The Clinical, Morphologic, and Molecular Spectrum of BRAF Fusion Spitz Tumors.

BRAF fusion Spitz neoplasms along with MAP3K8 fusions are among the subtypes of Spitz most likely to be diagnosed as Spitz melanoma. However, the current literature has only limited amounts of clinical follow-up on these cases. In this study, we share our experience with 39 BRAF fusion Spitz neoplasms and provide the greatest number of cases with available clinical follow-up. Among 24 patients with clinical follow-up (mean duration of 26.1 mo), none developed metastatic disease. Detailed biomarker assessment with FISH studies, TERT promoter mutational analysis, PRAME and p16 IHC also strongly favored a benign process. Only 2 of 17 cases were positive by FISH, 37 of 38 were negative for TERT promoter mutations, 24 of 24 were negative for PRAME and 16 of 21 had retained staining with p16. In addition, we identify and describe several distinct morphologic patterns, some of which are highly spitzoid in cytomorphology while others lack convincingly spitzoid cytomorphology. We address classification of those cases with less than classic spitzoid cytomorphology with a nonsupervised PCA plot which shows that independent of how spitzoid the cytomorphology is, BRAF fusions strongly cluster with other subtypes of Spitz neoplasms. In conclusion, we show with clinical follow-up, a meta-analysis of the current literature, and our biomarker analysis, that most BRAF fusion Spitz neoplasms have an indolent course and should be considered either benign or of intermediate grade. Further, our morphologic assessment and PCA plot suggest the morphologic spectrum of Spitz neoplasms may need to be expanded.

PRKC Fusion Melanocytic Tumors, a Subgroup of Melanocytic Tumors More Closely Aligned to Blue Nevi Than to PRKAR1A-inactivated Pigmented Epithelioid Melanocytomas.

Tumors morphologically classified as pigmented epithelioid melanocytomas (PEMs) are genomically diverse, with the 2 most common genomic subtypes being PRKC fusions or PRKAR1A inactivating mutations. PRKC fusions activate the Gα q/11 pathway similar to blue nevi. Conversely, inactivating mutations in PRKAR1A activate the Gα s pathway. We hypothesize that PRKC fusions have greater genomic overlap with blue nevi compared with PRKAR1A-inactivated PEMs. We characterized the clinical and morphologic features of 21 PRKC and PRKACB fusion melanocytic tumors and compared this to PRKAR1A mutated PEMs. To test our hypothesis regarding greater genomic overlap between PRKC fusions and blue nevi relative to PRKAR1A mutated PEMs, we performed a principal component analysis (PCA) using mRNA expression data. Lastly, we performed a meta-analysis focusing on the outcome data of PRKC fusions. PRKC fusions occur at a younger median age than PRKAR1A mutated PEMs (16 vs. 27). Histologically, PRKC fusions have solid aggregates of epithelioid melanocytes not typical of PRKAR1A mutated PEMs. The PCA plot showed no overlap between the PRKC fusion group and the PRKAR1A-mutated PEMs. There was a significant overlap between PRKC fusions and blue nevi. A meta-analysis of PRKC fusion cases in the literature suggests melanoma is uncommon, but the loss of BAP-1 nuclear expression may be associated with an adverse prognosis as in tumors from the blue nevus family. PRKC fusion melanocytic tumors have greater genomic overlap with blue nevi compared with PRKAR1A mutated PEMs. We recommend categorizing benign PRKC fusion melanocytic tumors as blue fusion nevi/tumors.

Molecular effects of indoor tanning.

Background: Tanning bed users have a significantly increased risk of melanoma, but it remains unclear how indoor tanning drives melanomagenesis. Tanning bed radiation is often thought of as a substitute for natural UV radiation despite differences in the maximum doses, UV content, body sites exposed, and patterns of melanoma that arise. Methods: To better understand the epidemiologic trends and etiology of melanoma associated with tanning bed use, we described the patterns of melanoma in patients with quantifiable tanning bed usage and performed exome sequencing of 182 melanocytes from normal skin of a subset of these patients. Results: Tanning bed users were more likely than non-users to have melanoma on body sites with low cumulative levels of sun damage and were more likely to have multiple melanomas. The melanocytes in normal appearing skin from tanning bed users had higher mutation burdens, a higher proportion of melanocytes with pathogenic mutations, and distinct mutational signatures. These differences were most prominent over body sites that experience comparatively less exposure to natural sunlight. Conclusions: We conclude that tanning bed radiation induces melanoma by increasing the mutation burden of melanocytes and by mutagenizing a broader field of melanocytes than are typically exposed to natural sunlight. The unique signatures of mutations in skin cells of tanning users may be attributable to the distinct spectra of radiation emitted from solariums.

