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Purpose: Activation of the mitogen-activated protein kinase (MAPK) pathway and the phosphatidylinositol 3-kinase/AKT pathway seems to be critical for melanoma proliferation. Components of these pathways are client proteins of heat-shock protein 90 (hsp90), suggesting that inhibition of hsp90 could have significant antimelanoma effects. We conducted a phase II trial using the hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) in melanoma patients. The primary end points were clinical responses and whether treatment inhibited MAPK pathway activity.Experimental Design: Melanoma patients with measurable disease were stratified on the basis of whether or not their tumor harbored a V600E BRAF mutation. The hsp90 inhibitor 17-AAG was administered i.v. once weekly ×6 weeks at 450 mg/m2. Tumor biopsies were obtained pretreatment and 18 to 50 hours after the first dose of 17-AAG, and were snap-frozen.Results: Fifteen evaluable patients were treated; nine had BRAF mutations and six were wild-type. No objective responses were observed. Western blot analysis of tumor biopsies showed an increase in hsp70 and a decrease in cyclin D1 expression in the posttreatment biopsies but no significant effect on RAF kinases or phospho–extracellular signal-regulated kinase expression. Plasma analyzed by mutant-specific PCR for V600E BRAF showed 86% sensitivity and 67% specificity in predicting tumor DNA sequencing results.Conclusions: At this dose and schedule of 17-AAG, the effects of 17-AAG on RAF kinase expression were short-lived, and no objective antimelanoma responses were seen. Future trials in melanoma should focus on a more potent hsp90 inhibitor or a formulation that can be administered chronically for a more prolonged suppression of the MAPK pathway.

Purpose: Both phosphatidylinositol 3-kinase/AKT and RAS/mitogen-activated protein kinase signal transduction pathways mediate 4E-BP1 phosphorylation, releasing 4E-BP1 from the mRNA cap and permitting translation initiation. Given the prevalence of PTEN and BRAF mutations in melanoma, we first examined translation initiation, as measured by phosphorylated 4E-BP1 (p-4E-BP1), in metastatic melanoma tissues and cell lines. We then tested the association between amounts of total and p-4E-BP1 and patient survival.Experimental Design: Seven human metastatic melanoma cells lines and 72 metastatic melanoma patients with accessible metastatic tumor tissues and extended follow-up information were studied. Expression of 4E-BP1 transcript, total 4E-BP1 protein, and p-4E-BP1 was examined. The relationship between 4E-BP1 transcript and protein expression was assessed in a subset of patient tumors (n = 41). The association between total and p-4E-BP1 levels and survival was examined in the larger cohort of patients (n = 72).Results: 4E-BP1 was hyperphosphorylated in 4 of 7 melanoma cell lines harboring both BRAF and PTEN mutations compared with untransformed melanocytes or RAS/RAF/PTEN wild-type melanoma cells. 4E-BP1 transcript correlated with 4E-BP1 total protein levels as measured by the semiquantitative reverse-phase protein array (P = 0.012). High levels of p-4E-BP1 were associated with worse overall and post-recurrence survival (P = 0.02 and 0.0003, respectively).Conclusion: Our data show that translation initiation is a common event in human metastatic melanoma and correlates with worse prognosis. Therefore, effective inhibition of the pathways responsible for 4E-BP1 phosphorylation should be considered to improve the treatment outcome of metastatic melanoma patients.

Purpose: In certain cancers, MDM2 SNP309 has been associated with early tumor onset in women. In melanoma, incidence rates are higher in women than in men among individuals less than 40 years of age, but among those older than 50 years of age, melanoma is more frequent in men than in women. To investigate this difference, we examined the association among MDM2 SNP309, age at diagnosis, and gender among melanoma patients.Experimental Design: Prospectively enrolled melanoma patients (N = 227) were evaluated for MDM2 SNP309 and the related polymorphism, p53 Arg72Pro. DNA was isolated from patient blood samples, and genotypes were analyzed by PCR-restriction fragment length polymorphism. Associations among MDM2 SNP309, p53 Arg72Pro, age at diagnosis, and clinicopathologic features of melanoma were analyzed.Results: The median age at diagnosis was 13 years earlier among women with a SNP309 GG genotype (46 years) compared with women with TG+TT genotypes (59 years; P = 0.19). Analyses using age dichotomized at each decade indicated that women with a GG genotype had significantly higher risks of being diagnosed with melanoma at ages P = 0.01). Similar observations were not seen among men.Conclusions: Our data suggest that MDM2 may play an important role in the development of melanoma in women. The MDM2 SNP309 genotype may help identify women at risk of developing melanoma at a young age.

