Luis Beltran

Distinct Genomic Alterations in Prostate Cancers in Chinese and Western Populations Suggest Alternative Pathways of Prostate Carcinogenesis

Prostate cancer is significantly more common in Western men than in Asian men, but the basis for this difference remains unknown. Because genomic studies of Asian prostate cancer are very limited, we used a genome-wide approach to reveal the genomic alterations in Chinese prostate cancers. We found a significant reduction in the frequency of certain somatic genomic changes that are commonly found in Western prostate cancers, including the 21q22.2-22.3 deletion, which involves the TMPRSS2:ERG fusion gene, and 10q deletion, which causes PTEN inactivation. Array results were confirmed by PCR-based molecular copy-number counting in selected samples. The different frequencies of these genomic changes were further evaluated by fluorescent in situ hybridization and immunohistochemistry analyses of tissue microarray samples. These alterations might be key genetic changes underlying the regional/ethnic difference in clinical incidence and might be induced by specific environmental and/or genetic risk factors that Western men are exposed to. Our findings suggest that tumors arise in Western and Chinese populations by alternative pathogenetic mechanisms.

The safety and efficacy of sunitinib before planned nephrectomy in metastatic clear cell renal cancer

Background: The safety and efficacy of upfront sunitinib, before nephrectomy in metastatic clear cell renal cancer (mCRC), has not been prospectively evaluated. Methods: Two prospective single-arm phase II studies investigated either two cycles (study A: n = 19) or three cycles (study B: n = 33) of sunitinib before nephrectomy in mCRC. Results: Overall, 38 of 52 (73%) of patients obtained clinical benefit (by RECIST) before surgery. The partial response rate of the primary tumour was 6% [median reduction in longest diameter of 12% (range 8%−35%)]. No patients became ineligible due to local progression of disease. A nephrectomy was carried out in 37 (71%) of patients. Necrosis (>50%) was a prominent feature at nephrectomy in 49%. Surgical complications (Clavien–Dindo classification) occurred in 10 (27%) patients, including one death (3%). The median blood loss and surgical time were 725 (90–4200) ml and 189 (70–420) min, respectively. The median progression-free survival was 8 months (95% confidence interval 6–15 months). A comparison of two versus three pre-surgery cycles showed no significant difference in terms of surgical complications or efficacy. Conclusions: Nephrectomy after upfront sunitinib can be carried out safely. It obtains control of disease. Randomised studies are required to address if this approach is beneficial.

The Outcome of Patients Treated with Sunitinib Prior to Planned Nephrectomy in Metastatic Clear Cell Renal Cancer

BACKGROUND The role of cytoreductive nephrectomy in metastatic clear cell renal cell carcinoma (ccRCC) is controversial. OBJECTIVE To determine the outcome of patients with metastatic ccRCC who receive sunitinib prior to planned nephrectomy. DESIGN, SETTING, AND PARTICIPANTS The study combined the data from two prospective phase 2 studies that assessed upfront sunitinib (12-16 wk) prior to nephrectomy in previously untreated patients with metastatic renal cell carcinoma (RCC). Sunitinib was discontinued during the perioperative period (median: 29 d). INTERVENTION Sunitinib 50mg in six weekly cycles (4 wk on, 2 wk off). MEASUREMENTS Progression-free (PFS) and overall survival (OS) using the Kaplan-Meier method. RESULTS AND LIMITATIONS Twenty-one patients (32%) had Memorial Sloan-Kettering Cancer Centre (MSKCC) poor-risk disease; 45 (68%) had intermediate-risk disease. Nephrectomy was not performed in 19 (29%), most commonly due to disease progression (n = 12). The PFS for the cohort was 6.3 mo (95% confidence interval [CI], 5.1-8.5). Seventeen (36%) patients progressed during the treatment break, 13 (76%) of whom stabilised upon reinitiating of sunitinib. The OS for the cohort was 15.2 mo (95% CI, 10.3-NA). The OS for the intermediate MSKCC risk group was significantly longer than that for the poor-risk group (26.0 mo [95% CI, 13.6-NA] and 9.0 mo [95% CI, 5.8-20.5], respectively; p < 0.01). In multivariate analysis, progression of disease prior to planned nephrectomy (hazard ratio [HR]: 5.34; 95% CI, 3.17-13.27), high Fuhrman grade (HR 3.27; 95% CI, 1.38-7.72), and MSKCC poor risk at diagnosis (HR 4.75; 95% CI, 2.05-11.02) were associated with short survival (p < 0.01). However, in the absence of randomised studies it is not possible to determine if this approach is beneficial. CONCLUSIONS Upfront sunitinib prior to planned nephrectomy in intermediate-risk disease is associated with a median survival of >2 yr despite frequent progression during treatment break. Progression in metastatic sites prior to planned surgery and MSKCC poor-risk disease was associated with a poor outcome.

