Germline Variants in Patients Affected by Both Uveal and Cutaneous Melanoma.

Uveal melanoma (UM) and nonacral cutaneous melanoma (CM) are distinct entities with varied genetic landscapes despite both arising from melanocytes. There are, however, similarities in that they most frequently affect people of European ancestry, and high penetrance germline variants in BAP1, POT1 and CDKN2A have been shown to predispose to both UM and CM. This study aims to further explore germline variants in patients affected by both UM and CM, shedding light on the underlying genetic mechanism causing these diseases. Using exome sequencing we analysed germline DNA samples from a cohort of 83 Australian patients diagnosed with both UM and CM. Eight (10%) patients were identified that carried pathogenic mutations in known melanoma predisposition genes POT1, MITF, OCA2, SLC45A2 and TYR. Three (4%) patients carried pathogenic variants in genes previously linked with other cancer syndromes (ATR, BRIP1 and MSH6) and another three cases carried monoallelic pathogenic variants in recessive cancer genes (xeroderma pigmentosum and Fanconi anaemia), indicating that reduced penetrance of phenotype in these individuals may contribute to the development of both UM and CM. These findings highlight the need for further studies characterising the role of these genes in melanoma susceptibility.

Iris melanoma in an Australian cohort.

BACKGROUND: To report the clinicopathological features and epidemiology of iris melanoma in Queensland, Australia. METHODS: This was a retrospective study of 86 patients with iris melanoma treated between 2001 and 2022 at the Queensland Ocular Oncology Service, Brisbane, Australia. Main outcome measures included demographics, clinical and phenotypic features, age-adjusted incidence and relative survival. RESULTS: Eighty-six patients (63% female) were included. Mean age was 54 years (range 17-82 years). The majority of patients (97%) were Caucasian, with blue eyes, fair skin and Fitzpatrick Skin Type I or II. Demographic features and clinical history showed a tendency for high ultraviolet radiation (UVR) exposure in the cohort. Histopathology was available in 69 cases (82%), and of these, 77% tumours were of spindle cell origin, with low-risk genetic profiles. Patients were followed for a mean of 8 years (median 7, range 1-21 years) after diagnosis, and only one case of metastasis was documented. CONCLUSIONS: The association of iris freckles, history of UVR exposure and dermatologic findings supports the role of UVR in iris melanoma. Occupation and avocation history, as well as evaluation of iris freckles may offer an easily accessible way of stratifying the risk of an individual for development of UVR-related uveal melanoma.

Combined photodynamic therapy and transpupillary thermotherapy for small choroidal melanoma.

OBJECTIVE: The objective of this study was to determine whether combining verteporfin-based photodynamic therapy (PDT) and transpupillary thermotherapy (TTT) achieves adequate tumour control while maintaining visual acuity in individuals with small choroidal melanoma of amelanotic, melanotic, and variable pigmentation. DESIGN: Individuals with posterior choroidal melanomas up to 3 mm in height underwent verteporfin-based PDT followed by immediate TTT. Further combined laser therapy was performed if a poor response was noted at 12 weeks or beyond. Tumours that demonstrated significant further growth were treated with brachytherapy or enucleation. A total of 37 eyes of 37 patients from the Terrace Eye Centre in Brisbane, Australia were studied. Average age of participants was 59.62 ± 12.45 years, and 17 of 37 participants were female (46%). METHODS: This was a retrospective, noncomparative interventional study. RESULTS: Seven of the 37 participants (19%) had recurrence of their tumour requiring further brachytherapy or enucleation. There was no statistically significant difference in visual acuity before and after treatment. There were no baseline characteristics that predicted treatment outcome. Ten individuals developed complications including epiretinal membrane (16%), scotoma (8%), cataract (3%), and macular edema (3%). No individuals experienced extraocular extension or progressed to metastatic disease. The mean follow-up time was 49 months. CONCLUSION: Combined PDT and TTT achieved 81% tumour control in this study while preserving visual acuity. However, higher rates of local recurrence compared with brachytherapy warrant close follow-up to identify recurrences early.

