Whole Exome Sequencing Identifies Candidate Genes Associated with Hereditary Predisposition to Uveal Melanoma.

PURPOSE: To identify susceptibility genes associated with hereditary predisposition to uveal melanoma (UM) in patients with no detectable germline BAP1 alterations. DESIGN: Retrospective case series from academic referral centers. PARTICIPANTS: Cohort of 154 UM patients with high risk of hereditary cancer defined as patients with 1 or more of the following: (1) familial UM, (2) young age (<35 years) at diagnosis, (3) personal history of other primary cancers, and (4) family history of 2 or more primary cancers with no detectable mutation or deletion in BAP1 gene. METHODS: Whole exome sequencing, a cancer gene panel, or both were carried out. Probands included 27 patients with familial UM, 1 patient with bilateral UM, 1 patient with congenital UM, and 125 UM patients with strong personal or family histories, or both, of cancer. Functional validation of variants was carried out by immunohistochemistry, reverse-transcriptase polymerase chain reaction, and genotyping. MAIN OUTCOME MEASURES: Clinical characterization of UM patients with germline alterations in known cancer genes. RESULTS: We identified actionable pathogenic variants in 8 known hereditary cancer predisposition genes (PALB2, MLH1, MSH6, CHEK2, SMARCE1, ATM, BRCA1, and CTNNA1) in 9 patients, including 3 of 27 patients (11%) with familial UM and 6 of 127 patients (4.7%) with a high risk for cancer. Two patients showed pathogenic variants in CHEK2 and PALB2, whereas variants in the other genes each occurred in 1 patient. Biallelic inactivation of PALB2 and MLH1 was observed in tumors from the respective patients. The frequencies of pathogenic variants in PALB2, MLH1, and SMARCE1 in UM patients were significantly higher than the observed frequencies in noncancer controls (PALB2: P = 0.02; odds ratio, 8.9; 95% confidence interval, 1.5-30.6; MLH1: P = 0.04; odds ratio, 25.4; 95% confidence interval, 1.2-143; SMARCE1: P = 0.001; odds ratio, 2047; 95% confidence interval, 52-4.5e15, respectively). CONCLUSIONS: The study provided moderate evidence of gene and disease association of germline mutations in PALB2 and MLH1 with hereditary predisposition to UM. It also identified several other candidate susceptibility genes. The results suggest locus heterogeneity in predisposition to UM. Genetic testing for hereditary predisposition to cancer is warranted in UM patients with strong personal or family history of cancers, or both.

Germline large deletion of BAP1 and decreased expression in non-tumor choroid in uveal melanoma patients with high risk for inherited cancer.

Uveal melanoma (UM) is the most common phenotype in patients with germline BAP1 mutation. This study aimed to identify selection criteria for BAP1 germline testing and assessed the role of large deletion/duplication and epigenetic inactivation. One hundred seventy-two UM patients with high risk of hereditary cancer were included. Germline variants in BAP1 were assessed by direct sequencing and large deletion/duplication by multiplex ligation-dependent probe amplification. BAP1 expression in unaffected choroid tissue from a patient with UM was assessed by quantitative RT-PCR and methylation by pyrosequencing. Twenty-eight patients had one or more germline sequence variants in BAP1; seven of these were pathogenic. One hundred forty patients were assessed for large deletion/duplication and in one BAP1 whole gene deletion was detected. In total, eight patients (4.7%) had pathogenic alterations in BAP1 with the highest frequencies of in patients with a personal/family history of ≥2 BAP1-related cancers 6/16 (38%), age of onset <35 years 4/21 (19%) and familial UM 6/34 (18%). One of 19 non-tumor choroid tissues tested showed uncharacteristically low expression as compared to the controls decrease in BAP1 RNA expression but no evidence of constitutional promotor hypermethylation was detected. UM patients with a strong personal or family history of cancers associated with BAP1, early age of onset and familial UM should be assessed for germline variants in BAP1, including large deletions.

