Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2010 Oct 19;5(10):e13500.
doi: 10.1371/journal.pone.0013500.

Bypass mechanisms of the androgen receptor pathway in therapy-resistant prostate cancer cell models

Rute B Marques  1 , Natasja F Dits, Sigrun Erkens-Schulze, Wytske M van Weerden, Guido Jenster
Affiliations

Bypass mechanisms of the androgen receptor pathway in therapy-resistant prostate cancer cell models

Rute B Marques et al. PLoS One. .

Abstract

Background: Prostate cancer is initially dependent on androgens for survival and growth, making hormonal therapy the cornerstone treatment for late-stage tumors. However, despite initial remission, the cancer will inevitably recur. The present study was designed to investigate how androgen-dependent prostate cancer cells eventually survive and resume growth under androgen-deprived and antiandrogen supplemented conditions. As model system, we used the androgen-responsive PC346C cell line and its therapy-resistant sublines: PC346DCC, PC346Flu1 and PC346Flu2.

Methodology/principal findings: Microarray technology was used to analyze differences in gene expression between the androgen-responsive and therapy-resistant PC346 cell lines. Microarray analysis revealed 487 transcripts differentially-expressed between the androgen-responsive and the therapy-resistant cell lines. Most of these genes were common to all three therapy-resistant sublines and only a minority (∼5%) was androgen-regulated. Pathway analysis revealed enrichment in functions involving cellular movement, cell growth and cell death, as well as association with cancer and reproductive system disease. PC346DCC expressed residual levels of androgen receptor (AR) and showed significant down-regulation of androgen-regulated genes (p-value = 10(-7)). Up-regulation of VAV3 and TWIST1 oncogenes and repression of the DKK3 tumor-suppressor was observed in PC346DCC, suggesting a potential AR bypass mechanism. Subsequent validation of these three genes in patient samples confirmed that expression was deregulated during prostate cancer progression.

Conclusions/significance: Therapy-resistant growth may result from adaptations in the AR pathway, but androgen-independence may also be achieved by alternative survival mechanisms. Here we identified TWIST1, VAV3 and DKK3 as potential players in the bypassing of the AR pathway, making them good candidates as biomarkers and novel therapeutical targets.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Differentially-expressed genes in PC346DCC, PC346Flu1 and PC346Flu2 sublines compared to the androgen-responsive PC346C.
PC346C was cultured in complete medium with 0.1 nM R1881, whereas the hormone-refractory sublines were culture in dextran-coated charcoal stripped medium (PC346DCC), supplemented with 1 µM of the antiandrogen hydroxyflutamide (PC346Flu1 and PC346Flu2). A) Heat-map representation: red and green colors represent up-regulation and down-regulation, respectively, whereas black indicates no difference between sublines and parental PC234C cells. Grey squares indicate missing data, either due to low expression levels, poor data quality or absence of probes for the respective transcript in the array platform used for the study. B) Venn-diagram of the number of regulated genes in the different sublines.
Figure 2
Figure 2. Activation state of the AR pathway in the PC346DCC, PC346Flu1 and PC346Flu2.
Differentially-expressed genes in PC346 hormone-refractory sublines versus parental PC346C were linked to a previously established androgen-response gene signature (see Materials and Methods section). (A) Heat-map representation of androgen-responsive genes deregulated in any of the PC346 hormone-refractory sublines. Color scheme as described in Fig. 1. (B) Venn-diagram and respective statistics.
Figure 3
Figure 3. Biological processes deregulated in the hormone-refractory sublines.
Top 5 biological functions enriched in the therapy-resistant sublines: (A) diseases and disorders, (B) molecular and cellular functions. (C) Example of Network analysis for PC346DCC showing deregulation of hormone and growth-factor receptor signaling: up-regulated genes are represented in red and repressed genes in green. Analysis was performed using Ingenuity Pathway Analysis software (www.ingenuity.com).
Figure 4
Figure 4. Expression of the androgen-independent PC346DCC signature genes in prostate cancer samples from patient tumors.
The 276-gene signature from PC346DCC was linked to data from 7 prostate cancer microarray databases of primary (Lapointe, Varambally, Tomlins, Yu), metastatic (Chandran, Lapointe, Varambally, Tomlins, Yu) and hormone-therapy refractory tumors (Tamura, Tomlins and Best). Only genes present in at least 5/7 databases (209 genes) and deregulated in at least 3/7 (111 genes) were included in the analysis. Heat-map representation of (A) 72 overexpressed and (B) 39 repressed genes in PC346DCC. (C) Deregulated genes selected for further qPCR analysis. Color scheme as described in Fig. 1. Grey squares indicate missing data, either due to low expression levels, poor data quality or absence of probes for the respective transcript in the array platform used for the study. PC-NAP: prostate cancer minus normal adjacent prostate; MET-PC: metastasis minus primary prostate tumors; HR-HN: hormone-therapy refractory minus hormone-naïve tumors.
Figure 5
Figure 5. Quantitative RT-PCR analysis of TWIST1, DKK3 and VAV3 in an independent set of prostate samples.
Prostate tumor samples were obtained by radical prostatectomy or transurethral resection of the prostate of patients being operated at Erasmus MC clinic. This panel contains 21 benign prostate tissue samples and 74 adenocarcinomas at different disease stages. (A) TWIST1; (B) DKK3; (C) VAV3 expression in prostate samples; (D) VAV3 metastasis-free survival analysis. NAP: normal adjacent prostate; PC: primary prostate cancer; LNmet: lymph node metastasis; PC-Met: non-progressive organ-confine prostate cancer; PC+Met: primary tumor from progressive prostate cancer that either had or developed metastasis during subsequent follow-up; HN: hormone-naïve; HR: hormone-therapy refractory; (*) p-value ≤0.0001 and (**) p-value ≤0.005 using Mann-Whitney two-tailed test. (***) p-value ≤0.0001 with Post linear-trend test.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Crawford ED, Eisenberger MA, McLeod DG, Spaulding JT, Benson R, et al. A controlled trial of leuprolide with and without flutamide in prostatic carcinoma. N Engl J Med. 1989;321:419–424. - PubMed
    1. Eisenberger MA, Blumenstein BA, Crawford ED, Miller G, McLeod DG, et al. Bilateral orchiectomy with or without flutamide for metastatic prostate cancer. N Engl J Med. 1998;339:1036–1042. - PubMed
    1. Jenster G. The role of the androgen receptor in the development and progression of prostate cancer. Semin Oncol. 1999;26:407–421. - PubMed
    1. van der Kwast TH, Schalken J, Ruizeveld de Winter JA, van Vroonhoven CC, Mulder E, et al. Androgen receptors in endocrine-therapy-resistant human prostate cancer. Int J Cancer. 1991;48:189–193. - PubMed
    1. Ruizeveld de Winter JA, Janssen PJ, Sleddens HM, Verleun-Mooijman MC, Trapman J, et al. Androgen receptor status in localized and locally progressive hormone refractory human prostate cancer. Am J Pathol. 1994;144:735–746. - PMC - PubMed

Publication types