Drivers of melanoma susceptibility

Cutaneous melanoma is a cancer of melanocytes, the pigment-producing cells in our skin. It is one of the most aggressive human malignancies, constituting only about 2% of all dermatological cancers but being responsible for over 75% of all deaths from skin cancer. It has recently become a major public health problem, as it is now the fifth most common cancer in the United Kingdom after its incidence more than quadrupled in the last three decades. For these reasons, understanding the biological processes that are involved in its development is of great importance for devising novel treatments and for the management of patients in the clinic. The study of the genetic factors that influence melanoma risk can uncover mechanisms that are relevant in the transition from a benign melanocyte to a malignant melanoma. Approximately 10% of all melanoma cases are familial, and about half of these familial cases can be explained by pathogenetic variants in genes such as cyclin-dependent kinase inhibitor 2A (CDKN2A), cyclin-dependent kinase 4 (CDK4), breast cancer 2 (BRCA2), BRCA1-associated protein-1 (BAP1) and in the promoter of the telomerase reverse transcriptase (TERT). However, about 50% of all familial melanoma cannot be explained by mutations in known genes. In this dissertation, I detail the methodology I followed in an effort to uncover additional high-penetrance melanoma susceptibility genes. I analysed exome and genome sequence data from a total of 184 individuals that belong to 105 melanoma-prone families from the United Kingdom, The Netherlands and Australia that did not have any pathogenetic variants in known susceptibility genes. I applied different gene prioritisation strategies and developed novel software tools in order to devise a list of plausible melanoma susceptibility candidate genes; these analyses suggested that genes regulating telomere function could be influencing melanoma risk. After performing functional experimental analyses, our research team was able to determine that carriers of rare variants in the protection of telomeres (POT1) gene, a member of the shelterin complex that safeguards telomere integrity, are at high risk for developing melanoma. We successfully described the mechanism by which this happens, showing that the variants identified either disrupt POT1 mRNA splicing or abolish the ability of POT1 to bind to telomeres, and lead to increased telomere length in carriers when compared to melanoma cases with wild-type POT1. The main finding of the work described in this dissertation is the identification of telomere dysfunction as an important contributor to the risk of developing melanoma, and possibly other cancers. Our analyses suggest that POT1 is the second most commonly mutated high-penetrance melanoma susceptibility gene reported thus far, and moreover, that rare variants in this gene constitute the first hereditary mechanism for telomere lengthening in humans.This work was supported by a PhD studentship from the Wellcome Trust Sanger Institute and the Consejo Nacional de Ciencia y Tecnologia (CONACYT) of Mexico

An Overview of Strategies for Detecting Genotype-Phenotype Associations Across Ancestrally Diverse Populations

Genome-wide association studies (GWAS) have been very successful at identifying genetic variants influencing a large number of traits. Although the great majority of these studies have been performed in European-descent individuals, it has been recognised that including populations with differing ancestries enhances the potential for identifying causal SNPs due to their differing patterns of linkage disequilibrium. However, when individuals from distinct ethnicities are included in a GWAS, it is necessary to implement a number of control steps to ensure that the identified associations are real genotype-phenotype relationships. In this Review, we discuss the analyses that are required when performing multi-ethnic studies, including methods for determining ancestry at the global and local level for sample exclusion, controlling for ancestry in association testing, and post-GWAS interrogation methods such as genomic control and meta-analysis. We hope that this overview provides a primer for those researchers interested in including distinct populations in their studies

Somatic drivers of B-ALL in a model of ETV6-RUNX1; Pax5(+/-) leukemia.

BACKGROUND: B-cell precursor acute lymphoblastic leukemia (B-ALL) is amongst the leading causes of childhood cancer-related mortality. Its most common chromosomal aberration is the ETV6-RUNX1 fusion gene, with ~25% of ETV6-RUNX1 patients also carrying PAX5 alterations. METHODS: We have recreated this mutation background by inter-crossing Etv6-RUNX1 (Etv6 (RUNX1-SB)) and Pax5(+/-) mice and performed an in vivo analysis to find driver genes using Sleeping Beauty transposon-mediated mutagenesis and also exome sequencing. RESULTS: Combination of Etv6-RUNX1 and Pax5(+/-) alleles generated a transplantable B220 + CD19+ B-ALL with a significant disease incidence. RNA-seq analysis showed a gene expression pattern consistent with arrest at the pre-B stage. Analysis of the transposon common insertion sites identified genes involved in B-cell development (Zfp423) and the JAK/STAT signaling pathway (Jak1, Stat5 and Il2rb), while exome sequencing revealed somatic hotspot mutations in Jak1 and Jak3 at residues analogous to those mutated in human leukemias, and also mutation of Trp53. CONCLUSIONS: Powerful synergies exists in our model suggesting STAT pathway activation and mutation of Trp53 are potent drivers of B-ALL in the context of Etv6-RUNX1;Pax5(+/-)

A Genome-Wide Association Study for Regulators of Micronucleus Formation in Mice.