Known and novel ocular toxicities of biologics, targeted agents, and traditional chemotherapeutics.

PURPOSE: Increases in cancer with an aging population and the rapid development of new chemotherapeutics underscore the need for ophthalmologists to identify and manage potential ocular toxicities. This retrospective case series reports the ocular side effects of traditional and novel chemotherapeutic agents from a large center. METHODS: The medical records of 3537 adult patients 18 years and older who presented to an academic ophthalmology department on high-risk medications identified by ICD-9 search between January 2010 and February 2015 were reviewed. A cancer diagnosis, as well as a temporal association with chemotherapeutic use and ocular side effect, was deemed necessary for inclusion in the study. The main measures were ocular side effects in cancer patients taking chemotherapy, ocular imaging abnormalities, and the outcome of each side effect. RESULTS: Of the 161 oncology patients referred to the ophthalmology clinic for chemotherapeutic screening or ocular side effect, 31 (19.3%) were identified as having an ocular adverse reaction due to a novel or traditional chemotherapeutic medication. A novel flattening of the corneal curvature with hyperopic shift and corneal microcysts was identified in a patient taking the antibody-drug conjugate mirvetuximab soravtansine and was reversible with topical steroids. A bilateral medium-vessel choroidal vasculopathy with serous retinal detachment was seen with ipilimumab. The most frequent medication with ocular toxicity was interferon-α(2b) (IFN-α(2b)) (6/31, 19.4%); headache was typical in these patients (83.3%). Ibrutinib ocular toxicity was second most common (5/31, 16.1%), usually causing red or dry eye, while one patient developed branch retinal artery occlusion. Retinal abnormalities documented on OCT imaging occurred with IFN-α(2b), ipilimumab, binimetinib, and docetaxel, while rod-cone ERG abnormality was seen with cisplatin. Inflammatory conditions included anterior scleritis with zoledronic acid, focal eyelid inflammation with veliparib, bilateral chemosis with R-CHOP, iritis, and blepharospasm with IFN-α(2b). AION occurred with pemetrexed, and transient vision loss with hyperemic disc OS was seen with FOLFOX. Two patients (2/31, 6.5%) developed permanent vision loss. Six patients were lost to follow-up, and the clinical course was unknown (6/31, 19.4%). CONCLUSIONS AND RELEVANCE: Cases of permanent visual loss were observed; yet, in the majority of side effects, they improved with topical therapy and/or holding the medication. Further research is needed to elucidate the incidence and the pathophysiology of these side effects and maximize patient quality of life.

Analysis of the exome aggregation consortium (ExAC) database suggests that the BAP1-tumor predisposition syndrome is underreported in cancer patients.

The BAP1-tumor predisposition syndrome (BAP1-TPDS) has been recently identified to predispose patients to a variety of cancers and preneoplastic lesions. About 130 unrelated probands have been identified worldwide; however, the impact of the syndrome is suspected to be much larger given the diversity of the cancer phenotype. To evaluate the frequency of germline BAP1 mutations in the general and cancer populations, we analyzed the Exome Aggregation Consortium (ExAC), a database that contains 53105 exomes of unrelated individuals unaffected by cancer (general population) and exomes of 7601 unrelated individuals affected by cancer provided by the Cancer Genome Atlas (TCGA, cancer subjects). BAP1 null variants were seen at much higher frequency in the cancer subjects (0.0526%) compared to the general population (0.00188%) with a relative risk of 27.93 and (P = 0.0011, [95% CI: 3.122-249.883], Fisher's exact test). We also studied a reported BAP1 null variant, c.1203T > G, p.T401* (rs200156887), observed commonly in the general population. Sequencing and restriction fragment polymorphism of the RT-4 cell line that contains this variant revealed that it is in fact a 3bp deletion/insertion, c.1201_1203delinsGAG, a likely benign missense alteration p.Y401E explaining the relative high frequency of this variant in the general population. In conclusion, germline null mutations in BAP1 have a significantly higher frequency in cancer patients than the general population. Given the low frequency of reported families with BAP1-TPDS, our results suggest that the syndrome is underreported especially in patients with cancer.

