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Over the past several decades, the study of Spitz neoplasms has flourished, with expanded conceptualization and refined terminology, providing a framework for the assessment and classification of Spitz nevi, atypical Spitz Tumors, and Spitz melanoma. Cancer genomics have generated concepts such as driver and passenger genes and clonal evolution, which can be applied to Spitz tumors. Herein, we provide a historical perspective, followed by a summary of current knowledge and clinical approaches for these challenging tumors.

Eruptive melanocytic nevi (EMN) describes the sudden onset of cutaneous nevi over weeks or months. Such a clinical event is generally seen in young adult patients and may be related to several possible causes. We report here a case of EMN in an old male patient followed up for a thick amelanotic cutaneous melanoma. A few months after the eruption, multiple hepatic masses, diagnosed as melanoma metastasis, were detected. The presented case may suggest that EMN may be a paraneoplastic phenomenon of alert in patients being followed for melanoma or other malignancies.

Studies on the genomic aberrations in melanocytic neoplasms have shown a complex genomic landscape. In nevi and melanomas, a MAP-kinase pathway activation was generally found, produced by different chromosomal aberrations, including BRAF, NRAS, HRAS, GNAQ, GNA11, BAP1, CTNNB1, MAP2K1, PRKAR1A, and NF1 mutations, and ALK, ROS1, NTRK1, RET, MET, BRAF, NTRK3, and PRKCA fusions. Melanomas also showed a variable number of additional mutations ablating tumor-suppression mechanisms and activating other oncogenic pathways, including CDKN2A loss, PTEN loss, as well as TP53 and TERT-promoter mutations. Moreover, borderline melanocytic tumors displayed the same chromosomal aberrations, but more mutations than nevi and fewer than melanomas. In this context, the notion that melanocytic neoplasms can be classified as benign/malignant is hardly supportable, because all neoplasms harbor a certain number of mutations and the progression risk, that is, the malignant potential, is related and proportional to the burden of pathogenic mutations. Moreover, from the genomic analysis, in parallel to the current diagnostic categories of "nevi," "melanomas," and "melanocytomas," some aggregations or classes of tumors based on the characteristic types of driver mutations/fusions emerge as possible and more rationale, including Spitzoid neoplasms, blue neoplasms, BAP1-inactivated melanocytic neoplasms, deep penetrating melanocytic neoplasms, pigment-synthesizing melanocytic neoplasms, and "common" melanocytic neoplasms. Each of these classes, showing the same driver mutations/fusions, demonstrates to have the same pathogenesis and may be genetically considered as a single tumor, although with a variable amount of progression risk. Histologic features, being an expression of the mutational state, could be used to obtain an approximate risk assessment in each single tumor.

Simple Summary The recent WHO classification of melanocytic tumors requires the implementation of combined phenotypic–genotypic diagnostics. For rare tumors, such as atypical deep penetrating nevi, there is insufficient information regarding genetic status, and it is not yet clear whether the observed unusual morphological cyto-architectures reflect a distinct genomic profile or are associated with an increased metastatic potential and aggressive clinical behavior. We report herein a comprehensive next-generation sequencing (NGS) analysis of a series of atypical DPNs, showing their mutational profile with some specific signatures for these rare and diagnostically challenging tumors. Abstract Deep penetrating nevi (DPNs) are rare melanocytic neoplasms consisting of pigmented spindled or epithelioid melanocytes with a distinctive wedge-shaped configuration showing activation of the WNT pathway, with unusual cyto-architectural features. It is unclear whether they show a distinct genomic profile associated with a diverse metastatic potential. We describe herein a cohort of 21 atypical DPNs analyzed by next-generation sequencing using the Ion AmpliSeq™ Comprehensive Cancer Panel. We found that β-catenin exon 3 was mutated in 95% and MAP kinase pathway genes in 71% of the cases. Less frequent mutations were observed in HRAS (19%) and MAP2K1 (24%). Isocitrate dehydrogenases 1 (IDH1) mutations, including R132C, V178I, and S278L, were identified in 38% of cases and co-existed with BRAF/HRAS mutations. The only case with progressive nodal disease carried alterations in the β-catenin pathway and mutations in IDH1 and NRAS (codon 61). By a comprehensive mutation analysis, we found low genetic heterogeneity and a lack of significant associations between specific gene mutations and histopathological features, despite atypical features. Whether the acquisition of an NRAS or IDH1 mutation in an atypical DPN may represent a molecular evolution implying a pathway to melanoma progression should be confirmed in a larger series.