Absolute Quantitation of DNA Methylation of 28 Candidate Genes in Prostate Cancer Using Pyrosequencing

Aberrant DNA methylation plays a pivotal role in carcinogenesis and its mapping is likely to provide biomarkers for improved diagnostic and risk assessment in prostate cancer (PCa). We quantified and compared absolute methylation levels among 28 candidate genes in 48 PCa and 29 benign prostate hyperplasia (BPH) samples using the pyrosequencing (PSQ) method to identify genes with diagnostic and prognostic potential. RARB, HIN1, BCL2, GSTP1, CCND2, EGFR5, APC, RASSF1A, MDR1, NKX2-5, CDH13, DPYS, PTGS2, EDNRB, MAL, PDLIM4, HLAa, ESR1 and TIG1 were highly methylated in PCa compared to BPH (p < 0.001), while SERPINB5, CDH1, TWIST1, DAPK1, THRB, MCAM, SLIT2, CDKN2a and SFN were not. RARB methylation above 21% completely distinguished PCa from BPH. Separation based on methylation level of SFN, SLIT2 and SERPINB5 distinguished low and high Gleason score cancers, e.g. SFN and SERPINB5 together correctly classified 81% and 77% of high and low Gleason score cancers respectively. Several genes including CDH1 previously reported as methylation markers in PCa were not confirmed in our study. Increasing age was positively associated with gene methylation (p < 0.0001). Accurate quantitative measurement of gene methylation in PCa appears promising and further validation of genes like RARB, HIN1, BCL2, APC and GSTP1 is warranted for diagnostic potential and SFN, SLIT2 and SERPINB5 for prognostic potential.

The Effect of VEGF-Targeted Therapy on Biomarker Expression in Sequential Tissue from Patients with Metastatic Clear Cell Renal Cancer

Purpose: To investigate how biologically relevant markers change in response to antiangiogenic therapy in metastatic clear cell renal cancer (mRCC) and correlate these changes with outcome. Experimental Design: The study used sequential tumor tissue and functional imaging (taken at baseline and 12–16 weeks) obtained from three similar phase II studies. All three studies investigated the role of VEGF tyrosine kinase inhibitors (TKI) before planned nephrectomy in untreated mRCC (n = 85). The effect of targeted therapy on ten biomarkers was measured from sequential tissue. Comparative genomic hybridization (CGH) array and DNA methylation profiling (MethylCap-seq) was performed in matched frozen pairs. Biomarker expression was correlated with early progression (progression as best response) and delayed progression (between 12–16 weeks). Results: VEGF TKI treatment caused a significant reduction in vessel density (CD31), phospho-S6K expression, PDL-1 expression, and FOXP3 expression (P < 0.05 for each). It also caused a significant increase in cytoplasmic FGF-2, MET receptor expression in vessels, Fuhrman tumor grade, and Ki-67 (P < 0.05 for each). Higher levels of Ki-67 and CD31 were associated with delayed progression (P < 0.05). Multiple samples (n = 5) from the same tumor showed marked heterogeneity of tumor grade, which increased significantly with treatment. Array CGH showed extensive intrapatient variability, which did not occur in DNA methylation analysis. Conclusion: TKI treatment is associated with dynamic changes in relevant biomarkers, despite significant heterogeneity in chromosomal and protein, but not epigenetic expression. Changes to Ki-67 expression and tumor grade indicate that treatment is associated with an increase in the aggressive phenotype of the tumor. Clin Cancer Res; 19(24); 6924–34. ©2013 AACR.

