InterMEL: An international biorepository and clinical database to uncover predictors of survival in early-stage melanoma.

INTRODUCTION: We are conducting a multicenter study to identify classifiers predictive of disease-specific survival in patients with primary melanomas. Here we delineate the unique aspects, challenges, and best practices for optimizing a study of generally small-sized pigmented tumor samples including primary melanomas of at least 1.05mm from AJTCC TNM stage IIA-IIID patients. We also evaluated tissue-derived predictors of extracted nucleic acids' quality and success in downstream testing. This ongoing study will target 1,000 melanomas within the international InterMEL consortium. METHODS: Following a pre-established protocol, participating centers ship formalin-fixed paraffin embedded (FFPE) tissue sections to Memorial Sloan Kettering Cancer Center for the centralized handling, dermatopathology review and histology-guided coextraction of RNA and DNA. Samples are distributed for evaluation of somatic mutations using next gen sequencing (NGS) with the MSK-IMPACTTM assay, methylation-profiling (Infinium MethylationEPIC arrays), and miRNA expression (Nanostring nCounter Human v3 miRNA Expression Assay). RESULTS: Sufficient material was obtained for screening of miRNA expression in 683/685 (99%) eligible melanomas, methylation in 467 (68%), and somatic mutations in 560 (82%). In 446/685 (65%) cases, aliquots of RNA/DNA were sufficient for testing with all three platforms. Among samples evaluated by the time of this analysis, the mean NGS coverage was 249x, 59 (18.6%) samples had coverage below 100x, and 41/414 (10%) failed methylation QC due to low intensity probes or insufficient Meta-Mixed Interquartile (BMIQ)- and single sample (ss)- Noob normalizations. Six of 683 RNAs (1%) failed Nanostring QC due to the low proportion of probes above the minimum threshold. Age of the FFPE tissue blocks (p<0.001) and time elapsed from sectioning to co-extraction (p = 0.002) were associated with methylation screening failures. Melanin reduced the ability to amplify fragments of 200bp or greater (absent/lightly pigmented vs heavily pigmented, p<0.003). Conversely, heavily pigmented tumors rendered greater amounts of RNA (p<0.001), and of RNA above 200 nucleotides (p<0.001). CONCLUSION: Our experience with many archival tissues demonstrates that with careful management of tissue processing and quality control it is possible to conduct multi-omic studies in a complex multi-institutional setting for investigations involving minute quantities of FFPE tumors, as in studies of early-stage melanoma. The study describes, for the first time, the optimal strategy for obtaining archival and limited tumor tissue, the characteristics of the nucleic acids co-extracted from a unique cell lysate, and success rate in downstream applications. In addition, our findings provide an estimate of the anticipated attrition that will guide other large multicenter research and consortia.

Risk Factors for Keratinocyte Carcinoma in Recipients of Allogeneic Hematopoietic Cell Transplants.