In mammals the regulation of genomic instability plays a key role in tumor suppression and also controls genome plasticity, which is important for recombination during the processes of immunity and meiosis. Most studies to identify regulators of genomic instability have been performed in cells in culture or in systems that report on gross rearrangements of the genome, yet subtle differences in the level of genomic instability can contribute to whole organism phenotypes such as tumor predisposition. Here we performed a genome-wide association study in a population of 1379 outbred Crl:CFW(SW)-US_P08 mice to dissect the genetic landscape of micronucleus formation, a biomarker of chromosomal breaks, whole chromosome loss, and extranuclear DNA. Variation in micronucleus levels is a complex trait with a genome-wide heritability of 53.1%. We identify seven loci influencing micronucleus formation (false discovery rate <5%), and define candidate genes at each locus. Intriguingly at several loci we find evidence for sexual dimorphism in micronucleus formation, with a locus on chromosome 11 being specific to males.This work was supported by Cancer Research UK and the Wellcome Trust.This is the final version of the article. It first appeared from the Genetics Society of America via http://dx.doi.org/10.1534/g3.116.03076

Germline TERT promoter mutations are rare in familial melanoma.

Germline CDKN2A mutations occur in 40 % of 3-or-more case melanoma families while mutations of CDK4, BAP1, and genes involved in telomere function (ACD, TERF2IP, POT1), have also been implicated in melanomagenesis. Mutation of the promoter of the telomerase reverse transcriptase (TERT) gene (c.-57 T>G variant) has been reported in one family. We tested for the TERT promoter variant in 675 multicase families wild-type for the known high penetrance familial melanoma genes, 1863 UK population-based melanoma cases and 529 controls. Germline lymphocyte telomere length was estimated in carriers. The c.-57 T>G TERT promoter variant was identified in one 7-case family with multiple primaries and early age of onset (earliest, 15 years) but not among population cases or controls. One family member had multiple primary melanomas, basal cell carcinomas and a bladder tumour. The blood leukocyte telomere length of a carrier was similar to wild-type cases. We provide evidence confirming that a rare promoter variant of TERT (c.-57 T>G) is associated with high penetrance, early onset melanoma and potentially other cancers, and explains <1 % of UK melanoma multicase families. The identification of POT1 and TERT germline mutations highlights the importance of telomere integrity in melanoma biology.The authors would like to thank the families for their willingness to participate; and Rajiv Kumar for the provision of mutation positive samples. The collection of samples in the UK population-ascertained sample set was funded by Cancer Research UK (awards C588/A19167 and C8216/A6129) and by the NIH (CA83115). The work of N.A.G. and R.v.D was supported by the Dutch Cancer Society (UL 2012-5489). D.J.A and C.D.R.E are supported by Cancer Research UK, ERC Combat Cancer and the Wellcome Trust. N.K.H is supported by a fellowship from the National Health and Medical Research Council of Australia. A.M.D. and K.A.P. were supported by CRUK grant (C8197/A16565) and The Isaac Newton Trust. K.M.B. is supported by the Intramural Research Program of the Division of Cancer Epidemiology and Genetics; National Cancer Institute; National Institutes of Health.This is the final version of the article. It first appeared from Springer via http://dx.doi.org/10.1007/s10689-015-9841-

Association of germline variants in telomere maintenance genes (POT1, TERF2IP, ACD, and TERT) with spitzoid morphology in familial melanoma: A multi-center case series

Spitzoid morphology in familial melanoma has been associated with germline variants in POT1, a telomere maintenance gene (TMG), suggesting a link between telomere biology and spitzoid differentiation. The aim is to assess if familial melanoma cases associated with germline variants in TMG (POT1, ACD, TERF2IP, and TERT ) commonly exhibit spitzoid morphology.Medicin

