Dominique Trudel

Spatial genomic heterogeneity within localized, multifocal prostate cancer

Herein we provide a detailed molecular analysis of the spatial heterogeneity of clinically localized, multifocal prostate cancer to delineate new oncogenes or tumor suppressors. We initially determined the copy number aberration (CNA) profiles of 74 patients with index tumors of Gleason score 7. Of these, 5 patients were subjected to whole-genome sequencing using DNA quantities achievable in diagnostic biopsies, with detailed spatial sampling of 23 distinct tumor regions to assess intraprostatic heterogeneity in focal genomics. Multifocal tumors are highly heterogeneous for single-nucleotide variants (SNVs), CNAs and genomic rearrangements. We identified and validated a new recurrent amplification of MYCL, which is associated with TP53 deletion and unique profiles of DNA damage and transcriptional dysregulation. Moreover, we demonstrate divergent tumor evolution in multifocal cancer and, in some cases, tumors of independent clonal origin. These data represent the first systematic relation of intraprostatic genomic heterogeneity to predicted clinical outcome and inform the development of novel biomarkers that reflect individual prognosis.

The influence of MMP-14, TIMP-2 and MMP-2 expression on breast cancer prognosis

IntroductionMatrix metalloproteinase (MMP)-2 is very active at degrading extracellular matrix. It is under the influence of an activator, membrane type 1 MMP (MMP-14), and the tissue inhibitor of metalloproteases (TIMP)-2. We hypothesized that the individual expression of these three markers or their balance may help to predict breast cancer prognosis.MethodsMMP-2, MMP-14 and TIMP-2 expression has been evaluated by 35S mRNA in situ hybridization on paraffin material of 539 breast cancers without distant metastasis at diagnosis and with a median follow-up of 9.2 years.ResultsMMP-2 and MMP-14 mRNA was detected primarily in reactive stromal cells whereas TIMP-2 mRNA was expressed by both stromal and cancer cells. Of the three molecules, an adjusted Cox model revealed that high MMP-14 mRNA (≥ 10% cells) alone predicted a significantly shorter overall survival (p = 0.031) when adjusted for clinical factors (tumor size and number of involved lymph nodes). Prognostic significance was lost when further adjusted for Her-2/neu and urokinase-type plasminogen activator (p = 0.284). Furthermore, when all three components were analyzed together, the survival was worst for patients with high MMP-2/high MMP-14/low TIMP-2 (5 year survival = 60%) and best with low MMP-2/low MMP-14/high TIMP-2 (5 year survival = 74%), but the difference did not reach statistical significance (p = 0.3285).ConclusionOf the MMP-14/TIMP-2/MMP-2 complex, MMP-14 was the factor most significantly associated with the outcome of breast cancer and was an independent factor of poor overall survival when adjusted for clinical prognostic factors, but not for certain ancillary markers.

Genomic hallmarks of localized, non-indolent prostate cancer

Prostate tumours are highly variable in their response to therapies, but clinically available prognostic factors can explain only a fraction of this heterogeneity. Here we analysed 200 whole-genome sequences and 277 additional whole-exome sequences from localized, non-indolent prostate tumours with similar clinical risk profiles, and carried out RNA and methylation analyses in a subset. These tumours had a paucity of clinically actionable single nucleotide variants, unlike those seen in metastatic disease. Rather, a significant proportion of tumours harboured recurrent non-coding aberrations, large-scale genomic rearrangements, and alterations in which an inversion repressed transcription within its boundaries. Local hypermutation events were frequent, and correlated with specific genomic profiles. Numerous molecular aberrations were prognostic for disease recurrence, including several DNA methylation events, and a signature comprised of these aberrations outperformed well-described prognostic biomarkers. We suggest that intensified treatment of genomically aggressive localized prostate cancer may improve cure rates.