Supplementary Data from Phase II Trial of 17-Allylamino-17-Demethoxygeldanamycin in Patients with Metastatic Melanoma

Supplementary Table S1 from Association of MDM2 SNP309, Age of Onset, and Gender in Cutaneous Melanoma

Supplementary Table 4 from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Supplementary Methods from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Supplementary Table 5 from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Supplementary Table 2 from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Supplementary Table 3 from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Supplementary Table 1 from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Supplementary Table 6 from Integrative Genomics Identifies Molecular Alterations that Challenge the Linear Model of Melanoma Progression

Introduction: Nevisense is a non-invasive device that measures electrical impedance spectroscopy (EIS) of individual skin lesions to aid in the diagnosis of melanoma. While EIS has demonstrated high sensitivity in diagnosing melanoma, its impact on a clinician’s diagnostic confidence remains unknown. Objective: To conduct a pilot study to evaluate whether clinician diagnostic confidence, sensitivity, specificity and accuracy can be increased by adding EIS measurement scores to clinical and dermoscopic images of lesions clinically suspicious for melanoma. Methods: Three pigmented lesions specialists and three 4th year medical students completed an online survey to evaluate 34 melanocytic lesions suspicious for melanoma. For each lesion, participants provided their diagnosis, biopsy recommendation, and confidence in diagnosing a lesion as benign or malignant based on history and clinical and dermoscopic images, and again after receiving an EIS score. Results: Addition of EIS scores increased mean biopsy sensitivity for melanoma/severely dysplastic nevi from 70% to 84% (p = .014) and mean diagnostic accuracy from 74% to 86% (p = .005). Mean diagnostic confidence increased for all histopathologic categories for both students and dermatologists (all p < .05). Conclusions: In this pilot study, EIS increased novice and expert diagnosticians’ confidence regarding dermoscopically equivocal melanocytic lesions. Further studies are needed to explore how EIS can help clinicians reassure patients regarding the management of clinically dysplastic melanocytic nevi.

ImportanceTherapy for advanced melanoma has transformed during the past decade, but early detection and prognostic assessment of cutaneous melanoma (CM) remain paramount goals. Best practices for screening and use of pigmented lesion evaluation tools and gene expression profile (GEP) testing in CM remain to be defined.ObjectiveTo provide consensus recommendations on optimal screening practices and prebiopsy diagnostic, postbiopsy diagnostic, and prognostic assessment of CM.Evidence ReviewCase scenarios were interrogated using a modified Delphi consensus method. Melanoma panelists (n = 60) were invited to vote on hypothetical scenarios via an emailed survey (n = 42), which was followed by a consensus conference (n = 51) that reviewed the literature and the rationale for survey answers. Panelists participated in a follow-up survey for final recommendations on the scenarios (n = 45).FindingsThe panelists reached consensus (≥70% agreement) in supporting a risk-stratified approach to melanoma screening in clinical settings and public screening events, screening personnel recommendations (self/partner, primary care provider, general dermatologist, and pigmented lesion expert), screening intervals, and acceptable appointment wait times. Participants also reached consensus that visual and dermoscopic examination are sufficient for evaluation and follow-up of melanocytic skin lesions deemed innocuous. The panelists reached consensus on interpreting reflectance confocal microscopy and some but not all results from epidermal tape stripping, but they did not reach consensus on use of certain pigmented lesion evaluation tools, such as electrical impedance spectroscopy. Regarding GEP scores, the panelists reached consensus that a low-risk prognostic GEP score should not outweigh concerning histologic features when selecting patients to undergo sentinel lymph node biopsy but did not reach consensus on imaging recommendations in the setting of a high-risk prognostic GEP score and low-risk histology and/or negative nodal status.Conclusions and RelevanceFor this consensus statement, panelists reached consensus on aspects of a risk-stratified approach to melanoma screening and follow-up as well as use of visual examination and dermoscopy. These findings support a practical approach to diagnosing and evaluating CM. Panelists did not reach consensus on a clearly defined role for GEP testing in clinical decision-making, citing the need for additional studies to establish the clinical use of existing GEP assays.

