Martin H. Voss

Adverse Outcomes in Clear Cell Renal Cell Carcinoma with Mutations of 3p21 Epigenetic Regulators BAP1 and SETD2 : A Report by MSKCC and the KIRC TCGA Research Network

Purpose: To investigate the impact of newly identified chromosome 3p21 epigenetic tumor suppressors PBRM1, SETD2, and BAP1 on cancer-specific survival (CSS) of 609 patients with clear cell renal cell carcinoma (ccRCC) from 2 distinct cohorts. Experimental Design: Select sequencing on 3p tumor suppressors of 188 patients who underwent resection of primary ccRCC at the Memorial Sloan-Kettering Cancer Center (MSKCC) was conducted to interrogate the genotype–phenotype associations. These findings were compared with analyses of the genomic and clinical dataset from our nonoverlapping The Cancer Genome Atlas (TCGA) cohort of 421 patients with primary ccRCC. Results: 3p21 tumor suppressors are frequently mutated in both the MSKCC (PBRM1, 30.3%; SETD2, 7.4%; BAP1, 6.4%) and the TCGA (PBRM1, 33.5%; SETD2, 11.6%; BAP1, 9.7%) cohorts. BAP1 mutations are associated with worse CSS in both cohorts [MSKCC, P = 0.002; HR 7.71; 95% confidence interval (CI)2.08–28.6; TCGA, P = 0.002; HR 2.21; 95% CI 1.35–3.63]. SETD2 are associated with worse CSS in the TCGA cohort (P = 0.036; HR 1.68; 95% CI 1.04–2.73). On the contrary, PBRM1 mutations, the second most common gene mutations of ccRCC, have no impact on CSS. Conclusion: The chromosome 3p21 locus harbors 3 frequently mutated ccRCC tumor suppressor genes. BAP1 and SETD2 mutations (6%–12%) are associated with worse CSS, suggesting their roles in disease progression. PBRM1 mutations (30%–34%) do not impact CSS, implicating its principal role in the tumor initiation. Future efforts should focus on therapeutic interventions and further clinical, pathologic, and molecular interrogation of this novel class of tumor suppressors. Clin Cancer Res; 19(12); 3259–67. ©2013 AACR.

Clinical and Pathologic Impact of Select Chromatin-modulating Tumor Suppressors in Clear Cell Renal Cell Carcinoma

BACKGROUND Historically, VHL was the only frequently mutated gene in clear cell renal cell carcinoma (ccRCC), with conflicting clinical relevance. Recent sequencing efforts have identified several novel frequent mutations of histone modifying and chromatin remodeling genes in ccRCC including PBRM1, SETD2, BAP1, and KDM5C. PBRM1, SETD2, and BAP1 are located in close proximity to VHL within a commonly lost (approximately 90%) 3p locus. To date, the clinical and pathologic significance of mutations in these novel candidate tumor suppressors is unknown. OBJECTIVE To determine the frequency of and render the first clinical and pathologic outcome associated with mutations of these novel candidate tumor suppressors in ccRCC. DESIGN, SETTING, AND PARTICIPANTS Targeted sequencing was performed in 185 ccRCCs and matched normal tissues from a single institution. Pathologic features, baseline patient characteristics, and follow-up data were recorded. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS The linkage between mutations and clinical and pathologic outcomes was interrogated with the Fisher exact test (for stage and Fuhrman nuclear grade) and the permutation log-rank test (for cancer-specific survival [CSS]). RESULTS AND LIMITATIONS PBRM1, BAP1, SETD2, and KDM5C are mutated at 29%, 6%, 8%, and 8%, respectively. Tumors with mutations in PBRM1 or any of BAP1, SETD2, or KDM5C (19%) are more likely to present with stage III disease or higher (p = 0.01 and p = 0.001, respectively). Small tumors (<4 cm) with PBRM1 mutations are more likely to exhibit stage III pathologic features (odds ratio: 6.4; p = 0.001). BAP1 mutations tend to occur in Fuhrman grade III-IV tumors (p = 0.052) and are associated with worse CSS (p = 0.01). Clinical outcome data are limited by the number of events. CONCLUSIONS Most mutations of chromatin modulators discovered in ccRCC are loss of function, associated with advanced stage, grade, and possibly worse CSS. Further studies validating the clinical impact of these novel mutations and future development of therapeutics remedying these tumor suppressors are warranted.