Germline BAP1 alterations in familial uveal melanoma.

Uveal melanoma (UM) is the most commonly diagnosed primary intraocular tumor in adults. Familial UM (FUM), defined as two or more family members diagnosed with UM, is rare and estimated at less than 1% of all UM. Currently, BAP1 is the only gene known to contribute significant risk for UM. In this study we aimed to estimate the frequency of BAP1 mutation in FUM and to characterize the family and personal histories of other cancers in these families. We identified 32 families with FUM, including seven families previously reported by our group. BAP1 mutation testing was carried out by direct sequencing of the coding exons and the adjacent untranslated regions of the gene. Germline deletion and duplication analysis of BAP1 was assessed by multiplex ligation-dependent probe amplification (MLPA). Germline BAP1 mutations were found in 6/32 (19%) families. No deletions or duplications were identified in any of the 24 samples tested by MLPA. Combined with published studies, the frequency of BAP1 mutations was 14/64 (22%) in FUM. FUM families without BAP1 mutations have distinct family histories with high rates of prostate cancer in first- and second-degree relatives. It is likely that additional genes conferring risk for FUM exist. It is important to understand key shared features of FUM to focus future research on identifying these additional tumor predisposition syndromes. Though BAP1 should be tested first in these families, FUM families without BAP1 mutation should be explored for additional predisposition genes. © 2016 Wiley Periodicals, Inc.

Measurement of Perceived Stress in Age-Related Macular Degeneration.

PURPOSE: To validate the Perceived Stress Scale (PSS) in patients with age-related macular degeneration (AMD) using Rasch analysis. METHODS: Study participants with AMD were recruited from the retina service of the Department of Ophthalmology at the Ohio State University during clinical visits for treatment or observation. Visual acuity with habitual distance correction was assessed. A 10-item version of the PSS was administered in large print or by reading the items to the patient. Rasch analysis was used to investigate the measurement properties of the PSS, including fit to the model, ability to separate between people with different levels of perceived stress, category response structure performance, and unidimensionality. RESULTS: A total of 137 patients with a diagnosis of AMD were enrolled. The mean (±SD) age of participants was 82 ± 9 years. Fifty-four percent were female. Median Early Treatment of Diabetic Retinopathy Study (ETDRS) visual acuity of the better eye was 65 letters (Snellen 20/50), with a range of approximately 20/800 to 20/15. Forty-seven percent of participants were receiving an anti-VEGF injection on the day of the study visit. The response category structure was appropriate. One item, "How often have you felt confident in your ability to handle your personal problems?" was removed due to poor fit statistics. The remaining nine items showed good fit to the model, acceptable measurement precision as assessed by the Rasch person separation statistic, and unidimensionality. There was some evidence of differential item functioning by age and visual acuity. CONCLUSIONS: The Perceived Stress Scale demonstrated acceptable measurement properties and may be useful for the measurement of perceived stress in patients with AMD.

Expanding the clinical phenotype of hereditary BAP1 cancer predisposition syndrome, reporting three new cases.

The clinical phenotype of BAP1 hereditary cancer predisposition syndrome (MIM 614327) includes uveal melanoma (UM), cutaneous melanoma (CM), renal cell carcinoma (RCC), and mesothelioma. However, the frequency of the syndrome in patients with UM and the association with other cancers are still not clear. In this study, we screened 46 previously untested, unrelated UM patients with high risk for hereditary cancer for germline mutation in BAP1. We also studied four additional patients with a personal or family history suggestive of BAP1 hereditary cancer syndrome. We identified three patients with germline pathogenic mutations (c.2050 C>T, pGln684*; c.1182C>G, p.Tyr394*, and c.1882_1885delTCAC, p. Ser628Profs*8) in BAP1. Two of these three patients presented with UM and the third with a metastatic adenocarcinoma likely from a hepatic cholangiocarcinoma. Reported family histories included UM, mesothelioma, RCC, CM, and several other internal malignancies. The results of this study confirm the association between germline BAP1 mutation and predisposition to UM, mesothelioma, CM and RCC. However, other cancers, such as cholangiocarcinoma and breast carcinoma may be part of the phenotype of this hereditary cancer predisposition syndrome. In addition, the results support the existence of other candidate genes in addition to BAP1 contributing to hereditary predisposition to UM.

