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

Marques RB, Dits NF, Erkens-Schulze S, van Weerden WM, Jenster G - PLoS ONE (2010)

Bottom Line: PC346DCC expressed residual levels of androgen receptor (AR) and showed significant down-regulation of androgen-regulated genes (p-value = 10(-7)).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.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Josephine Nefkens Institute, Erasmus Medical Center, Rotterdam, The Netherlands.

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.

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Related in: MedlinePlus

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.
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pone-0013500-g005: 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.

Mentions: Quantitative RT-PCR was performed on an independent set of prostate samples, obtained by radical prostatectomy or transurethral resection of the prostate of patients being operated at Erasmus MC clinic. This panel contained 21 benign prostate tissue samples and 74 adenocarcinomas at different disease stages. Quantitative PCR analysis showed up-regulation of TWIST1 in primary PCa samples and lymph node metastasis (P-value  = 0.0001 and 0.002, respectively). No difference was observed between hormone-refractory (HRPC) and hormone-naïve tumors (HNPC) (Fig. 5A). DKK3 expression was significantly decreased in PCa and lymph node metastasis (P-value ≤0.0001), although no difference was observed during progression from organ-confined to metastatic or hormone-refractory disease (Fig. 5B). VAV3 expression decreased gradually during PCa progression, with the lowest levels observed in metastatic prostate tumors (P-value  = 0.0001 for Post linear-trend test) and hormone-refractory samples (P-value  = 0.005 for HNPC vs. HRPC; Fig. 5C). Lymph node samples were removed from the VAV3 analysis in Fig. 5C, because VAV3 was highly expressed in normal lymph node compared to normal prostate tissues (data not shown). In these settings, the presence of remnants of normal lymph node tissue can lead to over-estimation of the real VAV3 quantity in lymph node metastasis. Kaplan-Meier analysis showed a direct correlation between VAV3 expression and metastasis-free survival (P-value  = 0.004 for Logrank trend test; Fig. 5D).


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

Marques RB, Dits NF, Erkens-Schulze S, van Weerden WM, Jenster G - PLoS ONE (2010)

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.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2957443&req=5

pone-0013500-g005: 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.
Mentions: Quantitative RT-PCR was performed on an independent set of prostate samples, obtained by radical prostatectomy or transurethral resection of the prostate of patients being operated at Erasmus MC clinic. This panel contained 21 benign prostate tissue samples and 74 adenocarcinomas at different disease stages. Quantitative PCR analysis showed up-regulation of TWIST1 in primary PCa samples and lymph node metastasis (P-value  = 0.0001 and 0.002, respectively). No difference was observed between hormone-refractory (HRPC) and hormone-naïve tumors (HNPC) (Fig. 5A). DKK3 expression was significantly decreased in PCa and lymph node metastasis (P-value ≤0.0001), although no difference was observed during progression from organ-confined to metastatic or hormone-refractory disease (Fig. 5B). VAV3 expression decreased gradually during PCa progression, with the lowest levels observed in metastatic prostate tumors (P-value  = 0.0001 for Post linear-trend test) and hormone-refractory samples (P-value  = 0.005 for HNPC vs. HRPC; Fig. 5C). Lymph node samples were removed from the VAV3 analysis in Fig. 5C, because VAV3 was highly expressed in normal lymph node compared to normal prostate tissues (data not shown). In these settings, the presence of remnants of normal lymph node tissue can lead to over-estimation of the real VAV3 quantity in lymph node metastasis. Kaplan-Meier analysis showed a direct correlation between VAV3 expression and metastasis-free survival (P-value  = 0.004 for Logrank trend test; Fig. 5D).

Bottom Line: PC346DCC expressed residual levels of androgen receptor (AR) and showed significant down-regulation of androgen-regulated genes (p-value = 10(-7)).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.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Josephine Nefkens Institute, Erasmus Medical Center, Rotterdam, The Netherlands.

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.

Show MeSH
Related in: MedlinePlus