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

Background: 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. Objective: To assess if familial melanoma cases associated with germline variants in TMG (POT1, ACD, TERF2IP, and TERT) commonly exhibit spitzoid morphology. Methods: In this case series, melanomas were classified as having spitzoid morphology if at least 3 of 4 dermatopathologists reported this finding in ≥25% of tumor cells. Logistic regression was used to calculate odds ratios (OR) of spitzoid morphology compared to familial melanomas from unmatched noncarriers that were previously reviewed by a National Cancer Institute dermatopathologist. Results: Spitzoid morphology was observed in 77% (23 of 30), 75% (3 of 4), 50% (2 of 4), and 50% (1 of 2) of melanomas from individuals with germline variants in POT1, TERF2IP, ACD, and TERT, respectively. Compared to noncarriers (n = 139 melanomas), POT1 carriers (OR = 225.1, 95% confidence interval: 51.7-980.5; P < .001) and individuals with TERF2IP, ACD, and TERT variants (OR = 82.4, 95% confidence interval: 21.3-494.6; P < .001) had increased odds of spitzoid morphology. Limitations: Findings may not be generalizable to nonfamilial melanoma cases. Conclusion: Spitzoid morphology in familial melanoma could suggest germline alteration of TMG.

Comparative Genomics Provides Etiologic and Biological Insight into Melanoma Subtypes.

Melanoma is a cancer of melanocytes, with multiple subtypes based on body site location. Cutaneous melanoma is associated with skin exposed to ultraviolet radiation; uveal melanoma occurs in the eyes; mucosal melanoma occurs in internal mucous membranes; and acral melanoma occurs on the palms, soles, and nail beds. Here, we present the largest whole-genome sequencing study of melanoma to date, with 570 tumors profiled, as well as methylation and RNA sequencing for subsets of tumors. Uveal melanoma is genomically distinct from other melanoma subtypes, harboring the lowest tumor mutation burden and with significantly mutated genes in the G-protein signaling pathway. Most cutaneous, acral, and mucosal melanomas share alterations in components of the MAPK, PI3K, p53, p16, and telomere pathways. However, the mechanism by which these pathways are activated or inactivated varies between melanoma subtypes. Additionally, we identify potential novel germline predisposition genes for some of the less common melanoma subtypes. SIGNIFICANCE: This is the largest whole-genome analysis of melanoma to date, comprehensively comparing the genomics of the four major melanoma subtypes. This study highlights both similarities and differences between the subtypes, providing insights into the etiology and biology of melanoma. This article is highlighted in the In This Issue feature, p. 2711.

Choroidal melanoma with synchronous Fuchs' adenoma and novel ATRX mutation.

BACKGROUND: To report a case of Fuchs' adenoma occurring in an eye with a large choroidal melanoma. We have reviewed the literature to describe the clinical presentation, ultrasound characteristics and pathological features of these entities. CASE PRESENTATION: A 69-year-old Caucasian man presented with vision loss from a large choroidal melanoma. Enucleation showed an incidental Fuchs' adenoma in the same eye. Whole-exome sequence analysis was also performed on the patient's blood and melanoma, which showed a rarely-reported ATRX mutation. CONCLUSIONS: Fuchs' adenoma is an under-diagnosed benign age-related hyperplasia of the non-pigmented ciliary epithelium (NPCE). Given its location and characteristics, it can be mistaken for choroidal melanoma and clinicians are reminded how to differentiate between these pathologies and that they may co-exist.

FRAMe: Familial Risk Assessment of Melanoma-a risk prediction tool to guide CDKN2A germline mutation testing in Australian familial melanoma.

Germline mutations in CDKN2A greatly increase risk of developing cutaneous melanoma. We have constructed a risk prediction model, Familial Risk Assessment of Melanoma (FRAMe), for estimating the likelihood of carrying a heritable CDKN2A mutation among Australian families, where the prevalence of these mutations is low. Using logistic regression, we analysed characteristics of 299 Australian families recruited through the Sydney site of GenoMEL (international melanoma genetics consortium) with at least three cases of cutaneous melanoma (in situ and invasive) among first-degree blood relatives, for predictors of the presence of a pathogenic CDKN2A mutation. The final multivariable prediction model was externally validated in an independent cohort of 61 melanoma kindreds recruited through GenoMEL Queensland. Family variables independently associated with the presence of a CDKN2A mutation in a multivariable model were number of individuals diagnosed with melanoma under 40 years of age, number of individuals diagnosed with more than one primary melanoma, and number of individuals blood related to a melanoma case in the first degree diagnosed with any cancer excluding melanoma and non-melanoma skin cancer. The number of individuals diagnosed with pancreatic cancer was not independently associated with mutation status. The risk prediction model had an area under the receiver operating characteristic curve (AUC) of 0.851 (95% CI 0.793, 0.909) in the training dataset, and 0.745 (95%CI 0.612, 0.877) in the validation dataset. This model is the first to be developed and validated using only Australian data, which is important given the higher rate of melanoma in the population. This model will help to effectively identify families suitable for genetic counselling and testing in areas of high ambient ultraviolet radiation. A user-friendly electronic nomogram is available at www.melanomarisk.org.au .