Background: Recent studies have shown an increasing incidence of cutaneous adnexal carcinomas (CACs). Objectives: The aim of our study was to evaluate incidence and survival for cases of CACs and investigate their association with other skin neoplasms. Methods: We conducted a population-based study. Data on incident cases of CACs were obtained from the Tuscany Cancer Registry between 1985 and 2010. In order to determine whether the occurrence of squamous cell carcinoma (SCC) among patients with CAC is higher or lower than expected in the general population, the standardized incidence ratio (SIR) was calculated. Results: A total of 242 patients with CAC were observed; the age-standardized incidence rate was 3·8 cases per million person-years. From 1997 to 2010 crude incidence rates increased by 159%. Age-specific incidence was higher in men over 80 years old than in women of the same age and younger individuals. Carcinomas of sweat gland origin prevailed; the most common histotype was porocarcinoma and the most frequently affected site was the head/neck. Overall, 88% of CACs were diagnosed at a localized stage. The 5-year overall survival and disease-specific survival rates were 59% [95% confidence interval (CI) 53–65] and 94% (95% CI 91–98), respectively. In the observation cohort, the number of SCCs was significantly higher than expected as the SIR was calculated to be 33·7 (P < 0·001). Conclusions: Increasing incidence warrants awareness and early diagnosis of CACs. Increased SCC incidence among patients with these tumours highlights the relevance of careful skin examination and follow-up.

Malignant melanoma is among the most aggressive cancers and its incidence is increasing worldwide. Targeted therapies and immunotherapy have improved the survival of patients with metastatic melanoma in the last few years; however, available treatments are still unsatisfactory. While the role of the BRAF-MEK1/2-ERK1/2 pathway in melanoma is well established, the involvement of mitogen-activated protein kinases MEK5-ERK5 remains poorly explored. Here we investigated the function of ERK5 signaling in melanoma. We show that ERK5 is consistently expressed in human melanoma tissues and is active in melanoma cells. Genetic silencing and pharmacological inhibition of ERK5 pathway drastically reduce the growth of melanoma cells and xenografts harboring wild-type (wt) or mutated BRAF (V600E). We also found that oncogenic BRAF positively regulates expression, phosphorylation, and nuclear localization of ERK5. Importantly, ERK5 kinase and transcriptional transactivator activities are enhanced by BRAF. Nevertheless, combined pharmacological inhibition of BRAFV600E and MEK5 is required to decrease nuclear ERK5, that is critical for the regulation of cell proliferation. Accordingly, combination of MEK5 or ERK5 inhibitors with BRAFV600E inhibitor vemurafenib is more effective than single treatments in reducing colony formation and growth of BRAFV600E melanoma cells and xenografts. Overall, these data support a key role of the ERK5 pathway for melanoma growth in vitro and in vivo and suggest that targeting ERK5, alone or in combination with BRAF-MEK1/2 inhibitors, might represent a novel approach for melanoma treatment.