Importance: Allogeneic hematopoietic cell transplant (alloHCT) is known to increase the risk for keratinocyte carcinoma. The extent to which host characteristics, including pigmentary phenotype and UV radiation exposure, contribute is unknown. Objective: To identify and validate independent risk factors for keratinocyte carcinoma after alloHCT, including those associated with the transplant and the host. Design, Setting, and Participants: This retrospective cohort study analyzed a consecutive sample of alloHCT recipients from January 1, 2000, to December 31, 2014, at the Mayo Clinic, Rochester, Minnesota (n = 872) and University Hospitals Cleveland Medical Center, Cleveland, Ohio (n = 147). Participants from the Mayo Clinic were randomly allocated (2:1) into discovery (n = 581) and validation (n = 291) cohorts. Time to first keratinocyte carcinoma and information about transplant- and host-associated risk factors were extracted. A multivariate keratinocyte carcinoma risk model was created using a stepwise Cox proportional hazards regression model with P ≤ .05 for entry that incorporated all covariates that were individually statistically significant at α = 0.05 in the discovery cohort. The risk model was first internally validated using the Mayo Clinic validation cohort and then externally validated using the independent cohort of alloHCT recipients at University Hospitals Cleveland Medical Center. Data were analyzed from March 13, 2018, to June 12, 2019. Exposures: Allogeneic hematopoietic cell transplant. Main Outcomes and Measures: The primary outcome was time to development of the first cutaneous keratinocyte carcinoma after alloHCT; secondary outcome, time to development of the first individual basal and/or squamous cell carcinoma after alloHCT. Results: Of the 872 alloHCT recipients identified in the Mayo Clinic cohort (520 men [59.6%]; mean [SD] age, 48.3 [12.6] years), 95 (10.9%) developed keratinocyte carcinoma after alloHCT during 5349 person-years of follow-up. Of the 147 alloHCT recipients in the exernal validation cohort (86 men [58.5%]; mean [SD] age, 47.9 [17.5] years), 18 (12.2%) developed keratinocyte carcinoma after alloHCT in 880 person-years of follow up. Risk factors independently associated with keratinocyte carcinoma after alloHCT included age (hazard ratio [HR] per 10 years, 1.72; 95% CI, 1.21-2.42), chronic lymphocytic leukemia (HR, 2.47; 95% CI, 1.20-5.09), clinically photodamaged skin (HR, 3.47; 95% CI, 1.87-6.41), and history of cutaneous squamous cell carcinoma (HR, 2.60; 95% CI, 1.41-5.91). Harrell concordance statistics were 0.81 (95% CI, 0.72-0.90) and 0.86 (95% CI, 0.74-0.98) for internal and external validation of the keratinocyte carcinoma risk model, respectively. Conclusions and Relevance: This study found validated independent risk factors for keratinocyte carcinoma after alloHCT that are enriched with host- compared with transplant-associated risk factors. These findings highlight the importance of assessing host-associated risk factors for keratinocyte carcinoma in patients eligible for alloHCT. Future studies should examine whether keratinocyte carcinoma risk stratification before alloHCT may inform long-term surveillance strategies.

Immune checkpoint inhibitor-induced sarcoidosis-like granulomas.

Immune checkpoint inhibitors targeting the cytotoxic T lymphocyte-associated antigen-4 and programmed cell death-1 receptors have transformed the treatment of melanoma and other cancers. These therapies are associated with a number of side effects, including immune-related adverse events. Sarcoidosis-like granulomas (SLGs) are important immune checkpoint inhibitor-related reactions to recognize as SLGs can mimic disease progression and accordingly impact treatment decisions. We systematically review reports of immune checkpoint inhibitor-induced SLGs in cancer patients and discuss potential underlying pathophysiological mechanisms.

Identification of gene expression levels in primary melanoma associated with clinically meaningful characteristics.

Factors influencing melanoma survival include sex, age, clinical stage, lymph node involvement, as well as Breslow thickness, presence of tumor-infiltrating lymphocytes based on histological analysis of primary melanoma, mitotic rate, and ulceration. Identification of genes whose expression in primary tumors is associated with these key tumor/patient characteristics can shed light on molecular mechanisms of melanoma survival. Here, we show results from a gene expression analysis of formalin-fixed paraffin-embedded primary melanomas with extensive clinical annotation. The Cancer Genome Atlas data on primary melanomas were used for validation of nominally significant associations. We identified five genes that were significantly associated with the presence of tumor-infiltrating lymphocytes in the joint analysis after adjustment for multiple testing: IL1R2, PPL, PLA2G3, RASAL1, and SGK2. We also identified two genes significantly associated with melanoma metastasis to the regional lymph nodes (PIK3CG and IL2RA), and two genes significantly associated with sex (KDM5C and KDM6A). We found that LEF1 was significantly associated with Breslow thickness and CCNA2 and UBE2T with mitosis. RAD50 was the gene most significantly associated with survival, with a higher level of expression associated with worse survival.

Stage IV melanoma of unknown primary: A population-based study in the United States from 1973 to 2014.

