Erin Flores

BAP1 cancer syndrome: malignant mesothelioma, uveal and cutaneous melanoma, and MBAITs

BackgroundBRCA1–associated protein 1 (BAP1) is a tumor suppressor gene located on chromosome 3p21. Germline BAP1 mutations have been recently associated with an increased risk of malignant mesothelioma, atypical melanocytic tumors and other neoplasms. To answer the question if different germline BAP1 mutations may predispose to a single syndrome with a wide phenotypic range or to distinct syndromes, we investigated the presence of melanocytic tumors in two unrelated families (L and W) with germline BAP1 mutations and increased risk of malignant mesothelioma.MethodsSuspicious cutaneous lesions were clinically and pathologically characterized and compared to those present in other families carrying BAP1 mutations. We then conducted a meta-analysis of all the studies reporting BAP1-mutated families to survey cancer risk related to the germline BAP1 mutation (means were compared using t-test and proportions were compared with Pearson χ2 test or two-tailed Fisher’s exact test).ResultsMelanocytic tumors: of the five members of the L family studied, four (80%) carried a germline BAP1 mutation (p.Gln684*) and also presented one or more atypical melanocytic tumors; of the seven members of W family studied, all carried a germline BAP1 mutation (p.Pro147fs*48) and four of them (57%) presented one or more atypical melanocytic tumors, that we propose to call “melanocytic BAP1-mutated atypical intradermal tumors” (MBAITs). Meta-analysis: 118 individuals from seven unrelated families were selected and divided into a BAP1-mutated cohort and a BAP1-non-mutated cohort. Malignant mesothelioma, uveal melanoma, cutaneous melanoma, and MBAITs prevalence was significantly higher in the BAP1-mutated cohort (p ≤ 0.001).ConclusionsGermline BAP1 mutations are associated with a novel cancer syndrome characterized by malignant mesothelioma, uveal melanoma, cutaneous melanoma and MBAITs, and possibly by other cancers. MBAITs provide physicians with a marker to identify individuals who may carry germline BAP1 mutations and thus are at high risk of developing associated cancers.

High Incidence of Somatic BAP1 Alterations in Sporadic Malignant Mesothelioma

Background: Breast cancer 1-associated protein 1 (BAP1) is a nuclear deubiquitinase that regulates gene expression, transcription, DNA repair, and more. Several findings underscore the apparent driver role of BAP1 in malignant mesothelioma (MM). However, the reported frequency of somatic BAP1 mutations in MM varies considerably, a discrepancy that appeared related to either methodological or ethnical differences across various studies. Methods: To address this discrepancy, we carried out comprehensive genomic and immunohistochemical (IHC) analyses to detect somatic BAP1 gene alterations in 22 frozen MM biopsies from U.S. MM patients. Results: By combining Sanger sequencing, multiplex ligation-dependent probe amplification, copy number analysis, and cDNA sequencing, we found alteration of BAP1 in 14 of 22 biopsies (63.6%). No changes in methylation were observed. IHC revealed normal nuclear BAP1 staining in the eight MM containing wild-type BAP1, whereas no nuclear staining was detected in the 14 MM biopsies containing tumor cells with mutated BAP1. Thus, IHC results were in agreement with those obtained by genomic analyses. We then extended IHC analysis to an independent cohort of 70 MM biopsies, of which there was insufficient material to perform molecular studies. IHC revealed loss of BAP1 nuclear staining in 47 of these 70 MM biopsies (67.1%). Conclusions: Our findings conclusively establish BAP1 as the most commonly mutated gene in MM, regardless of ethnic background or other clinical characteristics. Our data point to IHC as the most accessible and reliable technique to detect BAP1 status in MM biopsies.