The Impact of Next-generation Sequencing on Interobserver Agreement and Diagnostic Accuracy of Desmoplastic Melanocytic Neoplasms.

Next-generation sequencing (NGS) is increasingly being utilized as an ancillary tool for diagnostically challenging melanocytic neoplasms. It is incumbent upon the pathology community to perform studies assessing the benefits and limitations of these tools in specific diagnostic scenarios. One of the most challenging diagnostic scenarios faced by skin pathologists involves accurate diagnosis of desmoplastic melanocytic neoplasms (DMNs). In this study, 20 expert melanoma pathologists rendered a diagnosis on 47 DMNs based on hematoxylin and eosin sections with demographic information. After submitting their diagnosis, the experts were given the same cases, but this time with comprehensive genomic sequencing results, and asked to render a diagnosis again. Identification of desmoplastic melanoma (DM) improved by 7%, and this difference was statistically significant ( P <0.05). In addition, among the 15 melanoma cases, in the pregenomic assessment, only 12 were favored to be DM by the experts, while after genomics, this improved to 14 of the cases being favored to be DM. In fact, some cases resulting in metastatic disease had a substantial increase in the number of experts recognizing them as DM after genomics. The impact of the genomic findings was less dramatic among benign and intermediate-grade desmoplastic tumors (BIDTs). Interobserver agreement also improved, with the Fleiss multirater Kappa being 0.36 before genomics to 0.4 after genomics. NGS has the potential to improve diagnostic accuracy in the assessment of desmoplastic melanocytic tumors. The degree of improvement will be most substantial among pathologists with some background and experience in bioinformatics and melanoma genetics.

BRAF Mutated and Morphologically Spitzoid Tumors, a Subgroup of Melanocytic Neoplasms Difficult to Distinguish From True Spitz Neoplasms.

Drivers of Spitz neoplasms include activating point mutations in HRAS and Spitz-associated genomic fusions. It has become evident that some BRAF -mutated melanocytic neoplasms can morphologically mimic Spitz tumors (STs). These have been termed BRAF mutated and morphologically spitzoid (BAMS). In this study, 17 experts from the International Melanoma Pathology Study Group assessed 54 cases which included 40 BAMS and 14 true STs. The participants reviewed the cases blinded to the genomic data and selected among several diagnostic options, including BAMS, ST, melanoma, and other. A total of 38% of all diagnostic selections in the BAMS cases were for BAMS, whereas 32% were for ST. In 22 of the BAMS cases, the favored diagnosis was BAMS, whereas in 17 of the BAMS cases, the favored diagnosis was ST. Among the 20 cases in the total group of 54 with the highest number of votes for ST, half were BAMS. Of BAMS, 75% had a number of votes for ST that was within the SD of votes for ST seen among true ST cases. There was poor interobserver agreement for the precise diagnosis of the BAMS (kappa = 0.16) but good agreement that these cases were not melanoma (kappa = 0.7). BAMS nevi/tumors can closely mimic Spitz neoplasms. Expert melanoma pathologists in this study favored a diagnosis of ST in nearly half of the BAMS cases. There are BAMS cases that even experts cannot morphologically distinguish from true Spitz neoplasms.

Clinical, Morphologic, and Molecular Features of MAP3K8 Rearranged Spitz Neoplasms: A Retrospective Study Documenting That Bonafide Spitz Melanomas Are Rare.