Prognostic impact of intraductal carcinoma and large cribriform carcinoma architecture after prostatectomy in a contemporary cohort

BACKGROUND Intraductal carcinoma (IDC) of prostate is a distinct entity associated with higher Gleason score and poor prognosis. The prognostic significance of large cribriform Gleason pattern 4 (LC) in conjunction with IDC has not been previously investigated. The aim of our study was to determine the impact of IDC and LC on biochemical recurrence-free rate (bRFR) in a contemporary prostatectomy cohort. METHODS Prostate cancers of 246 prostatectomies, median follow-up 130.6 months, were graded with the International Society of Urological Pathology (ISUP) 2005 modified Gleason score (GS) and assessed for the presence of LC and/or IDC. In 57 cases with LC and/or IDC, immunostaining was performed to distinguish LC and IDC. The Kaplan-Meier (KM) method was used to estimate 5-year bRFR probabilities. Cox proportional hazards models were used to generate hazard ratios (HRs) and 95% confidence intervals (CIs). RESULTS Multivariable analysis showed that the presence of any amount of LC or IDC had a highly significant prognostic effect on bRFR (HR 2.98, 95% CI: 1.68-5.28, p=0.0002) after adjusting for GS, surgical margin status and pathological stage. Although IDC alone tended to be associated with a worse prognosis, LC and IDC did not appear to be associated with a difference in bRFR when analysed separately. CONCLUSION We demonstrate that the presence of any amount of LC/IDC is a significant prognostic factor after adjusting for Gleason score and T stage in determining patient outcome and we advocate including the presence of either in routine pathology reporting.

Randomized clinical trial of vitamin D3 doses on prostatic vitamin D metabolite levels and ki67 labeling in prostate cancer patients.

CONTEXT Vitamin D3 might benefit prostate cancer (PCa) patients because prostate cells can locally synthesize the active hormone calcitriol. OBJECTIVE Our objective was to determine the effects of oral vitamin D3 on vitamin D metabolites and PCa proliferative activity in prostate tissue. DESIGN AND SETTING We conducted a double-blind randomized clinical trial at surgical oncology clinics in Toronto, Canada. PATIENTS PCa patients (Gleason 6 or 7) participated in the study. Of 66 subjects who were enrolled, 63 completed the dosing protocol. INTERVENTION Vitamin D3 (400, 10 000, or 40 000 IU/d) was orally administered before radical prostatectomy. MAIN OUTCOME MEASURES We evaluated vitamin D metabolite levels and Ki67 labeling in surgical prostate tissue. Safety measures, PTH, and prostate-specific antigen (PSA) were also assessed. RESULTS Prostate tissue and serum levels of vitamin D metabolites, including calcitriol, increased dose dependently (P < .03) and were significantly higher in the 40 000-IU/d group than in every other dose group (P < .03). Prostate vitamin D metabolites correlated positively with serum levels (P < .0001). Ki67 measures did not differ significantly among vitamin D dose groups. However, cross-sectional analysis indicated that the calcitriol level attained in prostate was inversely associated with Ki67 intensity and Ki67 (3+) percent positive nuclei in PCa and benign tissue (P < .05). Safety measures did not change adversely with dosing. Compared with the 400-IU/d group, serum PTH and PSA were lower in the combined higher-dose groups at the end of the study (P < .02). CONCLUSIONS Oral vitamin D3 raised prostate calcitriol levels (level 1 evidence) and modestly lowered both PSA and PTH. Although Ki67 expression did not differ among dose groups, its levels correlated inversely with prostate calcitriol. These suggestions of clinical benefit justify continued clinical research.

Membrane-type-1 matrix metalloproteinase, matrix metalloproteinase 2, and tissue inhibitor of matrix proteinase 2 in prostate cancer: identification of patients with poor prognosis by immunohistochemistry.