Tumor Genetic Analyses of Patients with Metastatic Renal Cell Carcinoma and Extended Benefit from mTOR Inhibitor Therapy

Purpose: Rapalogs are allosteric mTOR inhibitors and approved agents for advanced kidney cancer. Reports of clonal heterogeneity in this disease challenge the concept of targeted monotherapy, yet a small subset of patients derives extended benefit. Our aim was to analyze such outliers and explore the genomic background of extreme rapalog sensitivity in the context of intratumor heterogeneity. Experimental Design: We analyzed archived tumor tissue of 5 patients with renal cell carcinoma, who previously achieved durable disease control with rapalogs (median duration, 28 months). DNA was extracted from spatially separate areas of primary tumors and metastases. Custom target capture and ultradeep sequencing was used to identify alterations across 230 target genes. Whole-exome sequence analysis was added to investigate genes beyond this original target list. Results: Five long-term responders contributed 14 specimens to explore clonal heterogeneity. Genomic alterations with activating effect on mTOR signaling were detected in 11 of 14 specimens, offering plausible explanation for exceptional treatment response through alterations in two genes (TSC1 and MTOR). In two subjects, distinct yet functionally convergent alterations activated the mTOR pathway in spatially separate sites. In 1 patient, concurrent genomic events occurred in two separate pathway components across different tumor regions. Conclusions: Analysis of outlier cases can facilitate identification of potential biomarkers for targeted agents, and we implicate two genes as candidates for further study in this class of drugs. The previously reported phenomenon of clonal convergence can occur within a targetable pathway which might have implications for biomarker development beyond this disease and this class of agents. Clin Cancer Res; 20(7); 1955–64. ©2014 AACR.

Phase 1 trial of everolimus plus sunitinib in patients with metastatic renal cell carcinoma

Simultaneous inhibition of the vascular epithelial growth factor (VEGF) and the mammalian target of rapamycin (mTOR) pathway may improve treatment response in advanced renal cell carcinoma (RCC). Everolimus, an oral mTOR inhibitor, and sunitinib, an oral tyrosine kinase inhibitor targeting VEGF, are standard agents in the management of metastatic RCC.

An Epidemiologic and Genomic Investigation Into the Obesity Paradox in Renal Cell Carcinoma

BACKGROUND Obesity increases risk for clear-cell renal cell carcinoma (ccRCC), yet obese patients appear to experience longer survival than nonobese patients. We examined body mass index (BMI) in relation to stage, grade, and cancer-specific mortality (CSM) while considering detection bias, nutritional status, and molecular tumor features. METHODS Data were available from 2119 ccRCC patients who underwent renal mass surgery at Memorial Sloan-Kettering Cancer Center between 1995 and 2012. Logistic regression models produced associations between BMI and advanced disease. Multivariable competing risks regression models estimated associations between BMI and CSM. Somatic mutation, copy number, methylation, and expression data were examined by BMI among a subset of 126 patients who participated in the Cancer Genome Atlas Project for ccRCC using the Kruskal-Wallis or Fisher exact tests. All statistical tests were two-sided. RESULTS Obese and overweight patients were less likely to present with advanced-stage disease compared with normal-weight patients (odds ratio [OR] = 0.61, 95% confidence interval [CI] = 0.48 to 0.79 vs OR = 0.65, 95% CI = 0.51 to 0.83, respectively). Higher BMI was associated with reduced CSM in univariable analyses (P < .005). It remained statistically significant after adjustment for comorbidities and albumin level, but it became non-statistically significant after adjusting for stage and grade (P > .10). Genome-wide interrogation by BMI suggested differences in gene expression of metabolic and fatty acid genes, including fatty acid synthase (FASN), consistent with the obesity paradox. CONCLUSIONS Our findings suggest that although BMI is not an independent prognostic factor for CSM after controlling for stage and grade, tumors developing in an obesogenic environment may be more indolent.

Hereditary leiomyomatosis and renal cell carcinoma syndrome-associated renal cancer: recognition of the syndrome by pathologic features and the utility of detecting aberrant succination by immunohistochemistry.

Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) syndrome is an autosomal dominant disorder in which germline mutations of fumarate hydratase (FH) gene confer an increased risk of cutaneous and uterine leiomyomas and renal cancer. HLRCC-associated renal cancer is highly aggressive and frequently presents as a solitary mass. We reviewed the clinicopathologic features of 9 patients with renal tumors presenting as sporadic cases but who were later proven to have FH germline mutations. Histologically, all tumors showed mixed architectural patterns, with papillary as the dominant pattern in only 3 cases. Besides papillary, tubular, tubulopapillary, solid, and cystic elements, 6 of 9 tumors contained collecting duct carcinoma–like areas with infiltrating tubules, nests, or individual cells surrounded by desmoplastic stroma. Prominent tubulocystic carcinoma–like component and sarcomatoid differentiation were identified. Although all tumors exhibited the proposed hallmark of HLRCC (large eosinophilic nucleolus surrounded by a clear halo), this feature was often not uniformly present throughout the tumor. Prior studies have shown that a high level of fumarate accumulated in HLRCC tumor cells causes aberrant succination of cellular proteins by forming a stable chemical modification, S-(2-succino)-cysteine (2SC), which can be detected by immunohistochemistry. We thus explored the utility of detecting 2SC by immunohistochemistry in the differential diagnosis of HLRCC tumors and other high-grade renal tumors and investigated the correlation between 2SC staining and FH molecular alterations. All confirmed HLRCC tumors demonstrated diffuse and strong nuclear and cytoplasmic 2SC staining, whereas all clear cell (184/184, 100%), most high-grade unclassified (93/97, 96%), and the large majority of “type 2” papillary (35/45, 78%) renal cell carcinoma cases showed no 2SC immunoreactivity. A subset of papillary (22%) and rare unclassified (4%) tumors showed patchy or diffuse cytoplasmic staining without nuclear labeling, unlike the pattern seen with confirmed HLRCC tumors. Sequencing revealed no germline or somatic FH alterations in 14 tumors that either exhibited only cytoplasmic 2SC staining (n=5) or were negative for 2SC (n=9), despite their HLRCC-like morphologic features. Our results emphasize the pivotal role of pathologic examination in the diagnosis of HLRCC patients and indicate immunohistochemical detection of 2SC as a useful ancillary tool in the differentiation of HLRCC renal tumors from other high-grade renal cell carcinomas.

mTOR inhibitors in advanced renal cell carcinoma.

Better understanding of the molecular biology of renal cell carcinoma (RCC) has led to the development of several targeted anti-cancer agents, several of which have since received approval for treatment of advanced disease. Two of these, the intravenous agent temsirolimus and the oral everolimus, exhibit antitumor effects through inhibition of the mammalian target of rapamycin (mTOR) pathway. This article reviews their mechanisms of action in the context of the current understanding of RCC pathophysiology, the clinical data leading to their approval, class-specific toxicities, potential molecular mechanisms behind treatment resistance and novel treatment approaches for RCC that incorporate mTOR blockade.

OncoKB: A Precision Oncology Knowledge Base

PURPOSE With prospective clinical sequencing of tumors emerging as a mainstay in cancer care, there is an urgent need for a clinical support tool that distills the clinical implications associated with specific mutation events into a standardized and easily interpretable format. To this end, we developed OncoKB, an expert-guided precision oncology knowledge base. METHODS OncoKB annotates the biological and oncogenic effect and the prognostic and predictive significance of somatic molecular alterations. Potential treatment implications are stratified by the level of evidence that a specific molecular alteration is predictive of drug response based on US Food and Drug Administration (FDA) labeling, National Comprehensive Cancer Network (NCCN) guidelines, disease-focused expert group recommendations and the scientific literature. RESULTS To date, over 3000 unique mutations, fusions, and copy number alterations in 418 cancer-associated genes have been annotated. To test the utility of OncoKB, we annotated all genomic events in 5983 primary tumor samples in 19 cancer types. Forty-one percent of samples harbored at least one potentially actionable alteration, of which 7.5% were predictive of clinical benefit from a standard treatment. OncoKB annotations are available through a public web resource (http://oncokb.org/) and are also incorporated into the cBioPortal for Cancer Genomics to facilitate the interpretation of genomic alterations by physicians and researchers. CONCLUSION OncoKB, a comprehensive and curated precision oncology knowledge base, offers oncologists detailed, evidence-based information about individual somatic mutations and structural alterations present in patient tumors with the goal of supporting optimal treatment decisions.