This study aimed to investigate the sentinel lymph node (SLN) tumour burden to predict the non-SLN positivity rate and the survival of melanoma patients to evaluate whether SLN microstaging could predict the prognosis, similar to what is currently performed by examining the lymph nodes excised by complete lymph node dissection. Of 1130 consecutive melanoma patients who underwent SLN biopsy, 226 were tumour-positive and 204 were included in this study. SLN metastases were classified on the basis of dimensional (Rotterdam) and topographic (Dewar) criteria either separately or combined. SLN metastases more than 1 mm in diameter had the highest non-SLN positivity rate (31%) compared with metastases 0.1-1 mm (10%) and less than 0.1 mm (4%). The non-SLN positivity rate was 45% for extensive metastases, 5% for subcapsular metastases and 23-29% for parenchymal, combined and multifocal classes, therefore suggesting a simplification of the parenchymal SLN metastases into only two classes: extensive and 'not extensive'. The dimension of the metastasis was correlated with a different non-SLN positivity rate only when the metastasis was in the parenchyma (20-36%) and not when it was in the subcapsular location (4-7%). Interestingly, the 5-year melanoma-specific survival (MSS) was 89% for patients with subcapsular less than 0.1 mm metastases and 45% for patients with nonsubcapsular more than 1 mm metastases (P=0.017). In the parenchyma, larger metastases (1 mm) were related to a lower 5-year MSS (46%) than smaller (1 mm) metastases (MSS 77%). SLN tumour burden characterization can be simplified and it can provide prognostic information on non-SLN positivity and survival, which is especially useful in patients who do not undergo complete lymph node dissection.

Electrochemotherapy (ECT) represents an effective local treatment for skin unresectable melanoma metastases with high overall objective response rate. ECT is based on the combination of anti-neoplastic drugs administration and cancer cells electroporation. Whether ECT can also activate the immune system is a matter of debate, however a significant recruitment of dendritic cells in melanoma treated metastases has been described. Herein we investigated immediate and late effects of ECT treatment on T cell subsets in ECT-treated lesions by fluorescent immunohistochemistry. Biopsies from melanoma patients (n = 10) were taken before ECT (t0), at d1 and d14 from treatment. At t0, CD3+CD4+ T cells were the most represented T cells, well detected in the perilesional dermis, particularly at tumour margin, while CD3+CD8+ T cells were less represented. CD4+FOXP3+ T regulatory (Treg) cells were present in the perilesional dermis and within the lesion. ECT induced a significant decrease of CD4+FOXP3+ Treg cells percentage in the perilesional dermis, observed at d1 and at d14 (p

Cutaneous melanoma is one of the most aggressive type of skin tumor. Early stage melanoma can be often cured by surgery; therefore current management guidelines dictate a different approach for thin (4mm) melanomas. We have carried out whole-exome sequencing in 5 thin and 5 thick fresh-frozen primary cutaneous melanomas. Unsupervised hierarchical clustering analysis of somatic copy number alterations (SCNAs) identified two groups corresponding to thin and thick melanomas. The most striking difference between them was the much greater abundance of SCNAs in thick melanomas, whereas mutation frequency did not significantly change between the two groups. We found novel mutations and focal SCNAs in genes that are embryonic regulators of axon guidance, predominantly in thick melanomas. Analysis of publicly available microarray datasets provided further support for a potential role of Ephrin receptors in melanoma progression. In addition, we have identified a set of SCNAs, including amplification of BRAF and ofthe epigenetic modifier EZH2, that are specific for the group of thick melanomas that developed metastasis during the follow-up. Our data suggest that mutations occur early during melanoma development, whereas SCNAs might be involved in melanoma progression.

T regulatory cells (Tregs), involved in tumour tolerance, can generate Adenosine by CD39/CD73 surface enzymes, which identify four Tregs subsets: CD39+CD73− nTregs, CD39+CD73+ iTregs, CD39−CD73+ oTregs and CD39−CD73− xTregs. In melanoma patients, increased Tregs levels are detected in peripheral blood (PB), sentinel lymph node (SLN) and tumour infiltrating lymphocytes (TILs), but Adenosine role was not investigated yet. We examined total Tregs and Adenosine subsets in PB, SLN and TILs from melanoma patients (n = 32) and PB from healthy donors (HD; n = 10) by flow cytometry. Total Tregs significantly increased in stage III-IV patients PB, in SLN and TILs, as compared to HD/stage I-II patients. Tregs subsets analyses showed that: 1) PB nTregs significantly increased in SLN and decreased in TILs; 2) iTregs significantly increased in stage III-IV patients PB and further significantly increased in SLN and TILs; 3) PB oTregs and xTregs significantly decreased in SLN and TILs. Patients clinical features did not significantly influence total Tregs, except SLN excision order. Results confirmed Tregs role in melanoma progression and indicate Adenosine generation as a novel escape mechanism, being nTregs and iTregs increased in PB/SLN/TILs.