Monosomy 3 status of uveal melanoma metastases is associated with rapidly progressive tumors and short survival.

The aim of the study was to investigate the molecular genetics of uveal melanoma (UM) metastases and correlate it with disease progression. Twelve pathologically confirmed UM metastases from 11 patients were included. Molecular genetic alterations in chromosomes 3 (including the BAP1 region), 8q, 6p, and 1p were investigated by microsatellite genotyping. Mutations in codon 209 of GNAQ and GNA11 genes were studied by restriction-fragment length polymorphism (RFLP). We identified monosomy of chromosome 3 in tumors from four patients with an average survival of 5 months (range 1-8 months) from time of diagnosis of metastatic disease. In contrast, tumors with either disomy or partial chromosome 3 alterations showed significantly slower metastatic disease progression with an average survival of 69 months (range 40-123 months, p = 0.003). Alterations in chromosomal arms 1p, 6p, and 8q and mutations in either GNAQ or GNA11 showed no association with disease progression. Prominent mononuclear inflammatory infiltrate was observed in tumors from patients with slowly progressive disease. In conclusion, in UM metastases, monosomy 3 is associated with highly aggressive, rapidly progressive disease while disomy or partial change of 3 and prominent mononuclear inflammatory infiltrate in the tumor is associated with better prognosis. These findings should be considered when designing clinical trials testing effectiveness of various therapies of metastatic UM.

Germline BAP1 mutation predisposes to uveal melanoma, lung adenocarcinoma, meningioma, and other cancers.

OBJECTIVE: To investigate the potential contribution of germline sequence alterations in the BAP1 gene in uveal melanoma (UM) patients with possible predisposition to hereditary cancer. DESIGN: A total of 53 unrelated UM patients with high risk for hereditary cancer and five additional family members of one proband were studied. Mutational screening was carried out by direct sequencing. RESULTS: Of the 53 UM patients studied, a single patient was identified with a germline BAP1 truncating mutation, c. 799 C→T (p.Q267X), which segregated in several family members and was associated with UM and other cancers. Biallelic inactivation of BAP1 and decreased BAP1 expression were identified in the UM, lung adenocarcinoma and meningioma tumours from three family members with this germline BAP1 mutation. Germline BAP1 variants of uncertain significance, likely non-pathogenic, were also identified in two additional UM patients. CONCLUSION: This study reports a novel hereditary cancer syndrome caused by a germline BAP1 mutation that predisposes patients to UM, lung carcinoma, meningioma, and possibly other cancers. The results indicate that BAP1 is the candidate gene in only a small subset of hereditary UM, suggesting the contribution of other candidate genes.

Atypical choroidal nevus in a subject with a germline PALB2 pathogenic variant.

Recent evidence suggests that PALB2 variants may increase risk for the development of uveal melanoma and uveal melanocytic neoplasms. Here we report a case of an atypical choroidal nevus in a patient with a personal history of cancer and pathogenic PALB2 germline variant. A 75-year-old white female presented with an elevated predominantly amelanotic choroidal lesion OS. On examination and ophthalmic imaging, the mass measured 8.8 mm × 6.5 mm × 1.5 mm. The mass showed predominantly medium to high reflectivity on diagnostic A-scan and acoustic hollowing on B-scan. OCT over the lesion showed no subretinal fluid. The patient has a personal history of breast cancer and gastric adenoma and a strong family history of cancer. The patient was found to have a pathogenic truncating variant in PALB2 (rs118203998 c.3549C > A, p.Y1183*). Together with our previous findings of pathogenic PALB2 variants in uveal melanoma patients, this new finding of an atypical choroidal nevus in a patient with a pathogenic PALB2 germline variant suggests that pathogenic PALB2 variants may be a risk factor for uveal melanocytic neoplasms. This finding warrants further assessment of the prevalence and progression of uveal melanocytic neoplasms in PALB2 pathogenic variant carriers.