BACKGROUND: Melanoma of unknown primary (MUP) is incompletely described on a population level. OBJECTIVE: We sought to characterize stage IV MUP in a population-based cancer registry. METHODS: We developed a novel search algorithm to identify cases of stage IV MUP in the Surveillance, Epidemiology, and End Results 18 registries from 1973 to 2014. Cases of stage IV melanoma of known primary (MKP) served as a comparison group. Age-standardized incidence rates, demographic characteristics, adjusted disease-specific survival, and Cox proportional hazard models were calculated for MUP and MKP. RESULTS: A total of 322 stage IV MUP cases and 12,796 stage IV MKP cases were identified in Surveillance, Epidemiology, and End Results 18 registries from 1973 to 2014. The incidence of stage IV MUP is increasing, particularly for patients younger than 30 years of age. In multivariate analyses, age older than 50 and a lack of surgical treatment were negative prognostic factors for stage IV MUP. Relative survival, but not 5-year adjusted disease-specific survival, was higher for stage IV MUP than for MKP. LIMITATIONS: Limitations include the retrospective study design and possible misclassification of MUP. CONCLUSIONS: The incidence of stage IV MUP is increasing, and stage IV MUP shares similar prognostic factors with stage IV MKP, including age and surgical treatment.

Primary bilateral uveal melanoma: a population-based study and systematic review.

IMPORTANCE: Primary bilateral uveal melanoma (UM) is a rare and incompletely described entity. It is not known how these patients compare to those with unilateral UM. BACKGROUND: We sought to comprehensively characterize and compare patients with primary bilateral and unilateral UM. DESIGN: Retrospective, population-based and systematic review. PARTICIPANTS: Patients with bilateral (n = 52) and unilateral UM (n = 8915). METHODS: We analysed cases of primary bilateral UM from three data sources: (i) the University Hospitals Cleveland Medical Center pathology database from 1996 to 2016 (n = 1); (ii) the Surveillance, Epidemiology and End-Results (SEER)-18 database from 1973 to 2013 (n = 5) and (iii) a systematic review of the English language literature (n = 46). Cases of unilateral UM were obtained from the SEER-18 database from 1973 to 2013 for comparison (n = 8915). MAIN OUTCOME MEASURES: Demographics, clinicopathological characteristics, treatments and survival. RESULTS: There were no differences in sex, race, mean age at diagnosis, site of uveal involvement, metastases at diagnosis, or treatment among patients with bilateral as compared to unilateral UM. Additionally, there were no clinicopathological differences between the two UMs in each patient with bilateral disease. Overall survival did not differ between unilateral and bilateral UM patients, or between bilateral UM patients who presented with, or subsequently developed, bilateral disease. CONCLUSIONS AND RELEVANCE: Bilateral and unilateral UM patients share similar demographics, clinicopathological characteristics, treatments and prognoses. Moreover, the development of bilateral disease does not portend a poorer prognosis and patients should be treated similarly to those with unilateral disease.

Melanoma subtypes demonstrate distinct PD-L1 expression profiles.

PD-L1 expression in the tumor immune microenvironment is recognized as both a prognostic and predictive biomarker in patients with cutaneous melanoma, a finding closely related to its adaptive (IFN-γ-mediated) mechanism of expression. Approximately 35% of cutaneous melanomas express PD-L1, however, the expression patterns, levels, and prevalence in rarer melanoma subtypes are not well described. We performed immunohistochemistry for PD-L1 and CD8 on 200 formalin-fixed paraffin-embedded specimens from patients with acral (n=16), mucosal (n=36), uveal (n=103), and chronic sun-damaged (CSD) (n=45) melanomas (24 lentigo maligna, 13 'mixed' desmoplastic, and 8 'pure' desmoplastic melanomas). CD8+ tumor-infiltrating lymphocyte (TIL) densities were characterized as mild, moderate, or severe, and their geographic association with PD-L1 expression was evaluated. Discrete lymphoid aggregates, the presence of a spindle cell morphology, and the relationship of these features with PD-L1 expression were assessed. PD-L1 expression was observed in 31% of acral melanomas, 44% of mucosal melanomas, 10% of uveal melanomas, and 62% of CSD melanomas (P<0.0001). Compared to our previously characterized cohort of cutaneous melanomas, the proportion of PD-L1(+) tumors was lower in uveal (P=0.0002) and higher in CSD (P=0.0073) melanomas, while PD-L1 expression in the acral and mucosal subtypes was on par. PD-L1 expression in all subtypes correlated with a moderate-severe grade of CD8+ TIL (all, P<0.003), supporting an adaptive mechanism of expression induced during the host antitumor response. The tumor microenvironments observed in CSD melanomas segregated by whether they were the pure desmoplastic subtype, which showed lower levels of PD-L1 expression when compared to other CSD melanomas (P=0.047). The presence of lymphoid aggregates was not associated with the level of PD-L1 expression, while PD-L1(+) cases with spindle cell morphology demonstrated higher levels of PD-L1 than those with a nested phenotype (P<0.0001). Our findings may underpin the reported clinical response rates for anti-PD-1 monotherapy, which vary by subtype.

