Abstract
Background:The androgen receptor (AR) is a major drug target in prostate cancer (PCa). We profiled the AR-regulated kinome to identify clinically relevant and druggable effectors of AR signaling.Methods:Using genome-wide approaches, we interrogated all AR regulated kinases. Among these, choline kinase alpha (CHKA) expression was evaluated in benign (n = 195), prostatic intraepithelial neoplasia (PIN) (n = 153) and prostate cancer (PCa) lesions (n = 359). We interrogated how CHKA regulates AR signaling using biochemical assays and investigated androgen regulation of CHKA expression in men with PCa, both untreated (n = 20) and treated with an androgen biosynthesis inhibitor degarelix (n = 27). We studied the effect of CHKA inhibition on the PCa transcriptome using RNA sequencing and tested the effect of CHKA inhibition on cell growth, clonogenic survival and invasion. Tumor xenografts (n = 6 per group) were generated in mice using genetically engineered prostate cancer cells with inducible CHKA knockdown. Data were analyzed with χ2 tests, Cox regression analysis, and Kaplan-Meier methods. All statistical tests were two-sided.Results:CHKA expression was shown to be androgen regulated in cell lines, xenografts, and human tissue (log fold change from 6.75 to 6.59, P = .002) and was positively associated with tumor stage. CHKA binds directly to the ligand-binding domain (LBD) of AR, enhancing its stability. As such, CHKA is the first kinase identified as an AR chaperone. Inhibition of CHKA repressed the AR transcriptional program including pathways enriched for regulation of protein folding, decreased AR protein levels, and inhibited the growth of PCa cell lines, human PCa explants, and tumor xenografts.Conclusions:CHKA can act as an AR chaperone, providing, to our knowledge, the first evidence for kinases as molecular chaperones, making CHKA both a marker of tumor progression and a potential therapeutic target for PCa.
Highlights
The androgen receptor (AR) is a major drug target in prostate cancer (PCa)
As with many other cancer types, resistance to therapy occurs in PCa in the form of progression to advanced castrationresistant prostate cancer (CRPC) [6,7] and is accompanied by reactivation or maintenance of AR signaling, which triggers a unique AR transcriptome [8]
Indirect mechanisms driving elevation of AR protein expression in PCa include the upregulation of heat shock proteins (HSPs) that act as chaperones for AR
Summary
All cell lines were verified by genetic profiling of polymorphic short tandem repeat (STR) loci as per ATCC standards. Recursive partitioning (RP) using a conditional inference framework was used to find a significant cutoff for each input gene for predicting recurrence-free survival using a prostate cancer gene expression dataset with accompanying survival data from 79 patients [27]. This was implemented using the ctree [28] function in the R (R Core Team, 2014). Kaplan-Meier survival curves and recursive partitioning plots were displayed for those genes that are predictive of recurrence-free survival, with P values of less than .05 (corrected for the testing of multiple cutoffs, but not genes).
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