Use and Perceptions of Advanced Driver Assistance Systems by Older Drivers With and Without Age-Related Macular Degeneration.

Purpose: Advanced driver assistance systems (ADAS) have been reported to improve the safety of elderly and normally sighted drivers. The purpose of this study was to assess exposure to, perceived safety of, comfort level with, and interest in using ADAS among drivers with age-related macular degeneration (AMD). Methods: Current drivers aged 60+ years were recruited at four US sites to complete a survey about ADAS and driving habits. Frequency of use and/or perceptions of eight ADAS were investigated. An avoidance score was generated using questions about difficult driving situations. Results: The survey was completed by 166 participants (80 with AMD vs. 86 without). Participants with AMD had worse self-rated vision than those without (34% vs. 2% poor or fair rating), and drove fewer weekly miles (median [interquartile range [IQR] 30 [15 to 75] vs. 60 [30 to 121] miles, P = 0.002). Participants with AMD reported more avoidance of difficult driving situations (P < 0.001). There was no difference in the number of ADAS used by AMD status (median [IQR for AMD = 2.5 [1 to 5] vs. 3 [2 to 4] without, P = 0.87). Greater reported number of ADAS used was associated with less avoidance of difficult situations (P = 0.02). The majority perceived improved safety with most ADAS. Conclusions: Many drivers with AMD utilize common ADAS, which subjectively improve their road safety and may help to reduce self-imposed restrictions for difficult situations and mileage. Translational Relevance: Drivers with AMD are adopting readily available ADAS, for which they reported potential benefits, such as safety and less restrictive driving.

Frequency, molecular pathology and potential clinical significance of partial chromosome 3 aberrations in uveal melanoma.

The clinical significance of partial chromosome 3 alteration in uveal melanoma is still not clear. Also, the reported frequencies vary considerably in the published literature from 0 to 48%. The aims of the following study were to identify the frequency, molecular pathology and potential clinical significance of partial chromosome 3 alteration in uveal melanoma. We studied 47 uveal melanomas with an average follow-up of 36 months. Of these, 14 had confirmed metastasis. Allelic imbalance/loss of heterozygosity was studied using microsatellite markers on chromosome 3 enriched in markers located in the previously reported smallest regions of deletion overlap. Chromosomal alterations were assessed by conventional cytogenetics or comparative genomic hybridization (CGH) in a subset of patients. Utilizing genotyping, partial chromosome 3 alteration was detected in 14/47 tumors (30%). In the 23 tumors with available cytogenetic/CGH, partial chromosome 3 alteration was detected in 8/23 (38%) and was caused by both gains (4/8) and losses (4/8) of chromosome 3 with high frequency of complex chromosome 3 aberrations detected by cytogenetics. Out of the 14 tumors with confirmed metastasis, only 1 showed partial chromosome 3 alteration and the remaining showed monosomy 3. By limiting the aggressive disease marker to monosomy 3, genotyping showed 93% sensitivity and 67% specificity for detection of aggressive uveal melanoma. In conclusion, partial chromosome 3 alterations are common in uveal melanoma and mostly caused by complex cytogenetic changes leading to partial gains and/or partial losses of chromosome 3. Partial chromosome 3 alteration is not likely to be associated with highly aggressive uveal melanoma that metastasizes within the first 3 years after treatment. Microsatellite-based genotyping of chromosome 3 is highly sensitive for detection of aggressive uveal melanoma.

Molecular genetic testing of uveal melanoma from routinely processed and stained cytology specimens.