Previous studies regarding the clinical behavior of Spitz neoplasms lack genomic characterization. We aim to assess our hypothesis that most MAP3K8 Spitz neoplasms are indolent despite MAP3K8 being the single most common driver of Spitz melanoma. Further, we aim to identify genomic features associated with aggressive behavior and to better characterize the morphology of these cases. We analyzed the outcomes of MAP3K8 Spitz neoplasms. We also performed a meta-analysis of the outcomes of MAP3K8 Spitz from the literature. Morphologic features were compared with other variants of Spitz using a Student t test and χ 2 test. Two of 35 cases resulted in local recurrence and one of these cases had local regional metastasis; all other cases had no evidence of recurrence (mean follow-up time: 33 mo). MAP3K8 Spitz only rarely results in aggressive behavior. Metastatic cases have genomic mutations associated with tumor progression. Morphologically, MAP3K8 Spitz neoplasms frequently showed nodular silhouette, large cell size, epithelioid morphology, and severe nuclear atypia resulting in more frequent diagnosis as Spitz melanoma. Most MAP3K8 Spitz neoplasms have excellent prognoses, apart from rare cases harboring additional genomic abnormalities associated with tumor progression.

Spitz tumor with RAF1 fusion: A report of 3 cases.

Spitz tumors are melanocytic neoplasms morphologically characterized by spindled and/or epithelioid cells and specific stromal and epidermal changes associated with mutually exclusive fusion kinases involving ALK, ROS1, NTRK1, NTRK2, NTRK3, MET and RET, BRAF and MAP3K8 genes or, less commonly, HRAS mutation. RAF1 fusions have been recently detected in cutaneous melanocytic neoplasms, including conventional melanoma, congenital nevus and BAP-1 inactivated tumors. We report herewith three Spitz neoplasms with a RAF1 fusion, including a previously reported CTDSPL::RAF1 fusion and two novel PPAP2B::RAF1 and ATP2B4::RAF1 fusions. Two cases were classified as Spitz nevus, while the remaining neoplasm was classified as Spitz melanoma at the time of the diagnosis, given 9p21 homozygous deletion and positive sentinel lymph node biopsy. We suggest that RAF1 fused melanocytic neoplasms can represent a novel subgroup of Spitz tumors, with a RAF1 fusion representing an oncogenic driver.

Clinical, Morphologic, and Molecular Features of Benign and Intermediate-grade Melanocytic Tumors With Activating Mutations in MAP2K1.

Activating mutations in MAP2K1 can be seen in benign and intermediate-grade melanocytic neoplasms with spitzoid morphology. We analyzed the clinical, histopathologic, and genetic features for 16 cases of benign and intermediate-grade melanocytic tumors harboring activating MAP2K1 mutations. We compared them to Spitz neoplasms with characteristic Spitz fusions or HRAS mutation. We also compared the mutational pattern of benign and intermediate-grade MAP2K1 -mutated neoplasms and melanomas with activating MAP2K1 mutations. Among the 16 cases, the favored morphologic diagnosis was Spitz nevus (8/16), atypical Spitz tumors (6/16), and deep penetrating nevus (2/16). The 2 most common architectural patterns seen included a plaque-like silhouette with fibroplasia around the rete reminiscent of a dysplastic nevus (n=7) or a wedge-shaped or nodular pattern with the plexiform arrangement of the nests aggregating around the adnexa or neurovascular bundle (n=8). The cases with dysplastic architecture and spitzoid cytology resembled dysplastic Spitz nevi. Compared with true Spitz neoplasms, MAP2K1 -mutated neoplasms occurred in older age groups and had more frequent pagetosis and a lower average mitotic count. The most common type of mutation in the benign and intermediate-grade cases in the literature involves an in-frame deletion, while, in melanomas, missense mutations are predominant. Benign and intermediate-grade melanocytic neoplasms with activating mutations in MAP2K1 can have morphologic overlap with Spitz neoplasms. A significant proportion of melanomas also have activating MAP2K1 mutations. In-frame deletions are predominantly seen in the benign and intermediate-grade cases, and missense mutations are predominantly seen in melanomas.

A reappraisal of the epidemiology of Spitz neoplasms in the molecular era: A retrospective cohort study.

