Androgen Suppresses the Proliferation of Androgen Receptor-Positive Castration-Resistant Prostate Cancer Cells via Inhibition of Cdk2, CyclinA, and Skp2

John M Kokontis,Chi-Jung Chung,Hui-Ping Lin,Chung-Ho Chang,Shutsung Liao,Junichi Fukuchi,Tzu-Min Chan,Richard A Hiipakka,Ching-Yu Lin,Shih Sheng Jiang,Chih-Pin Chuu,Allen Gao

doi:10.1371/journal.pone.0109170

Abstract

The majority of prostate cancer (PCa) patient receiving androgen ablation therapy eventually develop castration-resistant prostate cancer (CRPC). We previously reported that androgen treatment suppresses Skp2 and c-Myc through androgen receptor (AR) and induced G1 cell cycle arrest in androgen-independent LNCaP 104-R2 cells, a late stage CRPC cell line model. However, the mechanism of androgenic regulation of Skp2 in CRPC cells was not fully understood. In this study, we investigated the androgenic regulation of Skp2 in two AR-positive CRPC cell line models, the LNCaP 104-R1 and PC-3AR Cells. The former one is an early stage androgen-independent LNCaP cells, while the later one is PC-3 cells re-expressing either wild type AR or mutant LNCaP AR. Proliferation of LNCaP 104-R1 and PC-3AR cells is not dependent on but is suppressed by androgen. We observed in this study that androgen treatment reduced protein expression of Cdk2, Cdk7, Cyclin A, cyclin H, Skp2, c-Myc, and E2F-1; lessened phosphorylation of Thr14, Tyr15, and Thr160 on Cdk2; decreased activity of Cdk2; induced protein level of p27Kip1; and caused G1 cell cycle arrest in LNCaP 104-R1 cells and PC-3AR cells. Overexpression of Skp2 protein in LNCaP 104-R1 or PC-3AR cells partially blocked accumulation of p27Kip1 and increased Cdk2 activity under androgen treatment, which partially blocked the androgenic suppressive effects on proliferation and cell cycle. Analyzing on-line gene array data of 214 normal and PCa samples indicated that gene expression of Skp2, Cdk2, and cyclin A positively correlates to each other, while Cdk7 negatively correlates to these genes. These observations suggested that androgen suppresses the proliferation of CRPC cells partially through inhibition of Cyclin A, Cdk2, and Skp2.

Highlights

In 1941, Charles Huggins reported that androgen ablation therapy caused regression of primary and metastatic androgendependent prostate cancer (PCa) [1]
Activation of cyclin-dependent kinase 2 (Cdk2) complexes requires dephosphorylation of Thr14 and Tyr15 on Cdk2 by cdc25 phosphatase and phosphorylation of Thr160 on Cdk2 [26,27], which is mediated by CAK, a complex of Cdk7 and cyclin H [28]
Protein level of p27Kip1 was inverse-correlated to Sphase kinase-associated protein 2 (Skp2) in these castration-resistant prostate cancer (CRPC) cells, which was consistent with the fact that Skp2 targets p27Kip1 for ubiquitination and degradation [18,19,20]

Summary

Introduction

In 1941, Charles Huggins reported that androgen ablation therapy caused regression of primary and metastatic androgendependent prostate cancer (PCa) [1]. We previously reported that androgen treatment suppresses Sphase kinase-associated protein 2 (Skp2) and c-Myc through AR in LNCaP 104-R2 cells, inducing G1 cell cycle arrest and growth inhibition [15]. Androgen treatment decreased protein expression of Skp2, Cdk2, phospho-Cdk2 Tyr15, phospho-Cdk2 Thr14, phospho-Cdk2 Thr160, Cdk7, cyclin A, cyclin H, and c-Myc in LNCaP 104R1, PC-3AR, and PC-3LNCaPAR cells (Fig. 2).

Results

Conclusion