Introduction: Because CBZ-resistant CRPC has an extremely poor prognosis, establishment of a novel treatment for CBZ-resistant CRPC is greatly needed. The aim of this study was to explore drugs targeting the CBZ-resistance related genetic network by bioinformatic analysis and testing the antitumor effects of CBZ for resistant CRPC.
Methods: Two CRPC cell lines, DU145 and PC3, and two CBZ-resistant cell lines were used in this study. We incubated the cell lines with gradually increasing concentrations of CBZ to establish CBZ-resistant cell lines. We performed screening tests for candidate drugs to inhibit the genetic network in CBZ-resistant CRPC using in silico analysis, and identified a candidate drug, KHO931. The antitumor effect of KHO931 for CBZ-resistant cell lines was determined using mice xenograft tumor models.
Results: We incubated DU145 and PC3CR cells with gradually increasing concentrations of CBZ for 2 years and established the CBZ-resistant cell lines, DU145CR and PC3CR. These CBZ-resistant cell lines underwent cell division with 3 nM CBZ. We compared the cytotoxic effect of CBZ on DU145, PC3, DU145CR and PC3CR cells using a cell viability assay. The half maximal inhibitory concentration (IC50) of CBZ in CBZ-resistant cell lines was 3 times higher than that for parental cells. Cell cycle analysis demonstrated DU145CR and PC3CR had resistance to G2/M arrest induced by CBZ. Microarray analysis revealed gene clusters related to cell division were up-regulated in DU145CR and PC3CR. Next, we tested the anti-tumor effect of the candidate drug KHO931 for overcoming CBZ-resistance. KHO931 treatment had an anti-tumor effect in CBZ-resistant cell lines. Quantitative PCR revealed that KHO931 down-regulated mRNA expression of the Cytoskeleton (CSK)-related protein family, which regulates G2/M transition. A CSK-related protein inhibitor or genetic knockdown of CSK-related protein significantly suppressed cell proliferation of DU145CR and PC3CR. We tested the efficacy of KHO031 for CBZ resistance in mice xenograft models. KHO931 inhibited xenograft tumor growth in monotherapy. In addition, KHO931 and CBZ had a synergic effect on DU145CR and PC3CR xenograft tumors. These results suggest that KHO931 has potential for reprogramming CBZ-resistant CRPC.
Conclusions: Targeting the G2/M regulation mechanism with KHO931 can overcome CBZ resistance in CRPC. Source of
Funding: This study was supported by Grants-in-Aid for Scientific Research (#17K16813 to H. Hongo and #17K11158 to T. Kosaka) from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.