Managing the balance between accurately identifying early stage melanomas while avoiding obtaining biopsy specimens of benign lesions (ie, overbiopsy) is the major challenge of melanoma detection. Decision making can be especially difficult in patients with extensive atypical nevi. Recognizing that the primary screening modality for melanoma is subjective examination, studies have shown a tendency toward overbiopsy. Even low-risk routine surgical procedures are associated with morbidity, mounting health care costs, and patient anxiety. Recent advancements in noninvasive diagnostic modalities have helped improve diagnostic accuracy, especially when managing melanocytic lesions of uncertain diagnosis. Breakthroughs in artificial intelligence have also shown exciting potential in changing the landscape of melanoma detection. In the first article in this continuing medical education series, we review novel diagnostic technologies, such as automated 2- and 3-dimensional total body imaging with sequential digital dermoscopic imaging, reflectance confocal microscopy, and electrical impedance spectroscopy, and we explore the logistics and implications of potentially integrating artificial intelligence into existing melanoma management paradigms.

Survival outcomes in melanoma and their association with mutations in the telomerase reverse transcriptase gene TERT promoter remain uncertain. In addition, few studies have examined whether these associations are affected by a nearby common germline polymorphism or vary on the basis of melanoma histopathological subtype. We analyzed 408 primary tumors from a prospective melanoma cohort for somatic TERT

Detecting mutations in the plasma of patients with solid tumors is becoming a valuable method of diagnosing and monitoring cancer. The TERT promoter is mutated at high frequencies in multiple cancer types, most commonly at positions -124 and -146 (designated C228T and C250T, respectively). Detection of these mutations has been challenging because of the high GC content of this region (approximately 80%). We describe development of novel probe-based droplet digital PCR assays that specifically detect and quantify these two mutations, along with the less common 242-243 CC>TT mutation, and demonstrate their application using human tumor and plasma samples from melanoma patients. Assay designs and running conditions were optimized using cancer cell line genomic DNAs with the C228T or C250T mutations. The limits of detection were 0.062% and 0.051% mutant allele fraction for the C228T and C250T assays, respectively. Concordance of 100% was observed between droplet digital PCR and sequencing-based orthogonal methods in the detection of TERT mutant DNA in 32 formalin-fixed, paraffin-embedded melanoma tumors. TERT(mutant) DNA was also identified in 21 of 27 plasma samples (78%) from patients with TERT(mutant) tumors, with plasma mutant allele fractions ranging from 0.06% to 15.3%. There were no false positives in plasma. These data demonstrate the potential of these assays to specifically detect and quantify TERT(mutant) DNA in tumors and plasma of cancer patients.