Mesothelioma patients with germline BAP1 mutations have 7-fold improved long-term survival

BRCA1-associated protein-1 (BAP1) mutations cause a new cancer syndrome, with a high rate of malignant mesothelioma (MM). Here, we tested the hypothesis that MM associated with germline BAP1 mutations has a better prognosis compared with sporadic MM. We compared survival among germline BAP1 mutation MM patients with that of all MM (N = 10 556) recorded in the United States Surveillance, Epidemiology, and End Results (SEER) data from 1973 to 2010. We identified 23 MM patients--11 alive--with germline BAP1 mutations and available data on survival. Ten patients had peritoneal MM, ten pleural MM and three MM in both locations. Thirteen patients had one or more malignancies in addition to MM. Actuarial median survival for the MM patients with germline BAP1 mutations was 5 years, as compared with <1 year for the median survival in the United States SEER MM group. Five-year survival was 47%, 95% confidence interval (24-67%), as compared with 6.7% (6.2-7.3%) in the control SEER group. Analysis of the pooled cohort of germline BAP1 mutation MM showed that patients with peritoneal MM (median survival of 10 years, P = 0.0571), or with a second malignancy in addition to MM (median survival of 10 years, P = 0.0716), survived for a longer time compared with patients who only had pleural MM, or MM patients without a second malignancy, respectively. In conclusion, we found that MM patients with germline BAP1 mutations have an overall 7-fold increased long-term survival, independently of sex and age. Appropriate genetic counseling and clinical management should be considered for MM patients who are also BAP1 mutation carriers.

Minimal asbestos exposure in germline BAP1 heterozygous mice is associated with deregulated inflammatory response and increased risk of mesothelioma

Germline BAP1 mutations predispose to several cancers, in particular malignant mesothelioma. Mesothelioma is an aggressive malignancy generally associated with professional exposure to asbestos. However, to date, we found that none of the mesothelioma patients carrying germline BAP1 mutations were professionally exposed to asbestos. We hypothesized that germline BAP1 mutations might influence the asbestos-induced inflammatory response that is linked to asbestos carcinogenesis, thereby increasing the risk of developing mesothelioma after minimal exposure. Using a BAP1+/− mouse model, we found that, compared with their wild-type littermates, BAP1+/− mice exposed to low-dose asbestos fibers showed significant alterations of the peritoneal inflammatory response, including significantly higher levels of pro-tumorigenic alternatively polarized M2 macrophages, and lower levels of several chemokines and cytokines. Consistent with these data, BAP1+/− mice had a significantly higher incidence of mesothelioma after exposure to very low doses of asbestos, doses that rarely induced mesothelioma in wild-type mice. Our findings suggest that minimal exposure to carcinogenic fibers may significantly increase the risk of malignant mesothelioma in genetically predisposed individuals carrying germline BAP1 mutations, possibly via alterations of the inflammatory response.

Combined Genetic and Genealogic Studies Uncover a Large BAP1 Cancer Syndrome Kindred Tracing Back Nine Generations to a Common Ancestor from the 1700s

We recently discovered an inherited cancer syndrome caused by BRCA1-Associated Protein 1 (BAP1) germline mutations, with high incidence of mesothelioma, uveal melanoma and other cancers and very high penetrance by age 55. To identify families with the BAP1 cancer syndrome, we screened patients with family histories of multiple mesotheliomas and melanomas and/or multiple cancers. We identified four families that shared an identical BAP1 mutation: they lived across the US and did not appear to be related. By combining family histories, molecular genetics, and genealogical approaches, we uncovered a BAP1 cancer syndrome kindred of ~80,000 descendants with a core of 106 individuals, whose members descend from a couple born in Germany in the early 1700s who immigrated to North America. Their descendants spread throughout the country with mutation carriers affected by multiple malignancies. Our data show that, once a proband is identified, extended analyses of these kindreds, using genomic and genealogical studies to identify the most recent common ancestor, allow investigators to uncover additional branches of the family that may carry BAP1 mutations. Using this knowledge, we have identified new branches of this family carrying BAP1 mutations. We have also implemented early-detection strategies that help identify cancers at early-stage, when they can be cured (melanomas) or are more susceptible to therapy (MM and other malignancies).