BACKGROUND: Previous studies suggest that Spitz neoplasms occur primarily in younger patients, leading pathologists to shy away from diagnosing a benign Spitz neoplasm in the elderly. With the advent of genomic sequencing, there is a need for reappraisal of the epidemiology of Spitz neoplasms in the modern molecular era. OBJECTIVE: We aim to reassess the epidemiology of Spitz neoplasms incorporating next-generation sequencing. METHODS: We looked at 53,814 non-Spitz neoplasms and 1260 Spitz neoplasms including 286 Spitz neoplasms with next-generation sequencing testing and collected various epidemiologic data. RESULTS: In our general pool of cases, the proportion of Spitz neoplasm cases occurring is relatively the same in each of the first 4 decades of life with a precipitous drop in the fifth decade. In assessing a group of genomically verified cases of Spitz neoplasms, the drop was much less significant and up to 20% of all Spitz neoplasm cases occurred in patients over 50 years of age. LIMITATIONS: Limitations included the number of genetically verified Spitz neoplasm cases available and a possible bias as to which cases undergo genomic testing. CONCLUSION: Genomic verification may allow more confident diagnosis of Spitz neoplasms in patients over 50 years of age and avoid melanoma overdiagnosis.

NONO::TFE3 fusion cutaneous epithelioid and spindle cell tumor: A case series.

The NONO::TFE3 fusion has been described in MiT family translocation renal cell carcinomas as well as extracutaneous perivascular epithelioid cell tumors (PEComas). PEComas are known to express myogenic and melanocytic markers but SOX10 and p63 positivity has never been reported. We report two primary cutaneous tumors that morphologically and molecularly fit PEComas, both harboring the NONO::TFE3 fusion, but with an unusual immunophenotype of SOX10 and p63 positivity. One case was on an 80-year-old male's finger, and the other one was on a 72-year-old female's thigh. Both were well-circumscribed multinodular dermal tumors composed of nests of monotonous epithelioid to spindled cells with pale to vacuolated cytoplasm, some of which were arranged around blood vessels. Both tumors were positive for SOX10, S100, and p63, focally positive for Melan-A, and negative for myogenic markers. There are very little data regarding the molecular findings of primary cutaneous PEComas. While the NONO::TFE3 fusion has been identified in extracutaneous PEComas, it has never been reported in primary cutaneous cases. We believe these cases represent a previously undescribed subtype of cutaneous tumor which shows some immunophenotypic expression of melanocytic markers and we named these cases NONO::TFE3 fusion cutaneous epithelioid and spindle cell tumor. Further, we raise the question of whether this tumor should fall under the rubric of PEComa because of its morphology, partial expression of melanocytic markers, and the presence of the NONO::TFE3 fusion, or whether these tumors represent a separate novel class of tumors since the immunophenotypic expression of SOX10 and p63 is unusual for PEComas.

Agminated presentation of fusion-driven melanocytic neoplasms.

BACKGROUND: The conventionally understood pathogenesis of agminated Spitz nevi includes a mosaic HRAS mutation followed by copy number gains in 11p. However, we have recently observed agminated presentations of fusion-driven melanocytic neoplasms. METHODS: We retrieved cases from our database of benign fusion-induced melanocytic neoplasms with an agminated presentation. Both the primary lesion and the secondary lesion were sequenced. TERT-promoter mutational testing and the melanoma fluorescence in situ hybridization assay were also performed. RESULTS: Three cases were included. Two had a PRKCA fusion (partners ATP2B4 and MPZL1) and one had a ZCCHC8::ROS1 fusion. None of the cases met morphologic or molecular criteria for malignancy. There was no evidence of tumor progression in secondary lesions. The same fusion was identified in the primary and secondary lesions. None of the patients developed evidence of nodal or systemic metastasis. CONCLUSIONS: We present accumulating evidence that fusion-driven melanocytic neoplasms can present with an agminated presentation. The differential diagnosis of an agminated presentation versus a locally recurrent or potentially locally metastatic tumor is critical, and accurate diagnosis has significant prognostic and therapeutic consequences for the patient. As with HRAS mutations, fusion-driven melanocytic tumors may have an agminated presentation.

PRAME Immunohistochemical Expression and TERT Promoter Mutational Analysis as Ancillary Diagnostic Tools for Differentiating Proliferative Nodules From Melanoma Arising in Congenital Nevi.