Background: Analysis of cell-free circulating tumor DNA (ctDNA) from blood provides an opportunity to detect potential mechanisms of resistance to targeted therapies in melanoma patients. Approximately 20% and 17% of patients undergoing BRAF targeted therapy develop resistance associated with copy number increases in either BRAF or cylinD1 (CCND1), respectively. Detection of BRAF or CCND1 copy number gains prior to radiographic tumor progression could potentially prompt a therapeutic switch aimed at treating patients with smaller tumors burdens, which may improve survival. Conventional droplet digital PCR (ddPCR) for copy number analysis uses two probes, one target and one reference gene (2-plex). To improve the accuracy and sensitivity of the copy number analysis and account for possible genetic alterations in the reference gene, we performed a 5-probe multiplex (5-plex) reaction using three reference genes and two targets. Methods: We used cell lines with known copy number gains for the BRAF gene (M395-R) and CCND1 gene (SkMel-187) to identify copy number increases using ddPCR. To estimate the limit of detection of the copy number gains in plasma, high and low concentrations of the cell line DNAs were spiked into healthy plasma and divided into three replicate samples of 3ml each. DNA was extracted from plasma using the Circulating DSP NA Kit (Qiagen). The ddPCR reaction mix used 4X Supermix and five different copy number specific probes targeting: two genes of interest, BRAF (7q34) and CCND1 (11q13.3); and three control genes: FOXI3 (2p11.2), AGO1 (1p34.3) and TTC5 (14q11.2). Each probe was titrated to display a unique cluster location on the 2D plot. Probe concentrations were: BRAF: FAM 2X; CCND1: FAM 1X; FOXI3: HEX 2X; AGO1: HEX 1X; TTC5: FAM 0.7X/HEX 0.4X. Each extraction was divided into 4 replicate wells. Following droplet generation, gene copies were analyzed using the QX Manager 1.2 Standard Edition for advanced analysis. Results: Probe titration resulted in 32 quantifiably distinct ddPCR clusters on a single 2D plot. Copies/ul of the reference genes were used as a benchmark for diploid status. We determined that the M395-R cell line had 45 copies of the BRAF gene and that SKMel-187 had eight copies of the CCND1 gene compared to diploid PBMC DNA (Promega). Based on the spike-in experiments, we estimate that we can detect copy number gains in plasmas with 10% or 20% tumor fraction when tumor cells have approximately 45 or 8 copies of the target gene, respectively. Conclusion: We successfully multiplexed five copy number assays in a single ddPCR reaction, and showed that copy number changes could be identifiable with as little as 10% tumor fraction within plasma when the tumor-associated copy number gains were relatively high. Overall, the 5-plex assay has the potential to be excellent tool for copy number detection in blood, and can be customized as needed for other targets. Citation Format: Jennifer M. Wiggins, Mahrukh Syeda, David Polsky. Development of a novel 5-plex copy number ddPCR assay for ctDNA analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 539.

A diagnosis of late-stage melanoma is associated with significant mortality. From a public health perspective, the knowledge of geographic disparities in late-stage diagnoses can inform efforts to facilitate the diagnosis of the earlier stage, highly curable melanomas. We conducted a county-level analysis of melanoma in New York state to identify communities that may benefit from pilot health interventions to reduce the burden of late-stage melanoma. From 1995 to 2016, late-stage melanoma incidence increased from 1.5 to 2.8 cases per 100,000 in New York state. We found statistically significant associations between decreased county-level health system access (including physician density and resident educational status) and increased county incidence and proportion of late-stage disease among diagnosed cases (P0.001 for both). Increased county-level socioeconomic status, including measures of resident wealth and medical insurance status, was positively associated with greater late-stage incidence (P0.001). However, decreased county-level socioeconomic status was positively associated with a greater proportion of late-stage disease among cases at diagnosis (P = 0.009). Counties with reduced access to physician services and lower socioeconomic status may be suitable for pilot interventions promoting the recognition and diagnosis of early-stage melanomas to reduce late-stage diagnoses and associated mortality.

IMPORTANCE: Use of prognostic gene expression profile (GEP) testing in cutaneous melanoma (CM) is rising despite a lack of endorsement as standard of care. OBJECTIVE: To develop guidelines within the national Melanoma Prevention Working Group (MPWG) on integration of GEP testing into the management of patients with CM, including (1) review of published data using GEP tests, (2) definition of acceptable performance criteria, (3) current recommendations for use of GEP testing in clinical practice, and (4) considerations for future studies. EVIDENCE REVIEW: The MPWG members and other international melanoma specialists participated in 2 online surveys and then convened a summit meeting. Published data and meeting abstracts from 2015 to 2019 were reviewed. FINDINGS: The MPWG members are optimistic about the future use of prognostic GEP testing to improve risk stratification and enhance clinical decision-making but acknowledge that current utility is limited by test performance in patients with stage I disease. Published studies of GEP testing have not evaluated results in the context of all relevant clinicopathologic factors or as predictors of regional nodal metastasis to replace sentinel lymph node biopsy (SLNB). The performance of GEP tests has generally been reported for small groups of patients representing particular tumor stages or in aggregate form, such that stage-specific performance cannot be ascertained, and without survival outcomes compared with data from the American Joint Committee on Cancer 8th edition melanoma staging system international database. There are significant challenges to performing clinical trials incorporating GEP testing with SLNB and adjuvant therapy. The MPWG members favor conducting retrospective studies that evaluate multiple GEP testing platforms on fully annotated archived samples before embarking on costly prospective studies and recommend avoiding routine use of GEP testing to direct patient management until prospective studies support their clinical utility. CONCLUSIONS AND RELEVANCE: More evidence is needed to support using GEP testing to inform recommendations regarding SLNB, intensity of follow-up or imaging surveillance, and postoperative adjuvant therapy. The MPWG recommends further research to assess the validity and clinical applicability of existing and emerging GEP tests. Decisions on performing GEP testing and patient management based on these results should only be made in the context of discussion of testing limitations with the patient or within a multidisciplinary group.