Safety and Efficacy of Nivolumab in Combination With Ipilimumab in Metastatic Renal Cell Carcinoma: The CheckMate 016 Study

Purpose Combination treatment with immune checkpoint inhibitors has shown enhanced antitumor activity compared with monotherapy in tumor types such as melanoma. The open-label, parallel-cohort, dose-escalation, phase I CheckMate 016 study evaluated the efficacy and safety of nivolumab plus ipilimumab in combination, and nivolumab plus a tyrosine kinase inhibitor in metastatic renal cell carcinoma (mRCC). Safety and efficacy results from the nivolumab plus ipilimumab arms of the study are presented. Patients and Methods Patients with mRCC received intravenous nivolumab 3 mg/kg plus ipilimumab 1 mg/kg (N3I1), nivolumab 1 mg/kg plus ipilimumab 3 mg/kg (N1I3), or nivolumab 3 mg/kg plus ipilimumab 3 mg/kg (N3I3) every 3 weeks for four doses followed by nivolumab monotherapy 3 mg/kg every 2 weeks until progression or toxicity. End points included safety (primary), objective response rate, and overall survival (OS). Results All patients in the N3I3 arm (n = 6) were censored at the time of analysis as a result of dose-limiting toxicity or other reasons. Forty-seven patients were treated in both the N3I1 and the N1I3 arm, and baseline patient characteristics were balanced between arms. Grade 3 to 4 treatment-related adverse events were reported in 38.3% and 61.7% of the patients in the N3I1 and N1I3 arms, respectively. At a median follow-up of 22.3 months, the confirmed objective response rate was 40.4% in both arms, with ongoing responses in 42.1% and 36.8% of patients in the N3I1 and N1I3 arms, respectively. The 2-year OS was 67.3% and 69.6% in the N3I1 and N1I3 arms, respectively. Conclusion Nivolumab plus ipilimumab therapy demonstrated manageable safety, notable antitumor activity, and durable responses with promising OS in patients with mRCC.

Tumor immune microenvironment characterization in clear cell renal cell carcinoma identifies prognostic and immunotherapeutically relevant messenger RNA signatures.

BackgroundTumor-infiltrating immune cells have been linked to prognosis and response to immunotherapy; however, the levels of distinct immune cell subsets and the signals that draw them into a tumor, such as the expression of antigen presenting machinery genes, remain poorly characterized. Here, we employ a gene expression-based computational method to profile the infiltration levels of 24 immune cell populations in 19 cancer types.ResultsWe compare cancer types using an immune infiltration score and a T cell infiltration score and find that clear cell renal cell carcinoma (ccRCC) is among the highest for both scores. Using immune infiltration profiles as well as transcriptomic and proteomic datasets, we characterize three groups of ccRCC tumors: T cell enriched, heterogeneously infiltrated, and non-infiltrated. We observe that the immunogenicity of ccRCC tumors cannot be explained by mutation load or neo-antigen load, but is highly correlated with MHC class I antigen presenting machinery expression (APM). We explore the prognostic value of distinct T cell subsets and show in two cohorts that Th17 cells and CD8+ T/Treg ratio are associated with improved survival, whereas Th2 cells and Tregs are associated with negative outcomes. Investigation of the association of immune infiltration patterns with the subclonal architecture of tumors shows that both APM and T cell levels are negatively associated with subclone number.ConclusionsOur analysis sheds light on the immune infiltration patterns of 19 human cancers and unravels mRNA signatures with prognostic utility and immunotherapeutic biomarker potential in ccRCC.Tumor-infiltrating immune cells have been linked to prognosis and response to immunotherapy; however, the levels of distinct immune cell subsets and the signals that draw them into a tumor, such as the expression of antigen presenting machinery genes, remain poorly characterized. Here, we employ a gene expression-based computational method to profile the infiltration levels of 24 immune cell populations in 19 cancer types. We compare cancer types using an immune infiltration score and a T cell infiltration score and find that clear cell renal cell carcinoma (ccRCC) is among the highest for both scores. Using immune infiltration profiles as well as transcriptomic and proteomic datasets, we characterize three groups of ccRCC tumors: T cell enriched, heterogeneously infiltrated, and non-infiltrated. We observe that the immunogenicity of ccRCC tumors cannot be explained by mutation load or neo-antigen load, but is highly correlated with MHC class I antigen presenting machinery expression (APM). We explore the prognostic value of distinct T cell subsets and show in two cohorts that Th17 cells and CD8+ T/Treg ratio are associated with improved survival, whereas Th2 cells and Tregs are associated with negative outcomes. Investigation of the association of immune infiltration patterns with the subclonal architecture of tumors shows that both APM and T cell levels are negatively associated with subclone number. Our analysis sheds light on the immune infiltration patterns of 19 human cancers and unravels mRNA signatures with prognostic utility and immunotherapeutic biomarker potential in ccRCC.