ABSTRACT: Proliferative nodules (PNs) are benign melanocytic proliferations that typically develop within congenital melanocytic nevi. These tumors have overlapping histological features with melanoma. Ancillary immunohistochemistry and genomic sequencing are often used in diagnostically challenging cases. To assess the utility of preferentially expressed antigen in melanoma (PRAME) immunoreactivity and telomerase reverse transcriptase (TERT) promoter mutation analysis in distinguishing PNs from melanoma arising in congenital nevi cases. Twenty-one PNs and 2 melanomas arising in congenital nevi were immunohistochemically stained with PRAME. Cases with adequate tissue were also assessed for TERT promoter mutations through sequencing studies. The positivity rates in the PN cases were compared with those of the melanomas. Two of 21 PN cases were diffusely positive for PRAME (≥75% of the tumor cells positive). Two of 2 melanomas arising in congenital nevus cases were also diffusely PRAME positive. The difference was statistically significant using a Fisher exact test. None of the tumors harbored TERT promoter mutations. PRAME immunohistochemical marker may have diagnostic value in distinguishing diagnostically challenging PNs from melanoma, but diffuse expression is not specific for melanoma.

Routine imaging guided by a 31-gene expression profile assay results in earlier detection of melanoma with decreased metastatic tumor burden compared to patients without surveillance imaging studies.

Patients with early-stage disease typically have a good prognosis, but still have a risk of recurrence, even with negative sentinel lymph node biopsy (SLNB). This study explores the utility of routine imaging to detect metastases in patients with negative SLNB but high-risk 31 gene expression profile (31-GEP) scores. We retrospectively identified melanoma patients with negative SLNBs. Patients with high-risk GEP results were placed in the experimental group and patients without GEP testing were placed in the control group. Among both cohorts, recurrent melanoma groups were identified. The tumor burden at the time of recurrence and the time to recurrence were compared between experimental group patients with routine imaging and control group patients without imaging schedules. We identified 327 control patients and 307 experimental patients, of which 14.1% versus 20.5% had melanoma recurrence, respectively. Of the patients with recurrent melanoma, those in the experimental group were older (65.75 versus 59.20), had higher Breslow depths (3.72 mm versus 3.31 mm), and had advanced tumor staging (89.5% versus 71.4% of patients presenting clinical stage ≥ II) compared to the control group at primary diagnosis. However, melanoma recurrence was detected earlier (25.50 months versus 35.35 months) in the experimental group at a lower overall tumor burden (73.10 mm versus 27.60 mm). A higher percentage of experimental patients started immunotherapy when offered (76.3% and 67.9%). Patients who received routine imaging after high-risk GEP test scores had an earlier recurrence diagnosis with lower tumor burden, leading to better clinical outcomes.

TERT Promoter Mutational Analysis as an Ancillary Diagnostic Tool for Diagnostically Challenging Melanocytic Neoplasms.

ABSTRACT: Telomerase reverse transcriptase promoter mutations (TPMs) have been shown to be common in melanoma and uncommon in benign nevi. To assess the use of TPMs as an ancillary diagnostic tool, we report the concordance of the TPM status with the final diagnosis in clinical cases with distinct differential diagnostic scenarios: dysplastic nevus versus melanoma, atypical Spitz nevus versus melanoma, atypical deep penetrating nevus (DPN) versus melanoma, and atypical blue nevus versus malignant blue nevus. In a control cohort, we found a positive TPM in 51/70 (73%) of the total melanomas with the highest frequency in vertical growth phase melanoma cases. Conversely, only 2/35 (6%) dysplastic nevi in our control cases were TPM-positive and b were severely atypical dysplastic nevi. Our clinical cohort of 257 cases had a positive TPM in 24% of cases diagnosed as melanoma and in 1% of cases with a benign diagnosis. The overall concordance of the TPM status with the final diagnosis was 86%. The TPM status had the greatest concordance (95%) with the final diagnosis in the atypical DPN versus melanoma group, with the rest of the groups ranging between 50% and 88%. Overall, our results suggest that TPMs are most useful in the differential diagnosis of atypical DPN versus melanoma. It also has some value in the differential diagnosis of atypical Spitz tumor versus melanoma and dysplastic nevus versus melanoma, whereas in our cohort, it did not contribute meaningfully to differentiating malignant blue nevus and atypical blue nevus.

An update on genomic aberrations in Spitz naevi and tumours.