The CD63 tetraspanin is highly expressed in the early stages of melanoma and decreases in advanced lesions, suggesting it as a possible suppressor of tumor progression. We employed loss- and gain-of-gene-function approaches to investigate the role of CD63 in melanoma progression and acquisition of the epithelial-to-mesenchymal transition (EMT) program. We used two human melanoma cell lines derived from primary tumors and one primary human melanoma cell line isolated from a cutaneous metastasis, differing by levels of CD63 expression. CD63-silenced melanoma cells showed enhanced motility and invasiveness with downregulation of E-cadherin and upregulation of N-cadherin and Snail. In parallel experiments, transient and stable ectopic expression of CD63 resulted in a robust reduction of cell motility, invasiveness, and protease activities, which was proportional to the increase in CD63 protein level. Transfected cells overexpressing the highest level of CD63 when transplanted into immunodeficient mice showed a reduced incidence and rate of tumor growth. Moreover, these cells showed a reduction of N-cadherin, Vimentin, Zeb1, and a-SMA, and a significant resistance to undergo an EMT program both in basal condition and in the following stimulation with TGFβ. Thus, our results establish a previously unreported mechanistic link between the tetraspanin CD63 and EMT abrogation in melanoma.

Background. Scalp/neck melanomas have a poor progno- sis, possibly because of a rich vascular supply that prompts tumor cells' dissemination. Methods. We compared the accuracy of immunohistochemical (IHC) staining with morphology for the identification of lymphovascular inva- sion in 156 scalp/neck melanomas. We then analyzed the association of vessel invasion and density with pathological features and survival. Results. IHC-detected lymphatic vessel invasion (LVI) and blood vessel invasion (BVI) were identified in 34.6% and 13.5% of cases, respectively. IHC increased the LVI/BVI detection compared to morphology (40.4% vs 16.6%; p

Background Merkel cell polyomavirus (MCPyV) is associated with Merkel cell carcinoma and high viral load in the skin was proposed as a risk factor for the occurrence of this tumour. MCPyV DNA was detected, with lower frequency, in different skin cancers but since the viral load was usually low, the real prevalence of viral DNA could be underestimated. Objective To evaluate the performance of two assays (qPCR and ddPCR) for MCPyV detection and quantification in formalin fixed paraffin embedded (FFPE) tissue samples. Methods Both assays were designed to simultaneous detection and quantification of both MCPyV as well as house-keeping DNA in clinical samples. The performance of MCPyV quantification was investigated using serial dilutions of cloned target DNA. We also evaluated the applicability of both tests for the analysis of 76 FFPE cutaneous biopsies. Results The two approaches resulted equivalent with regard to the reproducibility and repeatability and showed a high degree of linearity in the dynamic range tested in the present study. Moreover, qPCR was able to quantify ≥10 5 copies per reaction, while the upper limit of ddPCR was 10 4 copies. There was not significant difference between viral load measured by the two methods The detection limit of both tests was 0,15 copies per reaction, however, the number of positive samples obtained by ddPCR was higher than that obtained by qPCR (45% and 37% respectively). Conclusions The ddPCR represents a better method for detection of MCPyV in FFPE biopsies, mostly these containing low copies number of viral genome.

