Login to Access Video or Poster Abstract: MP83-10
Sources of Funding: This work was supported by JSPS KAKENHI Grant Number 15K20109 and 26861299.

Introduction

Recently, cabazitaxel (CBZ) was identified as the most effective cytotoxic agent to demonstrate an improvement in survival in men with docetaxel-refractory CRPC. Accumulating evidence suggests EMT induced by androgen-deprivation therapy or docetaxel explains in part the aggressiveness of CRPC. The aim of this study was to screen for EMT inhibitory drugs by bioinformatic analysis and exploring the antitumor effect of CBZ for resistant CRPC.

Methods

PC3 and DU145 were used in this study. We incubated the cell lines with gradually increasing concentrations of CBZ to establish CBZ-resistant cell lines. In addition, we performed screening tests for candidate drugs to inhibit EMT of metastatic CRPC. The combined effect of CBZ and a candidate EMT inhibitor for CBZ-resistant cell lines was determined using a cell viability assay._x000D_

Results

We established a CBZ-resistant cell line, PC3-CBZR (Figure). PC3-CBZR cells underwent cell division with 3 nM CBZ. We compared the cytotoxic effect of CBZ on PC3 and PC3-CBZR cells using a cell viability assay. The IC50 of CBZ in PC3 and PC3-CBZR cells were 16.0 nM and 2.5 nM, respectively. Real-time PCR showed up-regulation of ZEB family and Vimentin and down-regulation of E-cadherin expression, suggesting PC3-CBZR cells had elevated metastatic potential. We screened for EMT inhibitors using The Broad institute&[prime]s connectivity map (CMAP) analysis, with human whole genome array data and identified a candidate drug, KOH513. The relative cell viabilities treated with 10 μM KOH513 in PC3 and PC3-CBZR cells were 60.0 ± 1.7%, 58.7 ± 0.7%, respectively. KOH513 was suggested to have an antitumor effect on CBZ-resistant prostate cancer. Real time PCR and immune fluorescence assay showed that KOH513 down-regulated ZEB family and Vimentin in PC3-CBZR cells at the protein and mRNA levels. We tested the efficacy of KOH513 in CBZ resistance using a cell viability assay. Single-agent administration of 6 nM CBZ or 3 μM KOH513 did not have an antitumor effect in PC3-CBZ cells. However, when treated with 6 nM CBZ and 3 µM KOH513 in combination, PC3-CBZ cell proliferation was significantly suppressed (relative cell viability 77.0 ± 1.7%). In the same manner, we established CBZ-resistant DU145 cells, DU145-CBZR. DU145-CBZR cells had also had an up-regulated EMT pathway, and cell proliferation was suppressed by EMT inhibitor KOH513. These results suggested that KOH513 had potential for reprogramming CBZ resistant CRPC cell lines into sensitive cell lines._x000D_

Conclusions

Targeting EMT signaling pathways with KOH513 can overcome CBZ resistance in CRPC.

Funding

This work was supported by JSPS KAKENHI Grant Number 15K20109 and 26861299.