In the following study we investigated the utility of molecular genetic testing of the DNA extracted from routinely stained and processed smears from uveal melanoma (UM). Smears from five uveal melanoma cell lines and 12 primary tumors were prepared and stained with Papanicolaou and Romanowsky stains. Genotyping was carried out utilizing 14 microsatellite markers on chromosomes 3, 6 and 8. Mutational screening for alterations in GNAQ and GNA11 genes was carried out by restriction fragment length polymorphism. The results were compared to those obtained through direct sequencing of frozen tumor tissues. High quality DNA was extracted from the stained slides with no difference in the efficiency of DNA extraction between the two staining techniques. The extracted DNA was of adequate quality for genotyping and mutational screening. DNA extracted from approximately 200 tumor cells is sufficient for reproducible testing of allelic imbalances and for studying the common somatic mutations in GNAQ and GNA11 genes. In conclusion, we presented the feasibility of utilizing routinely stained cytology smears from UM for molecular genetic testing. The DNA obtained is of sufficient quality to carry out genotyping for markers on chromosome 3, 6 and 8, as well as screening for somatic mutations in GNAQ and GNA11 genes.

Heterogeneity in Mitogen-Activated Protein Kinase (MAPK) Pathway Activation in Uveal Melanoma With Somatic GNAQ and GNA11 Mutations.

Purpose: The activation of the mitogen-activated protein kinase (MAPK) pathway has been suggested as the major downstream target when GNAQ and GNA11 (GNAQ/11) are mutated in uveal melanoma (UM). However, clinical trials with single agent MEK inhibitor showed no clinical significance in altering the overall outcome of the disease in UM; therefore, we investigated the correlation between naturally occurring mutations in GNAQ/11 and activation of MAPK pathway in vivo in primary UM. Methods: Screening for activating mutations in codons 183 and 209 of GNAQ/11 was carried out by sequencing and restriction fragment length polymorphism (RFLP) in a cohort of 42 primary UM. Activation of the MAPK pathway and other potential downstream signals was assessed by immunohistochemistry and/or Western blot analysis. Potential downstream signaling of mutant and wild type GNAQ/11 was studied by transient transfection assay in nonmutant cell lines. Results: Somatic mutations in GNAQ/11 were observed in 35/42 (83.3%) of primary UM. Tumors with GNAQ/11 mutations showed variations in the activation of ERK1/2 with significant tumor heterogeneity. Weak and undetectable ERK1/2 activation was observed in 4/35 (11.4%) and 8/35 (22.9%) of the GNAQ/11 mutant UM, respectively. Tumor heterogeneity of GNAQ/11 mutations was also observed in a subset of tumors. Conclusions: Our results indicate that there is marked variation in MAPK activation in UM with GNAQ/11 mutations. Thus, GNAQ/11 mutational status is not a sufficient biomarker to adequately predict UM patient responses to single-agent selective MEK inhibitor therapy.

Comprehensive Study of the Clinical Phenotype of Germline BAP1 Variant-Carrying Families Worldwide.

Background: The BRCA1-associated protein-1 (BAP1) tumor predisposition syndrome (BAP1-TPDS) is a hereditary tumor syndrome caused by germline pathogenic variants in BAP1 encoding a tumor suppressor associated with uveal melanoma, mesothelioma, cutaneous melanoma, renal cell carcinoma, and cutaneous BAP1-inactivated melanocytic tumors. However, the full spectrum of tumors associated with the syndrome is yet to be determined. Improved understanding of the BAP1-TPDS is crucial for appropriate clinical management of BAP1 germline variant carriers and their families, including genetic counseling and surveillance for new tumors. Methods: We collated germline variant status, tumor diagnoses, and information on BAP1 immunohistochemistry or loss of somatic heterozygosity on 106 published and 75 unpublished BAP1 germline variant-positive families worldwide to better characterize the genotypes and phenotypes associated with the BAP1-TPDS. Tumor spectrum and ages of onset were compared between missense and null variants. All statistical tests were two-sided. Results: The 181 families carried 140 unique BAP1 germline variants. The collated data confirmed the core tumor spectrum associated with the BAP1-TPDS and showed that some families carrying missense variants can exhibit this phenotype. A variety of noncore BAP1-TPDS -associated tumors were found in families of variant carriers. Median ages of onset of core tumor types were lower in null than missense variant carriers for all tumors combined (P < .001), mesothelioma (P < .001), cutaneous melanoma (P < .001), and nonmelanoma skin cancer (P < .001). Conclusions: This analysis substantially increases the number of pathogenic BAP1 germline variants and refines the phenotype. It highlights the need for a curated registry of germline variant carriers for proper assessment of the clinical phenotype of the BAP1-TPDS and pathogenicity of new variants, thus guiding management of patients and informing areas requiring further research.

