Table 1.
Clinical characteristics of patients with prostate cancer bone metastases obtained at orthopedic surgery.
Table 2.
Immunohistochemical data of castration resistant bone metastases stratified according to expression of AR splice variants (AR-V).
Table 3.
Detection of the full length androgen receptor (ARfl) RNA transcript and AR splice variants in prostate tissue and bone metastases.
Figure 1.
Normalized mRNA levels of the A) full length androgen receptor (ARfl), B) AR-V1 [14], and C) AR-V7 [14] in a total of 66 tissue samples from prostate cancer patients.
All mRNA levels were adjusted for corresponding housekeeping gene (RPL13a) mRNA levels, normalized to the median value for the primary tumors samples, and transformed by the natural logarithm. Levels in non-malignant prostate tissue (n = 13), primary prostate tumor (n = 13), hormone-naïve bone metastases (n = 10), and castration-resistant bone metastases (n = 30) samples are shown in white, light grey, dark grey, and black, respectively. Undetectable levels are not indicated, but represented by columns symbolizing the lowest measurable level in RT-PCR analysis of each transcript variant.
Figure 2.
Western blot analysis of the full length androgen receptor (ARfl) and ligand binding domain (LBD) truncated AR variants in castration resistant prostate cancer (CRPC) bone metastasis samples.
A) Antibodies (N-20 and PG-21) targeting the N-terminal domain (NTD) of the AR detected the ARfl and AR variants (AR-V; presumingly AR-V1, AR-V7, and/or AR-V567es). Antibody C-19 targeting the LBD detected the ARfl of ∼110 kDa, but not the LBD-truncated AR variants of approximately ∼80 kDa. B) Analysis of CRPC bone metastases representing samples with high levels of AR-V7 mRNA (levels in the highest quartile, Q4, according to RT-PCR analysis), as well as samples with AR-V7 mRNA levels in Q1, Q2, and Q3, by using the N-20 antibody. 22Rv1 cell extract served as positive control for ARfl and ∼80 kDa AR variants and a primary prostate cancer sample (P) served as negative control for the AR variants.
Figure 3.
Kaplan-Meier analysis of androgen receptor (AR) variant transcript levels in bone metastasis samples from castration-resistant prostate cancer (CRPC) patients versus cancer-specific survival after metastasis surgery.
Patients with A) AR-V7 [14] mRNA levels in the highest quartile (Q4), B) detectable AR-V567es [16] mRNA levels, and C) detectable AR-V567 and/or AR-V7 Q4 (AR-V high) mRNA levels had shorter survival than other patients with CRPC disease.
Figure 4.
Network of directly interacting gene products of differentially expressed genes between the AR-V high (detectable AR-V567 and/or AR-V7 in the highest quartile, Q4) and other castration-resistance prostate cancer (CRPC) bone metastases (P≤0.05 and fold change ≥1.5), generated by Metacore™ (Genego Inc, USA) bioinformatics analysis.
Red circles indicate higher and blue circles indicate lower transcript levels in the AR-V high than in other CRPC bone metastases. Note that many of the differentiating gene products are directly interacting via AR, C-MYC and CDK1.