Validation of an RNA cell cycle progression score for predicting death from prostate cancer in a conservatively managed needle biopsy cohort

Background:The natural history of prostate cancer is highly variable and difficult to predict accurately. Better markers are needed to guide management and avoid unnecessary treatment. In this study, we validate the prognostic value of a cell cycle progression score (CCP score) independently and in a prespecified linear combination with standard clinical variables, that is, a clinical-cell-cycle-risk (CCR) score.Methods:Paraffin sections from 761 men with clinically localized prostate cancer diagnosed by needle biopsy and managed conservatively in the United Kingdom, mostly between 2000 and 2003. The primary end point was prostate cancer death. Clinical variables consisted of centrally reviewed Gleason score, baseline PSA level, age, clinical stage, and extent of disease; these were combined into a single predefined risk assessment (CAPRA) score. Full data were available for 585 men who formed a fully independent validation cohort.Results:In univariate analysis, the CCP score hazard ratio was 2.08 (95% CI (1.76, 2.46), P<10−13) for one unit change of the score. In multivariate analysis including CAPRA, the CCP score hazard ratio was 1.76 (95% CI (1.44, 2.14), P<10−6). The predefined CCR score was highly predictive, hazard ratio 2.17 (95% CI (1.83, 2.57), χ2=89.0, P<10−20) and captured virtually all available prognostic information.Conclusions:The CCP score provides significant pretreatment prognostic information that cannot be provided by clinical variables and is useful for determining which patients can be safely managed conservatively, avoiding radical treatment.

Prognostic and Therapeutic Impact of Argininosuccinate Synthetase 1 Control in Bladder Cancer as Monitored Longitudinally by PET Imaging

Targeted therapies have yet to have significant impact on the survival of patients with bladder cancer. In this study, we focused on the urea cycle enzyme argininosuccinate synthetase 1 (ASS1) as a therapeutic target in bladder cancer, based on our discovery of the prognostic and functional import of ASS1 in this setting. ASS1 expression status in bladder tumors from 183 Caucasian and 295 Asian patients was analyzed, along with its hypothesized prognostic impact and association with clinicopathologic features, including tumor size and invasion. Furthermore, the genetics, biology, and therapeutic implications of ASS1 loss were investigated in urothelial cancer cells. We detected ASS1 negativity in 40% of bladder cancers, in which multivariate analysis indicated worse disease-specific and metastasis-free survival. ASS1 loss secondary to epigenetic silencing was accompanied by increased tumor cell proliferation and invasion, consistent with a tumor-suppressor role for ASS1. In developing a treatment approach, we identified a novel targeted antimetabolite strategy to exploit arginine deprivation with pegylated arginine deiminase (ADI-PEG20) as a therapeutic. ADI-PEG20 was synthetically lethal in ASS1-methylated bladder cells and its exposure was associated with a marked reduction in intracellular levels of thymidine, due to suppression of both uptake and de novo synthesis. We found that thymidine uptake correlated with thymidine kinase-1 protein levels and that thymidine levels were imageable with [(18)F]-fluoro-L-thymidine (FLT)-positron emission tomography (PET). In contrast, inhibition of de novo synthesis was linked to decreased expression of thymidylate synthase and dihydrofolate reductase. Notably, inhibition of de novo synthesis was associated with potentiation of ADI-PEG20 activity by the antifolate drug pemetrexed. Taken together, our findings argue that arginine deprivation combined with antifolates warrants clinical investigation in ASS1-negative urothelial and related cancers, using FLT-PET as an early surrogate marker of response.

Validation of a contemporary prostate cancer grading system using prostate cancer death as outcome

Background:Gleason scoring (GS) has major deficiencies and a novel system of five grade groups (GS⩽6; 3+4; 4+3; 8; ⩾9) has been recently agreed and included in the WHO 2016 classification. Although verified in radical prostatectomies using PSA relapse for outcome, it has not been validated using prostate cancer death as an outcome in biopsy series. There is debate whether an ‘overall’ or ‘worst’ GS in biopsies series should be used.Methods:Nine hundred and eighty-eight prostate cancer biopsy cases were identified between 1990 and 2003, and treated conservatively. Diagnosis and grade was assigned to each core as well as an overall grade. Follow-up for prostate cancer death was until 31 December 2012. A log-rank test assessed univariable differences between the five grade groups based on overall and worst grade seen, and using univariable and multivariable Cox proportional hazards. Regression was used to quantify differences in outcome.Results:Using both ‘worst’ and ‘overall’ GS yielded highly significant results on univariate and multivariate analysis with overall GS slightly but insignificantly outperforming worst GS. There was a strong correlation with the five grade groups and prostate cancer death.Conclusions:This is the largest conservatively treated prostate cancer cohort with long-term follow-up and contemporary assessment of grade. It validates the formation of five grade groups and suggests that the ‘worst’ grade is a valid prognostic measure.