Spitz neoplasms continue to be a diagnostic challenge for dermatopathologists and are defined by distinctive morphological and genetic features. With the recent advancements in genomic sequencing, the classification, diagnosis, and prognostication of these tumours have greatly improved. Several subtypes of Spitz neoplasms have been identified based on their specific genomic aberrations, which often correlate with distinctive morphologies and biological behaviour. These genetic driver events can be classified into four major groups, including: (1) mutations [HRAS mutations (with or without 11p amplification) and 6q23 deletions]; (2) tyrosine kinase fusions (ROS1, ALK, NTRK1-3, MET and RET); (3) serine/threonine kinase fusions and mutations (BRAF, MAP3K8, and MAP2K1); and (4) other rare genomic aberrations. These driver genomic events are hypothesised to enable the initial proliferation of melanocytes and are often accompanied by additional genomic aberrations that affect biological behaviour. The discovery of theses genomic fusions has allowed for a more objective definition of a Spitz neoplasm. Further studies have shown that the majority of morphologically Spitzoid appearing melanocytic neoplasms with aggressive behaviour are in fact BRAF or NRAS mutated tumours mimicking Spitz. Truly malignant fusion driven Spitz neoplasms may occur but are relatively uncommon, and biomarkers such as homozygous 9p21 (CDKN2A) deletions or TERT-p mutations can have some prognostic value in such cases. In this review, we discuss the importance and various methods of identifying Spitz associated genomic fusions to help provide more definitive classification. We also discuss characteristic features of the various fusion subtypes as well as prognostic biomarkers.

Next-generation Sequencing as a Potential Diagnostic Adjunct in Distinguishing Between Desmoplastic Melanocytic Neoplasms.

Desmoplastic melanomas (DMs) are often challenging to diagnose and ancillary tests, such as immunohistochemistry, have limitations. One challenge is distinguishing DM from benign desmoplastic melanocytic neoplasms. In this study, we explored the utility of next-generation sequencing data in the diagnosis of DMs versus desmoplastic Spitz nevi (DSN) and desmoplastic nevi (DN). We sequenced 47 cases and retrieved 12 additional previously sequenced clinical cases from our dermatopathology database. The 59 total cases were comprised of 21 DMs, 25 DSN, and 13 DN. The DMs had the highest tumor mutation burden at 22 mutations/megabase (m/Mb) versus the DSN (6 m/Mb) and DN (8 m/Mb). Truncating mutations in NF1 resulting in a loss-of-function were exclusive to the DM cohort, identified in 8/21 (38%) cases. Importantly, missense mutations in NF1 were nonspecific and seen with similar frequency in the different cohorts. Other mutations exclusive to the DMs included truncating mutations in TP53 , CDKN2A , and ARID2 . Among the DSN, 17/25 (68%) had an HRAS mutation or receptor tyrosine kinase fusion consistent with other Spitz tumors. Two cases in the DN cohort had missense mutations in BRAF without additional progression mutations and 2 other cases had mutations in GNAQ , supporting a diagnosis of a sclerosing blue nevus. The remainder of the DN had nonspecific mutations in various signaling pathways with few progression mutations. Overall, our study provides preliminary data that next-generation sequencing data may have the potential to serve as an ancillary diagnostic tool to help differentiate malignant and benign desmoplastic melanocytic neoplasms.

Next-generation sequencing improves agreement and accuracy in the diagnosis of Spitz and spitzoid melanocytic lesions.

BACKGROUND: Spitzoid melanocytic neoplasms can be challenging to diagnose on histopathology alone. Next-generation sequencing (NGS) offers promise as a valuable aid in the diagnosis. Recently, one study reported increased inter-rater agreement in the diagnosis of spitzoid melanocytic neoplasms among 20 expert melanoma pathologists after incorporating NGS data. We hypothesized that NGS would carry a similar utility in a broader group of dermatopathologists and general pathologists. METHODS: Sixty-three participants of a live online (www.Dermpedia.org) CME course rendered a diagnosis on 70 cases composed of melanocytic neoplasms with spitzoid features. In Survey 1, cases included H&E slides and demographic information only, while Survey 2 included NGS data. RESULTS: With NGS information, inter-rater agreement significantly improved from "fair" to "almost perfect" and from "fair" to "substantial" for categorizing lesions as Spitz versus non-Spitz and conventional melanoma versus not, respectively. There was also an increase in diagnostic accuracy, evidenced by improved recognition of three metastatic tumors as being conventional melanomas. CONCLUSION: The study supports the adoption of NGS as a valuable diagnostic adjunct for both expert and broader dermatopathologists in their assessments of spitzoid neoplasms.