Endogenous endophthalmitis and osteomyelitis associated with interleukin 17 inhibitor treatment for psoriasis in a patient with diabetes.

A 64-year-old man with type 2 diabetes mellitus and plaque psoriasis presented to the emergency room with 3 days of progressive right eye pain and decreased vision. After extensive workup and multidisciplinary team effort, the patient was diagnosed with and treated for unilateral endogenous methicillin-sensitive Staphylococcus aureus endophthalmitis, bacteraemia and osteomyelitis of the foot. The patient had been started on the interleukin 17 (IL-17) inhibitor secukinumab for his treatment-resistant plaque psoriasis 4 weeks prior to presentation. After treatment, his final vision was light perception and the foot infection resolved without sequelae. To our knowledge, this is the first reported case of both endogenous endophthalmitis and osteomyelitis associated with an IL-17 inhibitor.

Expression of vascular endothelial growth factor in uveal melanoma is independent of 6p21-region copy number.

PURPOSE: Overexpression of vascular endothelial growth factor (VEGF) and overrepresentation of the 6p region have been reported with a wide variation in uveal melanoma. The aim of the current study is to identify the frequency of copy number alteration in the 6p21 region and its correlation with the expression of VEGF in uveal melanoma. EXPERIMENTAL DESIGN: We studied 88 uveal melanomas for copy number change in the 6p region by comparative genomic hybridization and/or chromogenic in situ hybridization. Expression of VEGF protein was estimated by immunohistochemistry. In 15 tumors, VEGF mRNA expression was also studied by quantitative reverse transcription-PCR (RT-PCR) and VEGF splice variants were detected by RT-PCR. RESULTS: Copy number of the 6p21 region was successfully estimated in 37 tumors. In 10 (27%) of those, overrepresentation of the 6p21 region was detected. There was no statistically significant difference in VEGF expression between tumors with and without gain of 6p21 (P = 0.82). VEGF expression was not confined to the tumors and was also detected in the surrounding normal tissue. Expression of VEGF, detected by quantitative RT-PCR, was concordant with expression of VEGF protein. Different VEGF isoforms were expressed in different tumors with no obvious correlation with disease status. CONCLUSION: VEGF is overexpressed in a significant number of uveal melanomas. It should be noted that VEGF is not a candidate oncogene in uveal melanoma with 6p gain/amplification. VEGF overexpression other than structural amplification is probably significant in the pathogenesis of uveal melanomas, and its mechanism must be sought.

Germline BAP1 mutations misreported as somatic based on tumor-only testing.

We present three unrelated patients with germline mutations in BAP1 misreported as somatic mutations. All had strong family histories of cancer. One of these patients presented with an invasive breast cancer with the tumor tissue showing partial loss of the mutant rather than the wild type allele, suggesting that the germline BAP1 mutation didn't contribute to breast cancer development in this patient. This data highlights the importance of sequencing matching germline and tumor DNA for proper assessment of somatic versus germline mutation status. In patients with somatic mutations reported from laboratories carrying out tumor-only genomic testing, the possibility that a variant may be a germline mutation should be considered, especially if the personal and/or family history suggests hereditary cancer predisposition. Since tumor-only testing can reveal germline mutations, ethical issues for patients being tested should be considered including proper consent and genetic counseling.