Genomic analysis of adult case of ocular surface giant congenital melanocytic nevus and associated clinicopathological findings.

INTRODUCTION: Conjunctival nevi are the most common tumor of the ocular surface. There are some rare reports of so-called 'giant' conjunctival nevi. We report a case of a 47-year-old female with a cutaneous and ocular surface giant congenital melanocytic nevus and describe her clinical course. CASE DESCRIPTION: This is a retrospective case report of a single patient. A 47-year-old female with a history of biopsy-proven periorbital congenital melanocytic nevus, with an associated giant conjunctival nevus presented for structural and functional rehabilitation. Serial surgeries were performed and excised tissue was sent for histopathological and genetic examination. The conjunctival nevus had a low tumor mutation burden, and of the 647 somatic mutations, only one occurred within a protein coding region, namely NRAS p.Gln61Arg. CONCLUSION: This is the first reported adult case including genomic analysis of an ocular surface giant congenital melanocytic nevus. The case shows a possible association between periorbital congenital melanocytic nevi and giant conjunctival nevi, and underscores the possible role that targeted drug therapies may have in malignant transformation of these conditions.

Multiplex melanoma families are enriched for polygenic risk.

Cancers, including cutaneous melanoma, can cluster in families. In addition to environmental etiological factors such as ultraviolet radiation, cutaneous melanoma has a strong genetic component. Genetic risks for cutaneous melanoma range from rare, high-penetrance mutations to common, low-penetrance variants. Known high-penetrance mutations account for only about half of all densely affected cutaneous melanoma families, and the causes of familial clustering in the remainder are unknown. We hypothesize that some clustering is due to the cumulative effect of a large number of variants of individually small effect. Common, low-penetrance genetic risk variants can be combined into polygenic risk scores. We used a polygenic risk score for cutaneous melanoma to compare families without known high-penetrance mutations with unrelated melanoma cases and melanoma-free controls. Family members had significantly higher mean polygenic load for cutaneous melanoma than unrelated cases or melanoma-free healthy controls (Bonferroni-corrected t-test P = 1.5 × 10-5 and 6.3 × 10-45, respectively). Whole genome sequencing of germline DNA from 51 members of 21 families with low polygenic risk for melanoma identified a CDKN2A p.G101W mutation in a single family but no other candidate high-penetrance melanoma susceptibility genes. This work provides further evidence that melanoma, like many other common complex disorders, can arise from the joint action of multiple predisposing factors, including rare high-penetrance mutations, as well as via a combination of large numbers of alleles of small effect.

Whole genome landscapes of uveal melanoma show an ultraviolet radiation signature in iris tumours.

Uveal melanoma (UM) is the most common intraocular tumour in adults and despite surgical or radiation treatment of primary tumours, ~50% of patients progress to metastatic disease. Therapeutic options for metastatic UM are limited, with clinical trials having little impact. Here we perform whole-genome sequencing (WGS) of 103 UM from all sites of the uveal tract (choroid, ciliary body, iris). While most UM have low tumour mutation burden (TMB), two subsets with high TMB are seen; one driven by germline MBD4 mutation, and another by ultraviolet radiation (UVR) exposure, which is restricted to iris UM. All but one tumour have a known UM driver gene mutation (GNAQ, GNA11, BAP1, PLCB4, CYSLTR2, SF3B1, EIF1AX). We identify three other significantly mutated genes (TP53, RPL5 and CENPE).

A Panel of Circulating MicroRNAs Detects Uveal Melanoma With High Precision.

PURPOSE: To determine if a circulating microRNA (miRNA) panel could be used to distinguish between uveal melanoma and uveal nevi. METHODS: We report on a multicenter, cross-sectional study conducted between June 2012 and September 2015. The follow-up time was approximately 3 to 5 years. Blood was drawn from participants presenting with a uveal nevus (n = 10), localized uveal melanoma (n = 50), or metastatic uveal melanoma (n = 5). Levels of 17 miRNAs were measured in blood samples of study participants using a sensitive real-time PCR system. RESULTS: A panel of six miRNAs (miR-16, miR-145, miR-146a, miR-204, miR-211, and miR-363-3p) showed significant differences between participants with uveal nevi compared with patients with localized and metastatic uveal melanoma. Importantly, miR-211 was able to accurately distinguish metastatic disease from localized uveal melanoma (P < 0.0001; area under the curve = 0.96). When the six-miRNA panel was evaluated as a group it had the ability to identify uveal melanoma when four or more miRNAs (93% sensitivity and 100% specificity) reached or exceeded their cut-point. CONCLUSIONS: This miRNA panel, in tandem with clinical findings, may be suited to confirm benign lesions. In addition, due to the panel's high precision in identifying malignancy, it has the potential to augment melanoma detection in subsequent clinical follow-up of lesions with atypical clinical features. TRANSLATIONAL RELEVANCE: Uveal nevi mimic the appearance of uveal melanoma and their transformation potential cannot be definitively determined without a biopsy. This panel is most relevant at the nevus stage and in lesions with uncertain malignant potential as a companion diagnostic tool to assist in clinical decision-making.