Epidemiology of genitourinary melanoma in the United States: 1992 through 2012.

BACKGROUND: Primary melanoma arising in the genitourinary tract is rare and poorly characterized. OBJECTIVES: We sought to describe the epidemiology of genitourinary melanoma in the United States. METHODS: Incident case and population data were obtained for genitourinary melanoma from the Surveillance, Epidemiology, and End Results 13 Registries Database between 1992 and 2012. RESULTS: A total of 817 patients with genitourinary melanoma were identified; most cases occurred in the vulva. The incidence of genitourinary melanoma was much higher in women (1.74/1 million person-years) than men (0.17/1 million person-years). The highest rates occurred among non-Hispanic white women aged 85 years and older. Five-year melanoma-specific and overall survival were poor at 52.4% and 36.3%, respectively. Predictors of poor survival were increasing age, black race, and female sex. LIMITATIONS: The study population is small, therefore some rates reported may be unstable. In addition, cutaneous, mucosal, and other extracutaneous surfaces of the genitourinary tract cannot be reliably distinguished in Surveillance, Epidemiology, and End Results. Furthermore, melanomas may be underreported to cancer registries. CONCLUSION: From 1992 to 2012, genitourinary melanoma was 10 times more common in women than men. Survival was poor in women compared with men, which is different from cutaneous melanoma where women have a survival advantage.

Cutaneous manifestations associated with melanoma.

Melanoma is a malignancy most commonly arising from the skin; therefore, primary melanoma characteristics are usually the first cutaneous manifestations of melanoma. Cutaneous metastases, which can occur locally or diffusely, are important to detect in a timely manner as treatments for advanced melanoma that impact survival are now available. Melanoma can be associated with local or diffuse pigmentation changes, including depigmentation associated with the leukodermas and hyperpigmentation associated with diffuse melanosis cutis. The leukodermas occur frequently, illustrate the immunogenic nature of melanoma, and may impact prognosis. Paraneoplastic syndromes in association with melanoma are rare, though can occur.

Parotid melanoma of unknown primary.

PURPOSE: Rarely, melanoma is discovered in the parotid gland without an identifiable primary site. It is not known how patients with parotid melanoma of unknown primary (PMUP) compare to those with a known primary (PMKP). As such, we describe the largest series of patients with PMUP to date and compare them to patients with PMKP. METHODS: We analyzed cases from three sources: (1) the University Hospitals Case Medical Center pathology database (n = 45), (2) the Surveillance, Epidemiology, and End Results 18 database (n = 33), and (3) a comprehensive literature search (n = 32). RESULTS: PMUP patients were predominately male and presented at a mean age of 56 years. When compared to PMKP, PMUP cases were more likely to be diagnosed in the parotid parenchyma, present with stage IV disease, and develop distant metastases during follow-up in a shorter amount of time. However, there was no difference in overall survival between patients with PMUP and PMKP presenting with stage-matched disease. CONCLUSIONS: Overall survival is similar for stage-matched patients with parotid melanoma presenting with an unknown and known primary site.

Epidemiology of Anorectal Melanoma in the United States: 1992 to 2011.