Objectives. To determine the effect of new therapies and trends toward reduced mortality rates of melanoma. Methods. We reviewed melanoma incidence and mortality among Whites (the group most affected by melanoma) in 9 US Surveillance, Epidemiology, and End Results registry areas that recorded data between 1986 and 2016. Results. From 1986 to 2013, overall mortality rates increased by 7.5%. Beginning in 2011, the US Food and Drug Administration approved 10 new treatments for metastatic melanoma. From 2013 to 2016, overall mortality decreased by 17.9% (annual percent change [APC] = −6.2%; 95% confidence interval [CI] = −8.7%, −3.7%) with sharp declines among men aged 50 years or older (APC = −8.3%; 95% CI = −12.2%, −4.1%) starting in 2014. This recent, multiyear decline is the largest and most sustained improvement in melanoma mortality ever observed and is unprecedented in cancer medicine. Conclusions. The introduction of new therapies for metastatic melanoma was associated with a significant reduction in population-level mortality. Future research should focus on developing even more effective treatments, identifying biomarkers to select patients most likely to benefit, and renewing emphasis on public health approaches to reduce the number of patients with advanced disease.

BACKGROUND: Little is known on whether melanocortin 1 receptor (MC1R) associated cutaneous melanoma (CM) risk varies depending on histological subtype and body site, and whether tumour thickness at diagnosis (the most important prognostic factor for CM patients) differs between MC1R variant carriers and wild-type individuals. OBJECTIVE: We studied the association between MC1R variants and CM risk by histological subtype, body site, and Breslow thickness, using the database of the M-SKIP project. METHODS: We pooled individual data from fifteen case-control studies conducted during 2005–2015 in Europe and the USA. Study-specific, multi-adjusted odds ratios were pooled into summary odds ratios (SOR) and 95% confidence intervals (CI) using random-effects models. RESULTS: 6891 CM cases and 5555 controls were included. CM risk was increased among MC1R variant carriers vs. wild-type individuals. The increase in risk was comparable across histological subtypes (SOR for any variant vs. wild-type ranged between 1.57 and 1.70, always statistical significant) except acral lentiginous melanoma, for which no association emerged; and slightly greater on chronically (1.74, 95% CI 1.47–2.07) than intermittently (1.55, 95% CI 1.34–1.78) sun-exposed skin. CM risk was greater for those carrying ‘R’ vs. ‘r’ variants; correlated with the number of variants; and was more evident among individuals not showing the red hair colour phenotype. Breslow thickness was not associated with MC1R status. CONCLUSION: MC1R variants were associated with an increased risk of CM of any histological subtype (except ALM) and occurring on both chronically and intermittently sun-exposed skin.

This article, co-authored by a patient diagnosed with acral melanoma, discusses the patient’s experience of being diagnosed with and treated with surgery for this disease. The physician discusses the epidemiology, genetics, diagnosis, treatment, and prognosis of acral melanoma. Follow-up care plans are also discussed.

