Posted: November 1, 2016
How one investigator’s passion and devotion to science led to discovery of the “ghost gene” in MPM
Q&A with Michele Carbone, MD
Q: Many laboratories are now working on the link between BAP1 and mesothelioma. How did you come up with the idea that there was genetic susceptibility to developing mesothelioma?
A: Only a fraction of individuals exposed to asbestos develop mesothelioma, so we believed that if we could identify the reasons some people are more susceptible than others, we could develop preventive strategies. To this end, we studied an epidemic of mesothelioma in Cappadocia, Turkey, where about 50% of the population exposed to erionite fibers dies of mesothelioma. Over the course of 14 years, we discovered that susceptibility to mesothelioma was transmitted in a Mendelian fashion; we formulated the hypothesis that the cause of the epidemic environmental interaction with gene X.
Q: Who funded these studies?
A: Initially, I paid out of my own pocket, and I spent whatever vacation time I had working in Cappadocia. Then, after compiling preliminary data, I was awarded a grant from the American Cancer Society. Those studies allowed me to be awarded an NCI-P01 grant in 2006 to identify the hypothetical mesothelioma susceptibility gene(s), which we later identified as BAP1.
Q: How did you find the gene?
A: Initially, in the pre-NGS era, the gene eluded us. Dr. Nasu and other postdocs in my lab manually sequenced miles and miles of DNA looking for the putative mesothelioma-gene. I traveled around the country to be in the operating room of patients with a family history of mesothelioma to diagnose their tumors and to harvest tumor cells, from which I extracted the DNAs that Dr. Nasu and others sequenced. Four years into the grant, we had ruled out several DNA regions that genetic linkage analyses and comparative genomic hybridization— done by our P01 collaborators Dr. Cox at the University of Chicago and Dr. Testa at Fox Chase Cancer Center—had identified as possibly harboring a mesothelioma gene. We had significantly narrowed down potential chromosomal regions indicated by linkage analyses, but we were in the last year of the P01 with not enough time to sequence them all. We knew that we had little hope of renewing the grant and continuing this work, since we had nothing to show for it. There was also increasing skepticism that a mesothelioma gene existed. At a Chicago ASCO meeting, I was introduced with these words: “Now Dr. Carbone will tell us about his hunt for the ghost gene.” I started thinking that maybe I had committed a terrible error and that there was no such thing as a mesothelioma gene. The epiphany came when I received a phone call from Louisiana: a member of one of the families I was studying had developed an eye melanoma and a mesothelioma. I asked my colleague at University of Hawaii Cancer Center, biostatistician Dr. Pagano, “What is the chance of these two very rare tumors occurring in the same person, and of multiple mesotheliomas and eye melanomas in the same family?” He answered, “About 2 of 10 million cases.” I then concluded that this could not be a coincidence. I pulled out Principles and Practice of Oncology from my bookshelf, and found that eye melanomas and mesotheliomas shared frequent deletions in chromosome 3p, a region that was on Dr. Nasu’s to-do list, based on genetic linkage studies. A few days later, my collaborators Giovanni Gaudino and Haining Yang at the University of Hawaii Cancer Center walked into my office showing me a new paper by Dr. Harbour reporting a high incidence of somatic BAP1 mutations in eye melanoma, and Dr. Testa sent me an email about the same paper. BAP1 was on 3p. Was it BAP1? We were all quite excited. We discussed the possibility in a videoconference meeting with my P0-1 collaborator Dr. Pass at New York University. We were running out of grant time and this was our last chance, so we decided to focus sequencing on 3p. Dr. Testa’s lab helped speed up the process. The first clue came a few weeks later from Dr. Testa’s lab when he and his colleagues identified a truncating BAP1 mutation; soon after, Dr. Nasu in my lab obtained his sequence results and discovered the same mutation. There was still a possibility that this was a fluke, but that possibility was ruled out in the following weeks when DNA sequencing perfectly matched all affected members of two separate US families with BAP1 mutations, while none of the non-affected family members had inherited these mutations. Same results, two separate laboratories. Moreover, among 23 sporadic mesothelioma blood samples that Harvey Pass and Mary Hesdorffer at Mesothelioma Applied Research Foundation (MARF) sent us, we found germline mutations in two: the only two patients who previously had developed uveal melanoma. After more than 14 years of research, I could hardly believe that it was true. With Drs. Gaudino and Yang we reviewed all the data to rule out any possible mistake: they were very solid. We published that BAP1 was the mutated gene that caused mesothelioma and eye melanoma in some families. In the same issue of Nature Genetics, Speicher’s team reported that inherited BAP1 mutations caused benign melanocytic tumors. We checked our patients: they had those melanocytic lesions. Speicher and colleagues checked the family history of their patients and found several mesotheliomas. This was all quite reassuring!
