Gene expression profiling in bladder cancer identifies potential therapeutic targets.

Despite advances in management, bladder cancer remains a major cause of cancer related complications. Characterisation of gene expression patterns in bladder cancer allows the identification of pathways involved in its pathogenesis, and may stimulate the development of novel therapies targeting these pathways. Between 2004 and 2005, cystoscopic bladder biopsies were obtained from 19 patients and 11 controls. These were subjected to whole transcript-based microarray analysis. Unsupervised hierarchical clustering was used to identify samples with similar expression profiles. Hypergeometric analysis was used to identify canonical pathways and curated networks having statistically significant enrichment of differentially expressed genes. Osteopontin (OPN) expression was validated by immunohistochemistry. Hierarchical clustering defined signatures, which differentiated between cancer and healthy tissue, muscle-invasive or non-muscle invasive cancer and healthy tissue, grade 1 and grade 3. Pathways associated with cell cycle and proliferation were markedly upregulated in muscle-invasive and grade 3 cancers. Genes associated with the classical complement pathway were downregulated in non-muscle invasive cancer. Osteopontin was markedly overexpressed in invasive cancer compared to healthy tissue. The present study contributes to a growing body of work on gene expression signatures in bladder cancer. The data support an important role for osteopontin in bladder cancer, and identify several pathways worthy of further investigation.

Association of oestrogen receptor beta 2 (ER beta 2/ER beta cx) with outcome of adjuvant endocrine treatment for primary breast cancer--a retrospective study.

BACKGROUND: Oestrogen receptor beta (ERbeta) modulates ERalpha activity; wild type ERbeta (ERbeta1) and its splice variants may therefore impact on hormone responsiveness of breast cancer. ERbeta2/ERbetacx acts as a dominant negative inhibitor of ERalpha and expression of ERbeta2 mRNA has been proposed as a candidate marker for outcome in primary breast cancer following adjuvant endocrine therapy. We therefore now assess ERbeta2 protein by immunostaining and mRNA by quantitative RT-PCR in relation to treatment outcome. METHODS: ERbeta2-specific immunostaining was quantified in 141 primary breast cancer cases receiving adjuvant endocrine therapy, but no neoadjuvant therapy or adjuvant chemotherapy. The expression of mRNA for ERbeta2/ERbetacx was measured in 100 cases by quantitative RT-PCR. Statistical analysis of breast cancer relapse and breast cancer survival was performed using Kaplan Meier log-rank tests and Cox's univariate and multivariate survival analysis. RESULTS: High ERbeta2 immunostaining (Allred score >5) and high ERbeta2 mRNA levels were independently associated with significantly better outcome across the whole cohort, including both ERalpha positive and negative cases (Log-Rank P < 0.05). However, only ERbeta2 mRNA levels were significantly associated with better outcome in the ERalpha + subgroup (Log-Rank P = 0.01) and this was independent of grade, size, nodal status and progesterone receptor status (Cox hazard ratio 0.31 P = 0.02 for relapse; 0.17 P = 0.01 for survival). High ERbeta2 mRNA was also associated with better outcome in node negative cases (Log Rank P < 0.001).ERbeta2 protein levels were greater in ERalpha positive cases (T-test P = 0.00001), possibly explaining the association with better outcome. Levels of ERbeta2 protein did not correlate ERbeta2 mRNA levels, but 34% of cases had both high mRNA and protein and had a significantly better outcome (Log-Rank relapse P < 0.005). CONCLUSION: High ERbeta2 protein levels were associated with ERalpha expression. Although most cases with high ERbeta2 mRNA had strong ERbeta2 immunostaining, mRNA levels but not protein levels were independently predictive of outcome in tamoxifen-treated ERalpha + tumours. Post-transcriptional control needs to be considered when assessing the biological or clinical importance of ERbeta proteins.

Prognostic significance of osteopontin expression in human prostate cancer.

To test the hypothesis that expression of osteopontin (OPN), an integrin-binding glycoprotein, can independently predict the potential aggressiveness of prostate cancer, the status of OPN expression in benign and malignant prostate cancer cell lines and tissues was analysed by Western blot and immunohistochemistry. Amongst the four prostate cell lines analysed, the level of OPN expressed in the benign PNT-2 cells was set at 1, the relative level of OPN expressed in the weakly malignant cell line LNCaP was increased to 1.5. In the highly malignant cell lines Du-145 and PC-3, the level of OPN expression was further increased to 2.9 and 4.4, respectively. An increased expression of OPN was also observed in the prostate tissue samples. When the level of OPN in normal tissue was set at 1, its level in benign prostate hyperplasia (BPH) was similar at 0.99 +/- 0.2, whereas the OPN level in the highly malignant carcinoma tissue was greatly increased by nearly 6-fold to 5.9 +/- 0.3. Amongst the 116 cases examined immunocytochemically, of the 10 normal cases, 3 (30%) were unstained and 7 (70%) stained weakly positive (+). Amongst the 36 BPH samples, 32 (89%) stained weakly positive (+) and 4 (11%) were unstained (-). For the 70 carcinomas analysed, 31 (44%) stained strongly positive (+++), 20 (29%) stained moderately positive (++) and 19 (27%) stained weakly positive (+). These results showed that the level of OPN expressed between the normal and the BPH samples was not significantly different (Fisher's exact test, p = 0.16). However, in comparison to that in the BPH samples, the expression of OPN in the carcinoma tissues was significantly increased (Chi-square test, p < 0.0001). Kaplan-Meier survival analysis showed that the increased level of OPN expression was significantly (n = 70, p = 0.03) associated with reduced survival time of the patients. The OPN expression was increased with the increasing Gleason scores of the carcinomas (Chi-square test, p < 0.001). The results in our study support our hypothesis and suggest that the increased OPN level may be involved in the malignant transformation of prostate epithelial cells and OPN expression level is an important determinant for patient survival.