Melanoma represents an important public health problem, due to its high case-fatality rate. Identification of individuals at high risk would be of major interest to improve early diagnosis and ultimately survival. The aim of this study was to evaluate whether MC1R variants predicted melanoma risk independently of at-risk phenotypic characteristics.Data were collected within an international collaboration - the M-SKIP project. The present pooled analysis included data on 3,830 single, primary, sporadic, cutaneous melanoma cases and 2,619 controls from seven previously published case-control studies. All the studies had information on MC1R gene variants by sequencing analysis and on hair color, skin phototype, and freckles, ie, the phenotypic characteristics used to define the red hair phenotype.The presence of any MC1R variant was associated with melanoma risk independently of phenotypic characteristics (OR 1.60; 95% CI 1.36-1.88). Inclusion of MC1R variants in a risk prediction model increased melanoma predictive accuracy (area under the receiver-operating characteristic curve) by 0.7% over a base clinical model (P=0.002), and 24% of participants were better assessed (net reclassification index 95% CI 20%-30%). Subgroup analysis suggested a possibly stronger role of MC1R in melanoma prediction for participants without the red hair phenotype (net reclassification index: 28%) compared to paler skinned participants (15%).The authors suggest that measuring the MC1R genotype might result in a benefit for melanoma prediction. The results could be a valid starting point to guide the development of scientific protocols assessing melanoma risk prediction tools incorporating the MC1R genotype.

Background Complete removal of individual dysplastic nevi (DN) is often accomplished by a second surgical procedure after the initial biopsy. The choice to perform the second procedure is strongly influenced by histopathologic margin status of the initial biopsy specimen. Objective To evaluate the clinical and histopathologic outcomes of in toto biopsy of DN using a predetermined margin of normal skin. Methods We conducted a prospective study of a saucerization method using a defined 2-mm margin in patients undergoing biopsy of a pigmented skin lesion. Results We performed 151 biopsies in 138 patients. Overall, 137 of 151 lesions subjected to biopsy (90.7%) were melanocytic: 86 DN (57.0%), 40 nevi without atypia (26.5%), and 11 melanomas (7.3%). Of 78 DN, 68 (87.2%) were removed with clear histopathologic margins (8 DN were excluded because of inadequate processing). There was no clinical evidence of recurrence at any of the biopsy sites that were simply observed (i.e., not re-excised) over a median of 16.9 months. Limitations There were few biopsies performed on the face. Conclusions The complete histopathologic removal of nearly 9 of 10 DN using a peripheral margin of 2 mm of normal skin and a depth at the dermis and subcutaneous fat junction has the potential to decrease second procedures at DN biopsy sites, thereby decreasing patient morbidity and saving health care dollars.

Identifying new or changing melanocytic lesions, particularly in patients with numerous or atypical nevi, can be challenging. Total-body photography and sequential digital dermoscopy imaging, together known as digital follow-up, are 2 prominent forms of noninvasive imaging technology used in mole mapping that have been found to improve diagnostic accuracy, detect earlier-stage melanomas, and reduce costs. Digital follow-up, in combination with direct-to-consumer applications and teledermatology, is already revolutionizing the ways in which physicians and patients participate in melanoma surveillance and will likely continue to enhance early detection efforts.