BACKGROUND: Anorectal melanoma is a rare type of malignant melanoma and thus the epidemiology of patients with this tumor has been poorly defined. OBJECTIVE: To describe the epidemiology of anorectal melanoma in the United States. METHODS AND MATERIALS: We obtained case and population data from the Surveillance, Epidemiology, and End Results 13 Registries Database (SEER 13) between 1992 and 2011 using rectal diagnostic codes C20.9 to 21.8 and ICD-O-3 melanoma codes 8720 to 8721 and 8742 to 8746. RESULTS: There were 260 primary anorectal melanomas in SEER 13 from 1992 to 2011, occurring mostly in the rectum. The incidence of anorectal melanoma was higher among women than men with the highest rates occurring among white Hispanics ages 65 to 74 years. During this time period, the age-adjusted incidence rates rose significantly (p < .05) for both women and men with estimated annual percentage changes of 3.02% and 5.08%, respectively. Overall and melanoma-specific survival was poor irrespective of gender or ethnicity. CONCLUSION: Anorectal melanoma in the United States is increasing in both men and women, with the highest rates in elderly Hispanic white women. Hispanic whites were more likely to develop anorectal melanoma than non-Hispanic whites, suggesting that this population may be targeted for screening interventions. These results warrant further investigation to better understand the gender, racial, ethnic, and geographic variations for anorectal melanomas.

A systematic review and meta-analysis of animal-type melanoma.

BACKGROUND: Animal-type melanoma is a rare subtype of melanoma with heavily pigmented dermal epithelioid and spindled melanocytes. Its classification as a subtype of melanoma versus a borderline melanocytic tumor is debated. OBJECTIVES: Our primary objective was to characterize the demographics, clinical presentation, histopathology, management, and outcomes of patients with animal-type melanoma. METHODS: We performed a systematic review and meta-analysis of the English-language literature on animal-type melanoma. RESULTS: We identified 190 cases of animal-type melanoma. They occurred equally in men and women, with Caucasians (53.7%) most commonly affected. The median Breslow depth was 3.8 mm; ulceration was reported present in 15.8%; and dermal mitoses greater than or equal to 1/mm(2) was reported in 27.4%. The most common initial management was wide local excision with sentinel lymph node biopsy (55.7%). In all, 78 patients underwent sentinel lymph node biopsy with 41.0% positivity rate. A total of 32 patients underwent completion lymph node dissection with 34.4% positivity rate. Locoregional recurrence was reported in 15 patients, recurrence with distant metastases in 6 patients, and death in 5 patients. LIMITATIONS: Data were obtained from small studies with limited follow-up. There is no universally accepted definition of animal-type melanoma. CONCLUSION: Prospective studies with complete staging information and molecular profiling may allow further characterization of this tumor.

Cutaneous Malignant Neoplasms in Hematopoietic Cell Transplant Recipients: A Systematic Review.

IMPORTANCE: Hematopoietic cell transplantation has increased the survival of patients with several types of malignant hematologic disease and hematologic disorders; however, these patients have an increased risk of posttransplant cutaneous malignant neoplasms. Physicians should be aware of associated risk factors to provide appropriate patient screening and long-term care. OBJECTIVE: To identify the incidence and risk factors for cutaneous malignant neoplasms following hematopoietic cell transplantation. EVIDENCE REVIEW: A systematic review was conducted using Medline and Cochrane databases from January 1995 to December 2013. Retrospective and prospective reviews containing at least 100 patients who underwent hematopoietic cell transplantation reporting skin cancer as a primary outcome were included. Information regarding the entire cohort, data for the subset who developed cutaneous malignant neoplasms, and cutaneous malignant neoplasm risk factors were extracted from included articles. The level of evidence for each study was assessed using the Strength of Recommendation Taxonomy scale. FINDINGS: Patients who underwent hematopoietic cell transplantation had an increased risk of squamous cell carcinoma, basal cell carcinoma, and melanoma. Factors such as primary disease, chronic graft-vs-host disease, prolonged immunosuppression, radiation exposure, light skin color, sex, and T-cell depletion are risk factors for cutaneous malignant neoplasms. CONCLUSIONS AND RELEVANCE: Given the increased risk of cutaneous malignant neoplasms in hematopoietic cell transplant recipients, this population should be educated on skin self-examination and pursue regular follow-up with dermatologists.

Update on primary mucosal melanoma.

Mucosal melanomas are aggressive cancers of mucosal surfaces with clinical and pathologic characteristics distinct from cutaneous melanomas, warranting different staging systems and treatment approaches. Surgical resection is performed frequently for the primary tumor, although the utility of lymph node surgery and radiation therapy is not established. Therapies targeted against C-KIT activating mutations, identified in many mucosal melanomas, are emerging as promising treatments.