Objective To design a novel bromodomain-containing protein 4 (BRD4) and histone deacetylase (HDAC) dual-target inhibitor (11b), and to elucidate its therapeutic efficacy and mechanisms in suppressing prostate cancer through epigenetic regulation.

Methods BRD4 and HDAC expression levels were assessed via immunohistochemistry (IHC) using prostate cancer tissue microarrays. The inhibitory activity of 11b was screened across three prostate cancer cell lines, with the half-maximal inhibitory concentration (IC₅₀) determined by CCK-8 assay. Western blot was employed to analyze changes in the expression of target proteins, including BRD4, c-Myc proto-oncogene protein (c-Myc), and Ac-H3K27, with parallel comparisons to single-target agents, including suberoylanilide hydroxamic acid (SAHA), a HDAC inhibitor, and JQ-1, a BRD4 inhibitor. Cell invasion and proliferation were evaluated using Transwell and colony formation assays, and the autophagy mechanism was validated using 3-methyladenine (3-MA), an autophagy inhibitor. A PC-3 xenograft model was established in nude mice. Then, 11b (7.5 mg/kg or 15 mg/kg), normal saline, SAHA, and JQ-1 were administered via intraperitoneal injection, and their tumor growth inhibition effects were observed. The percentage of target protein-positive cells and the expression levels of target genes were quantified via IHC and RT-PCR, respectively.

Results BRD4 and HDAC expression levels were both higher in tumor tissues than those in normal tissues (P < 0.01). 11b exhibited the strongest inhibitory activity against PC-3 cells (IC₅₀ = 8.28 μmol/L), outperforming SAHA (22.61 μmol/L) and JQ-1 (22.09 μmol/L). Treatment with 11b reduced BRD4 and c-Myc expression by (41.58 ± 3.28)% and (63.21 ± 6.91)%, respectively (P < 0.01), and increased the Ac-H3K27 level to 6.52-fold that of the negative control (NC) group (P < 0.01), demonstrating greater modulation than either SAHA or JQ-1 did. The in vitro experiment showed that 8 μmol/L 11b treatment reduced PC-3 colony formation and migration by 97.5% and 96.3%, respectively (P < 0.001), and co-treatment with 3-MA reversed its cytotoxic effects. The in vivo experiment showed that 11b at both 7.5 mg/kg and 15 mg/kg significantly reduced tumor volume and weight compared with the control, SAHA, and JQ-1 groups (all P < 0.01), with the proportion of percentage of target protein-positive cells and the expression of target genes showing trends consistent with in vitro findings.

Conclusion The dual-target inhibitor 11b exerts potent antitumor effects in prostate cancer by synergistically modulating the BRD4/HDAC pathways. 11b demonstrates therapeutic efficacy superior to that of the single-target agents SAHA and JQ-1 in suppressing prostate cancer progression, highlighting its potential for clinical translation.