Background The American Joint Committee on Cancer (AJCC) maintains that the eighth edition of its Staging Manual (AJCC8) has improved accuracy compared with the seventh (AJCC7). However, there are concerns that implementation may disrupt analysis of active clinical trials for stage III patients. We used an independent cohort of melanoma patients to test the extent to which AJCC8 has improved prognostic accuracy compared with AJCC7. Methods We analyzed a cohort of 1315 prospectively enrolled patients. We assessed primary tumor and nodal classification of stage I–III patients using AJCC7 and AJCC8 to assign disease stages at diagnosis. We compared recurrence-free (RFS) and overall survival (OS) using Kaplan-Meier curves and log-rank tests. We then compared concordance indices of discriminatory prognostic ability and area under the curve of 5-year survival to predict RFS and OS. All statistical tests were two-sided. Results Stage IIC patients continued to have worse outcomes than stage IIIA patients, with a 5-year RFS of 26.5% (95% confidence interval [CI] = 12.8% to 55.1%) vs 56.0% (95% CI = 37.0% to 84.7%) by AJCC8 (P = .002). For stage I, removing mitotic index as a T classification factor decreased its prognostic value, although not statistically significantly (RFS concordance index [C-index] = 0.63, 95% CI = 0.56 to 0.69; to 0.56, 95% CI = 0.49 to 0.63, P = .07; OS C-index = 0.48, 95% CI = 0.38 to 0.58; to 0.48, 95% CI = 0.41 to 0.56, P = .90). For stage II, prognostication remained constant (RFS C-index = 0.65, 95% CI = 0.57 to 0.72; OS C-index = 0.61, 95% CI = 0.50 to 0.72), and for stage III, AJCC8 yielded statistically significantly enhanced prognostication for RFS (C-index = 0.65, 95% CI = 0.60 to 0.70; to 0.70, 95% CI = 0.66 to 0.75, P = .01). Conclusions Compared with AJCC7, we demonstrate that AJCC8 enables more accurate prognosis for patients with stage III melanoma. Restaging a large cohort of patients can enhance the analysis of active clinical trials.

Mutational heterogeneity can contribute to therapeutic resistance in solid cancers. In melanoma, the frequencies of intertumoral and intratumoral heterogeneity are controversial. We examined mutational heterogeneity within individual patients with melanoma using multiplatform analysis of commonly mutated driver and nonpassenger genes. We analyzed paired primary and metastatic tumors from 60 patients and multiple metastatic tumors from 39 patients whose primary tumors were unavailable (n = 271 tumors). We used a combination of multiplex SNaPshot assays, Sanger sequencing, mutation-specific PCR, or droplet digital PCR to determine the presence of BRAFV600, NRASQ61, TERT–124C>T, and TERT–146C>T mutations. Mutations were detected in BRAF (39%), NRAS (21%), and/or TERT (78%). Thirteen patients had TERTmutant discordant tumors; seven of these had a single tumor with both TERT–124C>T and TERT–146C>T mutations present at different allele frequencies. Two patients had both BRAF and NRAS mutations; one had different tumors and the other had a single tumor with both mutations. One patient with a BRAFmutant primary lacked mutant BRAF in at least one of their metastases. Overall, we identified mutational heterogeneity in 18 of 99 patients (18%). These results suggest that some primary melanomas may be composed of subclones with differing mutational profiles. Such heterogeneity may be relevant to treatment responses and survival outcomes.

The detection of cell-free, circulating tumor DNA (ctDNA) in the blood of patients with solid tumors is often referred to as "liquid biopsy." ctDNA is particularly attractive as a candidate biomarker in the blood. It is relatively stable after blood collection, can be easily purified, and can be quantitatively measured with high sensitivity and specificity using advanced technologies. Current liquid biopsy research has focused on detecting and quantifying ctDNA to (1) diagnose and characterize mutations in a patient's cancer to help select the appropriate treatment; (2) predict clinical outcomes associated with different treatments; and (3) monitor the response and/or progression of a patient's disease. The diagnostic use of liquid biopsies is probably greatest in tumors where the difficulty and/or risk of obtaining a tissue specimen for molecular diagnostics is high (e.g., lung, colon). In metastatic melanoma, however, obtaining a tissue sample for molecular diagnostics is not typically a major obstacle to patient care plans; rather predicting treatment outcomes and monitoring a patient's disease course during therapy are considered the current priorities for this cancer type. In this chapter we describe an approach to the validation of ctDNA detection assays for melanoma, focusing primarily on analytical validation, and provide methods to guide the use of droplet digital PCR assays for measuring ctDNA levels in plasma samples.