Abstract 1179: Combined genetic and genealogic studies uncover a large BAP1 cancer syndrome kindred, tracing back nine generations to a common ancestor from the 1700s

Germline BAP1 mutations cause a cancer syndrome characterized by high incidence of mesothelioma (MM), uveal melanoma and other cancers, and by very high penetrance, as all individuals carrying BAP1 mutations developed at least one, and usually several, malignancies throughout their lives. Through screening MM patients with histories of multiple cancers, we found four supposedly unrelated patients that shared an identical germline BAP1 mutation. We investigated whether this BAP1 mutation occurred in a ‘hot-spot’ for “de novo” mutations or whether these four MM patients shared a common ancestor. Using molecular genomics analyses we found that they are related. By genealogic studies we traced their ancestor to a couple that emigrated from Germany to North America in the early 1700’s; we traced the subsequent migration of their descendants, who are now living in at least three different US States. Our findings demonstrate that BAP1 mutations are transmitted among subsequent generations over the course of centuries. This knowledge and methodology is being used to identify additional branches of the family carrying BAP1 mutations. Our study shows that the application of modern genomic analyses, coupled with “classical” family histories collected by the treating physician, and with genealogical searches, offer a powerful strategy to identify high-risk germline BAP1 mutation carriers that will benefit from genetic counseling and early detection cancer screening. Citation Format: Michele Carbone, Erin G. Flores, Mitsuru Emi, Giovanni Gaudino, Sandra Pastorino, Haining Yang, Todd Johnson, Tatsuhiko Tsunoda, Mary Hesdorffer, Harvey I. Pass. Combined genetic and genealogic studies uncover a large BAP1 cancer syndrome kindred, tracing back nine generations to a common ancestor from the 1700s. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1179.

Abstract 4796: A founder mutation in the BAP1 gene among four caucasian families with high incidences of malignant peritoneal mesothelioma and uveal melanoma: a molecular and genealogical study in a 10-generation BAP1 cancer syndrome kindred

The BAP1 cancer syndrome is characterized by a high incidence of malignant mesothelioma (MM), uveal melanoma (UM), cutaneous melanoma (CM), clear cell renal cell carcinoma (ccRCC), and it is expected that the clinical phenotype will continue to broaden in scope. Molecular screening for BAP1 gene mutations among 29 patients selected for clinically apparent familial MM led to the identification of a heterozygous C base deletion mutation (c.1832delC, p.Leu573fs*3) shared by four of those patients. The frame shift deletion is predicted to truncate the BAP1 protein, and immunohistochemistry analysis of tumor specimens revealed predominant cytoplasmic staining. We genotyped 650K SNPs of the four MM samples and four controls and carried out principal component analysis and whole-genome identity-by-descent analysis using publicly available genotype data from the 1000 Genomes Project, UK10K Project, and NHIBL Exome Sequencing Project. These analyses showed that the four MM patients are of Central Europe ancestry and that some are related by a kinship coefficient of 0.0186. Haplotype analysis showed the presence of significantly shared segments around the BAP1 gene (LOD>37.1). The pairwise extent of shared segments between any of the four MM patients ranged in length from 9.1 to 34.2 Mbp and indicates the c.1832delC variant originated in a recent common ancestor within five to ten generations. Through a combined molecular genomic and genealogical approach, we ascertained, to our knowledge, the largest known genealogically connected BAP1 cancer syndrome kindred (K4), whose members can trace descent from a common ancestor in the 17th century. This pedigree provided a unique opportunity to examine effects of BAP1 alteration on tumor expression on various body sites and would facilitate study of gene-gene and gene-environment interaction involving the BAP1 gene. It also suggests that molecular screening of the BAP1 gene, coupled with genealogical research, would be an effective strategy for the early detection and early intervention for BAP1-associated malignancies. Citation Format: Erin Flores, Mitsuru Emi, Todd Johnson, Tatsuhiko Tsunoda, Dusty Behner, Harriet Hoffman, Mary Hesdorffer, Masaki Nasu, Andrea Napolitano, Francine Baumann, Haining Yang, Michele Carbone. A founder mutation in the BAP1 gene among four caucasian families with high incidences of malignant peritoneal mesothelioma and uveal melanoma: a molecular and genealogical study in a 10-generation BAP1 cancer syndrome kindred. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4796. doi:10.1158/1538-7445.AM2015-4796

Abstract 446: BAP1 mutation in mesothelioma and “BAP1 Cancer Syndrome”