Q: Why mesothelioma and uveal melanoma?
A: Using a BAP1+/- mouse model, we demonstrated that these mice develop mesothelioma following exposure to very low doses of asbestos. So we think that germline BAP1 mutation carriers may be more susceptible to even small amounts of asbestos exposure. Similarly, BAP1 mutations may increase sensitivity to ultraviolet (UV)-induced mutagenesis. Therefore, BAP1 is the first gene shown to modulate mineral fiber and environmental carcinogenesis.
Q: Do BAP1 mutation carriers develop other malignancies?
A: Yes. Subsequent to our research, other scientists have linked germline BAP1 mutations to renal cell and gallbladder carcinomas, basal cell carcinomas, various types of sarcomas, etc. I think BAP1 mutant carriers may be prone to many types of cancer, especially to cancers caused by environmental carcinogens. We named this condition “the BAP1 cancer syndrome.”
Q: What is the incidence of cancer in carriers of this BAP1 cancer syndrome?
A: So far, all carriers we studied have developed one or more cancers during their lifetime. However, we discovered that mesothelioma and other cancers that develop in a background of BAP1+/- germline mutations are often associated with prolonged survival of five to 10 or more years, which we think this may be related in part to early detection, since these families are closely monitored, and in part to a possibly less aggressive tumor phenotype.
Q: How many BAP1 families are there? Are there other genes that cause mesothelioma?
A: We have studied over 50 families in which several family members developed mesothelioma and often also other malignancies. Some of them do not have BAP1 mutations. Now, with support from a US Department of Defense grant, we are trying to identify a second mesothelioma gene. In a paper in press in PNAS, we identified novel genes that contribute to sporadic mesothelioma; we are now testing whether they are also mutated in the germline.
Q: How do BAP1 mutations originate?
A: The mutations are inherited and can be traced through centuries of ancestry. We identified four families who share the same BAP1 point mutation. We discovered that the four families are related and descend from a German couple who immigrated to the US in the early 1700s. From this extended family, we have built a family tree encompassing ~80,000 people, and we are identifying additional branches of this family who carry the mutation.
Q: How can medicine help carriers of BAP1 mutations?
A: Genetic counseling is imperative to inform carriers about prevention. Such individuals should avoid UV sunlight and trades in which asbestos exposure may occur. Patients should understand that their tumors are usually less aggressive than the same tumors when they occur sporadically. Moreover, we are enrolling BAP1+/- carriers in screening programs for early detection of eye and skin melanoma, which are cancers that are curable when detected early, and for biomarkers studies. We will soon start a prospective HMGB1 study following a recent discovery by Dr. Yang that this biomarker may allow us to identify asbestos-exposed individuals and those among them who are more likely to develop mesothelioma.
Q: Who helped you the most in this research?
A: The members of the high-risk mesothelioma families deserve most of the credit. Among them, the Cappadocia families in Turkey, and the Wisconsin and Louisiana families. Without their help, we would have never discovered that mesothelioma had a genetic component first, and later identified the gene.
Q: In your scientific team, who deserves most credit for this discovery?
A: Our success was the result of teamwork, and we would not have succeeded without the participation of all the scientists listed in our papers. I will also mention that some team members spent significant time traveling and working with me in Cappadocia, exposing themselves to the potent carcinogenic erionite fibers following my “crazy” idea that there was a mesothelioma gene to be found and that we could reduce the incidence of mesothelioma. They did it because of their passion and because they wanted to make a difference. To them, I am most grateful. Moreover, Dr. Murat Tuncer, then Director of Cancer Control at the Turkish Ministry of Health in Turkey, upon our request built a clinic in the village of Tuzkoy to help mesothelioma patients and our research.
Q: What lessons have you learned in Cappadocia?