High-resolution genome-wide copy-number analysis suggests a monoclonal origin of multifocal prostate cancer

Many human cancers present as multifocal lesions. Understanding the clonal origin of multifocal cancers is of both etiological and clinical importance. The molecular basis of multifocal prostate cancer has previously been explored using a limited number of isolated markers and, although independent origin is widely believed, the clonal origin of multifocal prostate cancer is still debatable. We attempted to address clonal origin using a genome‐wide copy‐number analysis of individual cancer and high‐grade prostatic intraepithelial neoplasia (HGPIN) lesions. Using Affymetrix array 6.0 copy‐number analysis, we compared the genomic changes detected in 48 individual cancer and HGPIN lesions, isolated from 18 clinically localized prostate cancer cases. Identical genomic copy‐number changes, shared by all same‐case cancer foci, were detected in all 13 informative cases displaying multiple tumor foci. In addition, individual HGPIN lesions in the two multifocal‐HGPIN cases available shared identical genomic changes. Commonly known genomic alterations, including losses at 6q15, 8p21.3‐8p21.2, 10q23.2‐10q23.31, 16q22.3, 16q23.2‐16q23.3 and 21q22.2‐21q22.3 regions and gain of 8q24.3 were the most frequently detected changes in this study and each was detected in all same‐case foci in at least one case. Microarray data were confirmed by fluorescence in situ hybridization in selected foci. Our high‐resolution genome‐wide copy‐number data suggest that many multifocal cases derive from a single prostate cancer precursor clone and that this precursor may give rise to separate HGPIN foci and may further progress to multifocal invasive prostate cancer. These findings, which demonstrate the monoclonal origin of multifocal prostate cancer, should significantly enhance our understanding of prostate carcinogenesis.

Safety and Efficacy of Pazopanib Therapy Prior to Planned Nephrectomy in Metastatic Clear Cell Renal Cancer

Importance The role of cytoreductive nephrectomy in patients with metastatic renal cancer in the era of targeted therapy is uncertain. Objective To establish the safety and efficacy of upfront pazopanib therapy prior to cytoreductive nephrectomy in previously untreated patients with metastatic clear cell renal cancer. Design, Setting, and Participants Single-arm phase 2 study of 104 previously untreated patients with metastatic clear cell renal cancer recruited between June 2008 and October 2012 at cancer treatment centers with access to nephrectomy services. The minimum follow-up was 30 months. Interventions Patients received 12 to 14 weeks of preoperative pazopanib therapy prior to planned cytoreductive nephrectomy and continued pazopanib therapy after surgery. Treatment was stopped at disease progression. Main Outcomes and Measures The primary end point was clinical benefit (using Response Evaluation Criteria in Solid Tumors, version 1.1) prior to surgery (at 12-14 weeks). Secondary end points included surgical complications, progression-free survival (PFS), overall survival (OS), and biomarker analysis. Results Of 104 patients recruited, 100 patients were assessable for clinical benefit prior to planned nephrectomy; 80 of 104 (76.9%) were men; median [interquartile range] age, 64 [56-71] years). Overall, 84 of 100 (84% [95% CI, 75%-91%]) gained clinical benefit before planned nephrectomy. The median reduction in the size of the primary tumor was 14.4% (interquartile range, 1.4%-21.1%). No patients were unable to undergo surgery as a result of local progression of disease. Nephrectomy was performed in 63 (61%) of patients; 14 (22%) reported surgical complications. The 2 most common reasons for not undergoing surgery were progression of disease (n = 13) and patient choice (n = 9). There was 1 postoperative surgical death. The median PFS and OS for the whole cohort were 7.1 (95% CI, 6.0-9.2) and 22.7 (95% CI, 14.3-not estimable) months, respectively. Patients with MSKCC poor-risk disease or progressive disease prior to surgery had a poor outcome (median OS, 5.7 [95% CI, 2.6-10.8] and 3.9 [95% CI, 0.5-9.1] months, respectively). Surgical complications were observed in 14 (22%) of the nephrectomies. Biomarker analysis from sequential tissue samples revealed a decrease in CD8 expression (20.00 vs 13.75; P = .05) and significant reduction in expression of von Hippel-Lindau tumor suppressor (100 vs 40; P < .001) and C-MET (300 vs 100; P < .001) and increased programmed cell death ligand 1 expression (0 vs 1.5; P < .001) in the immune component. No on-treatment biomarker correlated with response. Conclusions and Relevance Nephrectomy after upfront pazopanib therapy could be performed safely and was associated with good outcomes in patients with intermediate-risk metastatic clear cell renal cancer.