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Malignant mesothelioma (MM) is a lethal cancer whose pathogenesis results from complex interactions between host genetics and environmental carcinogens, such as asbestos and erionite fibers. Recently, BAP1 (BRCA associated protein 1) has been identified as a novel MM tumor suppressor gene located at 3p21, a region frequently deleted in MM, which encodes for a deubiquitinase enzyme known to target histones and other proteins. We discovered that germline BAP1 mutations cause a novel cancer syndrome characterized by a significant excess of both pleural and peritoneal MM, uveal and cutaneous melanoma and possibly other tumors. In the same study, we reported that 22% sporadic MM tumors, among the Caucasian population, harbored somatic BAP1 mutations. Several other studies supported a relevant role of BAP1 in MM. However, a significant variation in the frequency of BAP1 mutation was found across different studies and populations. In fact, one research group found BAP1 gene altered in 61% of tumor samples from a Japanese cohort, pointing out to a possible influence of ethnicity on the prevalence of BAP-1 alterations among MM patients. However, the limitations in tumor sample sizes and methodological differences across studies do not allow for conclusive associations. We are now further analyzing BAP1 status and possible clinicopathological associations using different and more sensitive methods, such as MPLA, DNA and RNA sequencing, DNA copy number and methylation. Our preliminary results indicate that, regardless of ethnicity, BAP1 plays a crucial role in MM pathogenesis. More experiments are urgently needed to see whether BAP1 expression could be used in diagnostic, prognostic, or therapeutic settings. The impact of this work will extend to other cancers with BAP1 mutations. Citation Format: Masaki Nasu, Andrea Napolitano, Sandra Pastorino, Mika Tanji, Erin Flores, Francine Baumann, Amy Powers, Giovanni Gaudino, Harvey I. Pass, Haining Yang, Michele Carbone. BAP1 mutation in mesothelioma and “BAP1 Cancer Syndrome”. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 446. doi:10.1158/1538-7445.AM2014-446

Abstract C187: Salicylates suppress tumor growth via inhibition of HMGB1.

High-Mobility Group Box 1 (HMGB1) is a chromatin-associated protein critically involved in nucleosome assembly. During cell necrosis, HMGB1 is first translocated from the nucleus to the cytoplasm, and then passively released to the extracellular space, where it initiates and propagates the inflammatory response. Immune cells (e.g. macrophages) and cancer cells are also able to actively secrete HMGB1. Extracellular HMGB1 is therefore considered a crucial molecule for both chronic inflammation and cancer development, and a promising therapeutic target. In this study, using a malignant mesothelioma (MM) model, we tested the hypothesis that the known anti-cancer effects of aspirin (acetyl salicylic acid), the most widely used anti-inflammatory drug, and salicylic acid, its major metabolite, could be due to HMGB1 inhibition. In vitro, we analyzed the effects of salicylates on HMGB1-induced MM cell proliferation, wound healing, migration, invasion and anchorage-independent colony formation. Moreover, using a MM xenograft model, we tested whether in vivo suppression of HMGB1 signaling by salicylates or BoxA, a specific HMGB1 inhibitor, resulted in reduced MM growth and increased survival. Our results demonstrated that salicylates, at doses normally used in clinical practice (≤1 mM), significantly suppress HMGB1-induced in vitro pro-tumorigenic effects on MM cells and inhibits in vivo MM growth, significantly extending mice survival. Our findings are critically relevant to the understanding of the poorly known anti-cancer effects of salicylates. From this perspective, they apply to more than 50 million people in the USA alone currently taking aspirin. Finally, these preclinical results could be translated to improve the current treatment of the usually lethal MM, and of other HMGB1-related malignancies as well. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C187. Citation Format: Andrea Napolitano, Laura Pellegrini, Sandro Jube, Cormac J. Jennings, Erin G. Flores, David Larson, Vishal S. Negi, Ian Pagano, Mika Tanji, Amy Powers, Sandra Pastorino, Harvey I. Pass, Marco E. Bianchi, Michele Carbone, Haining Yang. Salicylates suppress tumor growth via inhibition of HMGB1. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C187.