against wild-type and drug-resistant prostate cancer cells. Here, we reported the discovery and optimization of a series of biphenyl derivatives as androgen receptor antagonists and degraders. These biphenyl derivatives exhibited potent antiproliferative activity against LNCaP and 22Rv1 cells. Our discoveries enrich the diversity of small molecule AR degraders and offer insights for the development of novel AR degraders for the treatment of enzalutamide-resistant prostate cancer.

Background: Castration-resistant prostate cancer (CRPC), which has developed resistance to next-generation antiandrogens, such as enzalutamide (Enz), is a lethal disease. Furthermore, transcriptional regulation by super enhancers (SEs) is crucial for the growth and spread of prostate cancer, as well as drug resistance. The functions of SEs, a significant class of noncoding DNA cis-regulatory elements, have been the subject of numerous recent studies in the field of cancer research. Materials and Methods: The goal of this research was to identify SEs associated with Enz resistance in C4-2B cells using chromatin immunoprecipitation sequencing and cleavage under targets and tagmentation (CUT&Tag). Using HOMER analysis to predict protein/gene-binding motifs, we identified master transcription factors (TFs) that may bind to SE sites. Using small interfering RNA, WST-1 assays, and qRT-PCR, we then confirmed the associations between TFs of SEs and Enz resistance. Results: A total of 999 SEs were screened from C4-2B EnzR cells in total. Incorporating analysis with RNA-seq data revealed 41 SEs to be strongly associated with the promotion of Enz resistance. In addition, we finally predicted that master TFs bind to SE-binding regions. Subsequently, we selected zinc finger protein 467 (ZFP467) and SMAD family member 3 to confirm the functional connections of master TFs with Enz resistance through SEs (ZNF467). Conclusions: In this study, SMAD3 and ZNF467 were found to be closely related to Enz-resistant CRPC. Our research uncovered a sizable group of SEs linked to Enz resistance in prostate cancer, dissected the mechanisms underlying SE Enz resistance, and shed light on potential clinical uses for SEs.