Histopathological evaluation of melanocytic tumours usually allows reliable distinction of benign melanocytic naevi from melanoma. More difficult is the histopathological classification of Spitz tumours, a heterogeneous group of tumours composed of large epithelioid or spindle-shaped melanocytes. Spitz tumours are biologically distinct from conventional melanocytic naevi and melanoma, as exemplified by their distinct patterns of genetic aberrations. Whereas conventional naevi and melanoma often harbour BRAF mutations, NRAS mutations, or inactivation of NF1, Spitz tumours show HRAS mutations, inactivation of BAP1 (often combined with BRAF mutations), or genomic rearrangements involving the kinases ALK, ROS1, NTRK1, BRAF, RET, and MET. In Spitz naevi, which lack significant histological atypia, all of these mitogenic driver aberrations trigger rapid cell proliferation, but after an initial growth phase, various tumour suppressive mechanisms stably block further growth. In some tumours, additional genomic aberrations may abrogate various tumour suppressive mechanisms, such as cell-cycle arrest, telomere shortening, or DNA damage response. The melanocytes then start to grow in a less organised fashion, may spread to regional lymph nodes, and are termed atypical Spitz tumours. Upon acquisition of even more aberrations, which often activate additional oncogenic pathways or reduce and alter cell differentiation, the neoplastic cells become entirely malignant and may colonise and take over distant organs (spitzoid melanoma). The sequential acquisition of genomic aberrations suggests that Spitz tumours represent a continuous biological spectrum, rather than a dichotomy of benign versus malignant, and that tumours with ambiguous histological features (atypical Spitz tumours) might be best classified as low-grade melanocytic tumours. The number of genetic aberrations usually correlates with the degree of histological atypia and explains why existing ancillary genetic techniques, such as array comparative genomic hybridisation (CGH) or fluorescence in situ hybridisation (FISH), are capable of accurately classifying histologically benign and malignant Spitz tumours, but are not very helpful in the diagnosis of ambiguous melanocytic lesions. Nevertheless, we expect that progress in our understanding of tumour genomics and progression will refine the classification of melanocytic tumours in the near future. By integrating clinical, pathological, and genetic criteria, distinct tumour subsets will be defined within the heterogeneous group of Spitz tumours, which will eventually lead to improvements in diagnosis, prognosis and therapy.

Increasing evidences support the role of Merkel cell polyomavirus (MCPyV) and human papillomavirus (HPV) in non-cutaneous and cutaneous tumours. Porocarcinoma is a rare malignant neoplasm that arises from the intraepidermal ductal portion of the eccrine sweat glands. The aetiology of porocarcinoma is largely unknown and no systematic studies have been done to investigate the implication of infectious agents in the pathogenesis of this tumour.To investigate the possible association between MCPyV and/or HPV infection and porocarcinoma.Forty-four formalin-fixed paraffin-embedded (FFPE) porocarcinomas (40 primary and 4 metastatic) and 10 healthy skin specimens (controls), were analysed for the presence of MCPyV and HPV DNA using molecular detection methods.MCPyV DNA was found in 27/40 (68%) primary porocarcinomas and in 3/10 (30%) controls (Fisher exact test: p0.04). No significant difference in viral load was observed between tumours and healthy skin. Moreover, 2/40 primary porocarcinomas tested positive for high-risk HPV16. Cutaneous beta-HPV infection was detected in 16/40 (40%) porocarcinomas and in 6/10 (60%) controls. No particular beta-HPV types were significantly associated with tumour or with healthy skin. Two out of 4 metastatic biopsies were MCPyV DNA positive. All metastatic samples had mixed infections with cutaneous HPV types.This study demonstrated a significantly high prevalence of MCPyV and the presence of a broad spectrum of HPV types in porocarcinoma and provided the first available data about viral infections in this tumour. To understand the role, if any, of viral infections in the pathogenesis of porocarcinoma further studies are needed.

Introduction The diagnosis of cutaneous pigmented lesions remains a challenge for both dermatologists and pathologists. Our aim was to determine the diagnostic concordance between the conventional face-to-face diagnosis and the telediagnosis of 10 dermatologists with expertise in dermato-oncology of 10 challenging pigmented lesions. Methods Using a store-and-forward teledermatology method, clinical and dermoscopic digital images of all selected lesions were transmitted via e-mail to 10 dermatologists. Dermatologists were called to provide their telediagnoses with a step-by-step approach. When the dermatologists responded with their first clinical telediagnosis, they received a second email that contained dermoscopic images of the 10 cases. Final histopathological diagnosis was considered the gold standard for comparison with face-to-face and teledermatology diagnoses in statistical analysis. Results Face-to-face results indicated moderate agreement between clinical and histopathological diagnoses (K = 0.6). After the first clinical step, interobserver concordance of telediagnosis was lower than face-to-face diagnosis (K = 0.52). After the second dermoscopy step, the concordance declined further (K = 0.38). Conclusions Teledermatology was inferior to face-to-face dermatology. Moreover, the diagnostic concordance of telediagnosis decreased after the teledermoscopic step. This finding may be justified by the dermoscopic difficulty of the selected lesions, including Spitzoid proliferations and atypical melanocytic nevi of the elderly. These lesions may represent a potential diagnostic pitfall given their confounding dermoscopic aspects.