A: Many. Probably the most important was to listen to my heart rather than logic and to never give up. Once, as I was about to leave for Turkey a senior colleague, said, “Michele, if I were you, rather than wasting time looking for a ghost gene in Cappadocia, I would go to the Caribbean for the next few months.” Everybody around us laughed. But I had met the mesothelioma families in Cappadocia and in the US, I wanted to do something about their problem, and I hoped (although I was not sure) that my hypothesis of a meso-gene was correct. I was not after a grant: I wanted to fix a problem. Similarly, a few years later, with my Turkish collaborators Drs. Baris, Dogan and Emri we went to the Turkish Ministry of Health and asked them to build two new villages in Cappadocia and to relocate the residents of the erionite- containing villages to prevent further exposure in a population that was predisposed to mesothelioma. We never thought they would have listened to us, yet we wanted to try: we waited one week in Ankara to talk to the Ministry. Well, the Ministry listened to us, and they built two new erionite-free villages, likely saving many future lives from mesothelioma! ✦
Bibliography
Baumann F, Buck BJ, Metcalf RV, et al. The presence of asbestos in the natural environment is likely related to mesothelioma in young individuals and women from Southern Nevada. J Thorac Oncol. 2015; 10:731-737. Baumann F, Flores E, Napolitano A, et al. Mesothelioma Patients with Germline BAP1 Mutations Have Seven- Fold Improved Long-term Survival. Carcinogenesis. 2015; 36:76-81.
Carbone M, et al. Consensus Report of the 2015 Weinman International Conference on Mesothelioma. J Thorac Oncol. In press.
Carbone M, Baris YI, Bertino P, et al. Erionite exposure in North Dakota and Turkish villages with mesothelioma. Proc Natl Acad Sci USA. 2011; 108:13618-13623. Carbone M, Emri S, Dogan U, et al. A mesothelioma epidemic in Cappadocia: scientific developments and unexpected social outcomes. Nature Reviews Cancer. 2007; 7:147-154.
Carbone M, Ferris LK, Baumann F, et al. BAP1 cancer syndrome: malignant mesothelioma, uveal and cutaneous melanoma, and MBAITs. J Transl Med. 2012;10:179.
Carbone M, Flores EG, Emi M, et al. Combined genetic and genealogic studies uncover a large BAP1 cancer syndrome kindred, tracing back nine generations to a common ancestor from the 1700s. PLoS Genetics. 2015; 11(12):e1005633.
Carbone M, Yang H, Pass HI, et al. Bap1 and cancer. Nat Rev Cancer. 2013; 13:153-159. Dogan UA, Baris YI, Dogan M, et al. Genetic Predisposition to Fiber Carcinogenesis Causes a mesothelioma Epidemic in Turkey. Cancer Res. 2006; 66:5063-5068. Napolitano A, Antoine DJ , Pellegrini L, et al. HMGB1 and its hyper-acetylated isoform are sensitive and specific serum biomarkers to detect asbestos exposure and to identify mesothelioma patients. Clin Cancer Res. 2016; 22:3087-3096.
Napolitano A, Pellegrini L, Dei A, et al. Minimal asbestos exposure in germline BAP1 heterozygous mice is associated with deregulated inflammatory response and increased risk of mesothelioma. Oncogene. 2016; 35:1996-2002.
Nasu M, Emi M, Pastorino S, et al. High incidence of somatic BAP1 alterations in sporadic malignant mesothelioma. J Thorac Oncol. 2015;10:565-576.
Roushdy-Hammady I, Siegel J, Emri S, et al. A geneticsusceptibility factor malignant mesothelioma in the Cappadocian region of Turkey. The Lancet. 2001; 357:444-445. Testa JR, Cheung M, Pei J, et al. Germline BAP1 mutations predispose to malignant mesothelioma. Nat Genet. 2011; 43:1022-1025.
Yang H, Rivera Z, Jube S, et al. Programmed necrosis induced by asbestos in human mesothelial cells causes high-mobility group box 1 protein release and resultant inflammation. Proc Natl Acad Sci USA. 2010; 107:12611-12616.
Yoshikawa Y, Emi M, Hashimoto-Tamaoki T, et al. High-density array-CGH with targeted NGS unmask multiple non-contiguous minute deletions on chromosome 3p21 in mesothelioma. Proc Natl Aca Sci USA. 2016; in press.