Evaluation of the contribution of germline variants in BRCA1 and BRCA2 to uveal and cutaneous melanoma.

Germline mutations of BRCA1 and BRCA2 predispose individuals to a high risk of breast and ovarian cancer, and elevated risk of other cancers, including those of the pancreas and prostate. BRCA2 mutation carriers may have increased risk of uveal melanoma (UM) and cutaneous melanoma (CM), but associations with these cancers in BRCA1 mutation carriers have been mixed. Here, we further assessed whether UM and CM are associated with BRCA1 or BRCA2 by assessing the presence, segregation and reported/predicted pathogenicity of rare germline mutations (variant allele frequency < 0.01) in families with multiple members affected by these cancers. Whole-genome or exome sequencing was performed on 160 CM and/or UM families from Australia, the Netherlands, Denmark and Sweden. Between one and five cases were sequenced from each family, totalling 307 individuals. Sanger sequencing was performed to validate BRCA1 and BRCA2 germline variants and to assess carrier status in other available family members. A nonsense and a frameshift mutation were identified in BRCA1, both resulting in premature truncation of the protein (the first at p.Q516 and the second at codon 91, after the introduction of seven amino acids due to a frameshift deletion). These variants co-segregated with CM in individuals who consented for testing and were present in individuals with pancreatic, prostate and breast cancer in the respective families. In addition, 33 rare missense mutations (variant allele frequency ranging from 0.00782 to 0.000001 in the aggregated ExAC data) were identified in 34 families. Examining the previously reported evidence of functional consequence of these variants revealed all had been classified as either benign or of unknown consequence. Seeking further evidence of an association between BRCA1 variants and melanoma, we examined two whole-genome/exome sequenced collections of sporadic CM patients (total N = 763). We identified one individual with a deleterious BRCA1 variant, however, this allele was lost (with the wild-type allele remaining) in the corresponding CM, indicating that defective BRCA1 was not a driver of tumorigenesis in this instance. Although this is the first time that deleterious BRCA1 mutations have been described in high-density CM families, we conclude that there is an insufficient burden of evidence to state that the increased familial CM or UM susceptibility is because of these variants. In addition, in conjunction with other studies, we conclude that the previously described association between BRCA2 mutations and UM susceptibility represents a rare source of increased risk.

Correction to: Prolonged stable disease in a uveal melanoma patient with germline MBD4 nonsense mutation treated with pembrolizumab and ipilimumab.

The authors regret that the online version of this article contains an error. The MBD4 mutation in sample MM138 was given an incorrect dbSNP ID. The correct ID is rs769076971.

Germline variants in oculocutaneous albinism genes and predisposition to familial cutaneous melanoma.

Approximately 1%-2% of cutaneous melanoma (CM) is classified as strongly familial. We sought to investigate unexplained CM predisposition in families negative for the known susceptibility genes using next-generation sequencing of affected individuals. Segregation of germline variants of interest within families was assessed by Sanger sequencing. Several heterozygous variants in oculocutaneous albinism (OCA) genes: TYR, OCA2, TYRP1 and SLC45A2, were present in our CM cohort. OCA is a group of autosomal recessive genetic disorders, resulting in pigmentation defects of the eyes, hair and skin. Missense variants classified as pathogenic for OCA were present in multiple families and some fully segregated with CM. The functionally compromised TYR p.T373K variant was present in three unrelated families. In OCA2, known pathogenic variants: p.V443I and p.N489D, were present in three families and one family, respectively. We identified a likely pathogenic SLC45A2 frameshift variant that fully segregated with CM in a family of four cases. Another four-case family harboured cosegregating variants (p.A24T and p.R153C) of uncertain functional significance in TYRP1. We conclude that rare, heterozygous variants in OCA genes confer moderate risk for CM.

Estimating CDKN2A mutation carrier probability among global familial melanoma cases using GenoMELPREDICT.