The detection of cell-free, circulating tumor DNA (ctDNA) in the blood of patients with solid tumors is often referred to as "liquid biopsy." ctDNA is particularly attractive as a candidate biomarker in the blood. It is relatively stable after blood collection, can be easily purified, and can be quantitatively measured with high sensitivity and specificity using advanced technologies. Current liquid biopsy research has focused on detecting and quantifying ctDNA to (1) diagnose and characterize mutations in a patient's cancer to help select the appropriate treatment; (2) predict clinical outcomes associated with different treatments; and (3) monitor the response and/or progression of a patient's disease. The diagnostic use of liquid biopsies is probably greatest in tumors where the difficulty and/or risk of obtaining a tissue specimen for molecular diagnostics is high (e.g., lung, colon). In metastatic melanoma, however, obtaining a tissue sample for molecular diagnostics is not typically a major obstacle to patient care plans; rather predicting treatment outcomes and monitoring a patient's disease course during therapy are considered the current priorities for this cancer type. In this chapter we describe an approach to the validation of ctDNA detection assays for melanoma, focusing primarily on analytical validation, and provide methods to guide the use of droplet digital PCR assays for measuring ctDNA levels in plasma samples.

Managing the balance between accurately identifying early stage melanomas while avoiding obtaining biopsy specimens of benign lesions (ie, overbiopsy) is the major challenge of melanoma detection. Decision making can be especially difficult in patients with extensive atypical nevi. Recognizing that the primary screening modality for melanoma is subjective examination, studies have shown a tendency toward overbiopsy. Even low-risk routine surgical procedures are associated with morbidity, mounting health care costs, and patient anxiety. Recent advancements in noninvasive diagnostic modalities have helped improve diagnostic accuracy, especially when managing melanocytic lesions of uncertain diagnosis. Breakthroughs in artificial intelligence have also shown exciting potential in changing the landscape of melanoma detection. In the first article in this continuing medical education series, we review novel diagnostic technologies, such as automated 2- and 3-dimensional total body imaging with sequential digital dermoscopic imaging, reflectance confocal microscopy, and electrical impedance spectroscopy, and we explore the logistics and implications of potentially integrating artificial intelligence into existing melanoma management paradigms.

The growth of molecular technologies analyzing skin cells and inherited genetic variations has the potential to address current gaps in both diagnostic accuracy and prognostication in patients with melanoma or in individuals who are at risk for developing melanoma. In the second article in this continuing medical education series, novel molecular technologies are reviewed. These have been developed as adjunct tools for melanoma management and include the Pigmented Lesion Assay, myPath Melanoma, and DecisionDx-Melanoma tests, and genetic testing in patients with a strong familial melanoma history. These tests are commercially available and marketed as ancillary tools for clinical decision-making, diagnosis, and prognosis. We review fundamental principles behind each test, discuss peer-reviewed literature assessing their performance, and highlight the utility and limitations of each assay. The goal of this article is to provide a comprehensive, evidence-based foundation for clinicians regarding the management of patients with difficult pigmented lesions.

BACKGROUND: Patients diagnosed with metastatic melanoma have varied clinical courses, even in patients with similar disease characteristics. We examine the impact of initial stage of melanoma diagnosis, BRAF status of primary melanoma, and receiving adjuvant therapy on post metastatic survival. METHODS: We studied melanoma patients presenting to Perlmutter Cancer Center at New York University (NYU) and prospectively enrolled in NYU’s melanoma biospecimen database and followed up on protocol-driven schedule. Patients were stratified by stage at initial melanoma diagnosis as per AJCC 7(th) edition guidelines. Post-metastatic survival was determined using the Kaplan-Meier method, and Cox proportional hazards models were used to assess hazard ratios RESULTS: Three hundred and four out of 3204 patients developed metastatic disease over the time of follow up (median follow up 2.2 years, range 0.08–35.2 years). Patients diagnosed with stage I (n=96) melanoma had longer pmOS (29.5 months) than those diagnosed with stage II (n=99, pmOS 14.9 months) or stage III (n=109, pmOS 15.1 months) melanoma (p=0.036). Initial stage of diagnosis remained significant in multivariate analysis when controlling for LDH and site of metastases (primary diagnosis stage II (HR 1.44, p=0.046), stage III (HR 1.5, p=0.019)). Adjuvant treatment was associated with better survival but BRAF mutation status did not show an association. CONCLUSION: Our data challenge the general assumption that primary melanomas converge upon diagnosis of metastatic disease and behave uniformly. Primary stage of melanoma at the time of diagnosis may be prognostic of outcome, similar to LDH and metastatic disease sites.