Overexpression of the matrix metalloproteinase (MMP) 2 is associated with poor prognosis in many tumor types. Membrane-type-1 MMP (MMP14) activates MMP2 using pro-MMP2 specific inhibitor, tissue inhibitor of matrix proteinase 2 (TIMP2), as a receptor. We evaluated, by immunohistochemistry on 189 T3N0-2M0 prostate cancer (Pca) cases, the influence of MMP2, MMP14, and TIMP2 expression, individually and in association, on Pca disease-free survival (DFS). We evaluated marker expression separately in cancer, stromal, and benign epithelial (BE) cells according to a percentage scale (0%, <10%, 10%-50%, and >50%). Median follow-up was 4.61 years. In BE cells, there was an inverse relationship between initial prostate-specific antigen serum level and T3 stage with MMP14 expression (P = .003) and between pN stage and TIMP2 expression (P = .04). The most significant results with survival were obtained by dichotomizing the cases between those with less than 10% and at least 10% of cells expressing the marker, the latter category representing overexpression. TIMP2 overexpression in stromal cells was associated with a longer DFS with a hazard ratio of 0.573 (P = .02) for time to recurrence. MMP2 overexpression by BE cells correlated with a shorter DFS using a multivariate trend test (hazard ratio = 1.46, P = .02). Stromal cells expressing less than 10% TIMP2 and MMP2 overexpression was the only combination that was significantly associated with a shorter DFS (log-rank test, P = .0001). This study suggests that MMP14 is involved mostly in Pca implantation and that MMP2 and TIMP2 expression by reactive stromal cells might be used as predictors of DFS in T3N0-2M0 Pca.

Proteomic Profiling of Androgen-independent Prostate Cancer Cell Lines Reveals a Role for Protein S during the Development of High Grade and Castration-resistant Prostate Cancer

Background: The mechanisms that cause castration-resistant prostate cancer remain unknown. Results: Using high throughput proteomics and subsequent clinical validation, we identified Protein S as being elevated in high grade/advanced prostate cancer. Conclusion: Protein S is elevated in aggressive prostate cancer. Significance: Protein S expression could serve as a biomarker of aggressive prostate cancer. Androgen deprivation constitutes the principal therapy for advanced and metastatic prostate cancers. However, this therapeutic intervention usually results in the transition to a more aggressive androgen-independent prostate cancer. The elucidation of molecular alterations during the progression to androgen independence is an integral step toward discovering more effective targeted therapies. With respect to identifying crucial mediators of this transition, we compared the proteomes of androgen-independent (PC3, DU145, PPC1, LNCaP-SF, and 22Rv1) and androgen-dependent (LNCaP and VCaP) and/or normal prostate epithelial (RWPE) cell lines using mass spectrometry. We identified more than 100 proteins that were differentially secreted in the androgen-independent cell lines. Of these, Protein S (PROS1) was elevated in the secretomes of all of the androgen-independent prostate cancer cell lines, with no detectable secretion in normal and androgen-dependent cell lines. Using quantitative PCR, we observed significantly higher (p < 0.05) tissue expression levels of PROS1 in prostate cancer samples, further indicating its importance in prostate cancer progression. Similarly, immunohistochemistry analysis revealed elevation of PROS1 in high grade prostate cancer (Gleason grade ≥8), and further elevation in castration-resistant metastatic prostate cancer lesions. We also observed its significant (p < 0.05) elevation in high grade prostate cancer seminal plasma samples. Taken together, these results show that PROS1 is elevated in high grade and castration-resistant prostate cancer and could serve as a potential biomarker of aggressive disease.

Correlation of ERG Expression and DNA Methylation Biomarkers with Adverse Clinicopathologic Features of Prostate Cancer