BACKGROUND: Although rare in the general population, highly penetrant germline mutations in CDKN2A are responsible for 5%-40% of melanoma cases reported in melanoma-prone families. We sought to determine whether MELPREDICT was generalizable to a global series of families with melanoma and whether performance improvements can be achieved. METHODS: In total, 2116 familial melanoma cases were ascertained by the international GenoMEL Consortium. We recapitulated the MELPREDICT model within our data (GenoMELPREDICT) to assess performance improvements by adding phenotypic risk factors and history of pancreatic cancer. We report areas under the curve (AUC) with 95% confidence intervals (CIs) along with net reclassification indices (NRIs) as performance metrics. RESULTS: MELPREDICT performed well (AUC 0.752, 95% CI 0.730-0.775), and GenoMELPREDICT performance was similar (AUC 0.748, 95% CI 0.726-0.771). Adding a reported history of pancreatic cancer yielded discriminatory improvement (P < .0001) in GenoMELPREDICT (AUC 0.772, 95% CI 0.750-0.793, NRI 0.40). Including phenotypic risk factors did not improve performance. CONCLUSION: The MELPREDICT model functioned well in a global data set of familial melanoma cases. Adding pancreatic cancer history improved model prediction. GenoMELPREDICT is a simple tool for predicting CDKN2A mutational status among melanoma patients from melanoma-prone families and can aid in directing these patients to receive genetic testing or cancer risk counseling.

Prolonged stable disease in a uveal melanoma patient with germline MBD4 nonsense mutation treated with pembrolizumab and ipilimumab.

There is currently no effective treatment for metastasised uveal melanoma (UM). Recently, it was reported that a UM patient was responsive to checkpoint inhibitor (CI) treatment, due to a high tumour mutation burden correlated with a germline loss-of-function MBD4 mutation. Here, we report on another UM patient who carried an MBD4 germline nonsense variant (p.Leu563Ter) and her tumour showed a fivefold higher than average mutation burden. We confirmed the association between germline loss-of-function variant in MBD4 and CI response. The patient experienced stable disease (10 months) and survived 2 years with metastatic disease, which is twice as long as median survival. Additionally, the frequency of MBD4 loss-of-function variants in reported UM cohorts was > 20 times higher than in an aggregated population genome database (P < 5 × 10-5), implying a potential role as UM predisposition gene. These findings provide a strong basis for the inclusion of MBD4 in the screening of potential UM-prone families as well as stratification of immunotherapy.

Germline mutations in candidate predisposition genes in individuals with cutaneous melanoma and at least two independent additional primary cancers.

BACKGROUND: While a number of autosomal dominant and autosomal recessive cancer syndromes have an associated spectrum of cancers, the prevalence and variety of cancer predisposition mutations in patients with multiple primary cancers have not been extensively investigated. An understanding of the variants predisposing to more than one cancer type could improve patient care, including screening and genetic counselling, as well as advancing the understanding of tumour development. METHODS: A cohort of 57 patients ascertained due to their cutaneous melanoma (CM) diagnosis and with a history of two or more additional non-cutaneous independent primary cancer types were recruited for this study. Patient blood samples were assessed by whole exome or whole genome sequencing. We focussed on variants in 525 pre-selected genes, including 65 autosomal dominant and 31 autosomal recessive cancer predisposition genes, 116 genes involved in the DNA repair pathway, and 313 commonly somatically mutated in cancer. The same genes were analysed in exome sequence data from 1358 control individuals collected as part of non-cancer studies (UK10K). The identified variants were classified for pathogenicity using online databases, literature and in silico prediction tools. RESULTS: No known pathogenic autosomal dominant or previously described compound heterozygous mutations in autosomal recessive genes were observed in the multiple cancer cohort. Variants typically found somatically in haematological malignancies (in JAK1, JAK2, SF3B1, SRSF2, TET2 and TYK2) were present in lymphocyte DNA of patients with multiple primary cancers, all of whom had a history of haematological malignancy and cutaneous melanoma, as well as colorectal cancer and/or prostate cancer. Other potentially pathogenic variants were discovered in BUB1B, POLE2, ROS1 and DNMT3A. Compared to controls, multiple cancer cases had significantly more likely damaging mutations (nonsense, frameshift ins/del) in tumour suppressor and tyrosine kinase genes and higher overall burden of mutations in all cancer genes. CONCLUSIONS: We identified several pathogenic variants that likely predispose to at least one of the tumours in patients with multiple cancers. We additionally present evidence that there may be a higher burden of variants of unknown significance in 'cancer genes' in patients with multiple cancer types. Further screens of this nature need to be carried out to build evidence to show if the cancers observed in these patients form part of a cancer spectrum associated with single germline variants in these genes, whether multiple layers of susceptibility exist (oligogenic or polygenic), or if the occurrence of multiple different cancers is due to random chance.