POT1 mutations predispose to familial melanoma

Deleterious germline variants in CDKN2A account for around 40% of familial melanoma cases, and rare variants in CDK4, BRCA2, BAP1 and the promoter of TERT have also been linked to the disease. Here we set out to identify new high-penetrance susceptibility genes by sequencing 184 melanoma cases from 105 pedigrees recruited in the UK, The Netherlands and Australia that were negative for variants in known predisposition genes. We identified families where melanoma cosegregates with loss-of-function variants in the protection of telomeres 1 gene (POT1), with a proportion of family members presenting with an early age of onset and multiple primary tumors. We show that these variants either affect POT1 mRNA splicing or alter key residues in the highly conserved oligonucleotide/oligosaccharide-binding (OB) domains of POT1, disrupting protein-telomere binding and leading to increased telomere length. These findings suggest that POT1 variants predispose to melanoma formation via a direct effect on telomeres.D.J.A., C.D.R.-E., Z.D., J.Z.L., J.C.T., M.P. and T.M.K. were supported by Cancer Research UK and the Wellcome Trust (WT098051). C.D.R.-E. was also supported by the Consejo Nacional de Ciencia y Tecnología of Mexico. K.A.P. and A.M.D. were supported by Cancer Research UK (grants C1287/A9540 and C8197/A10123) and by the Isaac Newton Trust. N.K.H. was supported by a fellowship from the National Health and Medical Research Council of Australia (NHMRC). L.G.A. was supported by an Australia and New Zealand Banking Group Limited Trustees PhD scholarship. A.L.P. is supported by Cure Cancer Australia. The work was funded in part by the NHMRC and Cancer Council Queensland. The work of N.A.G. was in part supported by the Dutch Cancer Society (UL 2012-5489). M.H., J.A.N.-B. and D.T.B. were supported by Cancer Research UK (programme awards C588/A4994 and C588/A10589 and the Genomics Initiative). C.L.-O., A.J.R. and V.Q. are funded by the Spanish Ministry of Economy and Competitiveness through the Instituto de Salud Carlos III (ISCIII), the Red Temática de Investigación del Cáncer (RTICC) del ISCIII and the Consolider-Ingenio RNAREG Consortium. C.L.-O. is an investigator with the Botín Foundation.This is the author accepted manuscript. The final version is available from NPG via http://dx.doi.org/10.1038/ng.294

Population-based analysis of POT1 variants in a cutaneous melanoma case–control cohort

Article describes how pathogenic germline variants in the protection of telomeres 1 gene (POT1) have been associated with predisposition to a range of tumor types, including melanoma, glioma, leukemia and cardiac angiosarcoma. The authors sequenced all coding exons of the POT1 gene in 2928 European-descent melanoma cases and 3298 controls, identifying 43 protein-changing genetic variants

Cross-species genomic landscape comparison of human mucosal melanoma with canine oral and equine melanoma.

Mucosal melanoma is a rare and poorly characterized subtype of human melanoma. Here we perform a cross-species analysis by sequencing tumor-germline pairs from 46 primary human muscosal, 65 primary canine oral and 28 primary equine melanoma cases from mucosal sites. Analysis of these data reveals recurrently mutated driver genes shared between species such as NRAS, FAT4, PTPRJ, TP53 and PTEN, and pathogenic germline alleles of BRCA1, BRCA2 and TP53. We identify a UV mutation signature in a small number of samples, including human cases from the lip and nasal mucosa. A cross-species comparative analysis of recurrent copy number alterations identifies several candidate drivers including MDM2, B2M, KNSTRN and BUB1B. Comparison of somatic mutations in recurrences and metastases to those in the primary tumor suggests pervasive intra-tumor heterogeneity. Collectively, these studies suggest a convergence of some genetic changes in mucosal melanomas between species but also distinctly different paths to tumorigenesis

Cabbage and fermented vegetables : From death rate heterogeneity in countries to candidates for mitigation strategies of severe COVID-19

Large differences in COVID-19 death rates exist between countries and between regions of the same country. Some very low death rate countries such as Eastern Asia, Central Europe, or the Balkans have a common feature of eating large quantities of fermented foods. Although biases exist when examining ecological studies, fermented vegetables or cabbage have been associated with low death rates in European countries. SARS-CoV-2 binds to its receptor, the angiotensin-converting enzyme 2 (ACE2). As a result of SARS-CoV-2 binding, ACE2 downregulation enhances the angiotensin II receptor type 1 (AT(1)R) axis associated with oxidative stress. This leads to insulin resistance as well as lung and endothelial damage, two severe outcomes of COVID-19. The nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is the most potent antioxidant in humans and can block in particular the AT(1)R axis. Cabbage contains precursors of sulforaphane, the most active natural activator of Nrf2. Fermented vegetables contain many lactobacilli, which are also potent Nrf2 activators. Three examples are: kimchi in Korea, westernized foods, and the slum paradox. It is proposed that fermented cabbage is a proof-of-concept of dietary manipulations that may enhance Nrf2-associated antioxidant effects, helpful in mitigating COVID-19 severity.Peer reviewe