Purpose: Fusion of the TMPRSS2 gene with the ERG oncogene and aberrant DNA methylation patterns are commonly found in prostate cancer. The aim of this study was to analyze the relationship between ERG expression, DNA methylation of three biomarkers, and clinicopathologic features of prostate cancer. Experimental Design: Immunohistochemistry for ERG protein was conducted as a surrogate for TMPRSS2-ERG fusions. We analyzed methylation of CYP26A1, TBX15, and HOXD3 in 219 prostatectomy specimens by the quantitative MethyLight assay. DNA methylation was compared between ERG-positive and -negative cases and correlations of ERG and DNA methylation with clinicopathologic features were analyzed using χ2, Spearman correlation, logistic regression, and Cox regression. Results: ERG expression varied according to Gleason pattern (almost absent in pattern II, highest in pattern III, and lower in pattern IV/V) and showed a strong positive correlation with methylation levels of CYP26A1, TBX15, and HOXD3 (Spearman P < 0.005). TBX15 and HOXD3 methylation were significantly associated with pathologic stage, Gleason score, and Gleason pattern (P ≤ 0.015). In multivariate regression analysis, PSA, TBX15 high methylation, and HOXD3 high methylation were significantly associated with stage (P < 0.05), whereas ERG expression was negatively correlated with Gleason score (P = 0.003). In univariate time-to-recurrence analysis, a combination of HOXD3/TBX15 high methylation predicted recurrence in ERG-positive and -negative cases (P < 0.05). Conclusions: CYP26A1, TBX15, and HOXD3 are methylation markers of prostate cancer associated with ERG expression and clinicopathologic variables, suggesting that incorporation of these markers may be useful in a pre- and posttreatment clinical setting. Clin Cancer Res; 18(10); 2896–904. ©2012 AACR.

Green tea for ovarian cancer prevention and treatment: A systematic review of the in vitro, in vivo and epidemiological studies

OBJECTIVE This systematic review was conducted to examine the effects of green tea or green tea components on the prevention and progression of epithelial ovarian cancer. METHODS Using Medline, EMBASE and SciVerse (last researched: July 2011), we retrieved 22 articles including 5 epidemiological studies. RESULTS In epithelial ovarian cancer cell lines, green tea and green tea components have been shown to downregulate the expression of proteins involved in inflammation, cell signalization, cell motility and angiogenesis. Green tea and green tea components would induce apoptosis and could potentiate the effects of cisplatin, a chemotherapeutic agent. In human observational studies, significant associations between green tea intake and both decreased ovarian cancer occurrence and better prognosis were reported. CONCLUSIONS Available literature suggests potential molecular targets for green tea in ovarian cancer treatment and also provides data supporting the clinical evaluation of the role of green tea or green tea components in ovarian cancer prevention and treatment.

Visual and automated assessment of matrix metalloproteinase-14 tissue expression for the evaluation of ovarian cancer prognosis.

The purpose of this study was to evaluate whether the membrane type 1 matrix metalloproteinase-14 (or MT1-MMP) tissue expression, as assessed visually on digital slides and by digital image analysis, could predict outcomes in women with ovarian carcinoma. Tissue microarrays from a cohort of 211 ovarian carcinoma women who underwent a debulking surgery between 1993 and 2006 at the CHU de Québec (Canada) were immunostained for matrix metalloproteinase-14. The percentage of MMP-14 staining was assessed visually and with the Calopix software. Progression was evaluated using the CA-125 and/or the RECIST criteria according to the GCIG criteria. Dates of death were obtained by record linkage with the Québec mortality files. Adjusted hazard ratios of death and progression with their 95% confidence intervals were estimated using the Cox model. Comparisons between the two modalities of MMP-14 assessment were done using the box plots and the Kruskal–Wallis test. The highest levels of MMP-14 immunostaining were associated with nonserous histology, early FIGO stage, and low preoperative CA-125 levels (P<0.05). In bivariate analyses, the higher level of MMP-14 expression (>40% of MMP-14-positive cells) was inversely associated with progression using visual assessment (hazard ratio=0.39; 95% confidence interval: 0.18–0.82). A similar association was observed with the highest quartile of MMP-14-positive area assessed by digital image analysis (hazard ratio=0.48; 95% confidence interval: 0.28–0.82). After adjustment for standard prognostic factors, these associations were no longer significant in the ovarian carcinoma cohort. However, in women with serous carcinoma, the highest quartile of MMP-14-positive area was associated with progression (adjusted hazard ratio=0.48; 95% confidence interval: 0.24–0.99). There was no association with overall survival. The digital image analysis of MMP-14-positive area matched the visual assessment using three categories (>40% vs 21–40 vs <20%). Higher levels of MMP-14 immunostaining were associated with standard factors of better ovarian carcinoma prognosis. In women with serous carcinoma, high expression of MMP-14 was associated with lower progression.