We report a 20-year-old male patient with an atypical Spitz tumor, located in the dorsal aspect of his left hand, and a positive sentinel axillary lymph node. After lymphadenectomy, 1 of 23 non-sentinel lymph nodes excised was found to contain small multiple deposits of large spindle atypical melanocytes. Reviewing the pertinent literature, 5 of 29 patients with atypical Spitz tumors and positive sentinel nodes who had undergone lymphadenectomy have shown non-sentinel node involvement (17.2%), a proportion similar to that reported in melanoma patients. The exact nature of atypical Spitz tumors and the interpretation of cell deposits detected in sentinel nodes are still debated; data regarding the non-sentinel lymph node involvement in patients with atypical Spitz tumors may contribute to better understand the real biological potential of such tumors.

Dermatofibrosarcoma protuberans (DFSP) is a rare skin cancer with intermediate malignancy, characterized by a progressive local growth and a propensity for local recurrence. DFSP is most frequent in adults; however, in recent years, DFSP in childhood emerged to be more common than previously believed. Unfortunately DFSP in children may be misdiagnosed, leading to a delay in the treatment. The authors report two cases of childhood DFSP with unusual clinical presentation: a congenital nodular variant and an atrophic variant developed at 2 years of age, both with acral localization. They highlight the importance of an early diagnosis by pediatricians and dermatologists to ensure an appropriate complete excision and reduce the risks of recurrences.

Tubular adenoma (TA) and syringocystadenoma papilliferum (SCAP) may show histopathological overlap, with some lesions having features of both neoplasms (SCAP + TA). TA has been recently suggested to represent a carcinoma. Four observers blindly assessed 67 cases of TA, SCAP, and their lookalikes (poroma, apocrine adenoma, apocrine carcinoma; all lesions focally featuring a pseudopapillary pattern), and classified the lesions into one of four categories: (1) TA, (2) SCAP, (3) SCAP + TA, and (4) others. Lesions were also classified as benign or malignant. In only 29 cases was there unanimous agreement among the four observers, who classified 22 lesions as TA, three as SCAP, and four cases as others. Of the 38 cases where there was interobserver diagnostic variation, in 30, the diagnosis varied between TA or SCAP or SCAP + TA; the remainder fell in the others category. Analysis of the factors leading to interobserver variability indicated that diagnostic problems occurred when there were any of the following: epidermal acanthosis, papillomatosis, connection of the neoplastic tubules to the overlying epidermis and/or follicular infundibula, and plasma cell infiltration. These features accounted for the morphological overlap between TA and SCAP. All observers agreed that the lesions were benign; the only apocrine carcinoma included was recognized as such by all observers. From the study, it was concluded that TA may arise in the deep dermis without any epidermal connection, or, in other cases, it may be more superficially located with or without an epidermal connection. It may be reasonably inferred that, possibly as a response to infection, there may be accompanying plasma cells and variable acanthosis and papillomatosis, such that the appearances are those of "pure" SCAP, or lesions may have features "intermediate" or overlapping between TA and SCAP.

Background Mucosal melanomas (MM) represent a heterogeneous tumour population that exhibits site-specific molecular profiles. Aims In a multicentre retrospective study, we investigated KIT aberrations in primary anorectal (AR) melanomas compared with melanoma metastatic to the gastrointestinal (GI) tract. Methods Primary AR MM (n=31) and GI metastatic melanoma (n=27) were studied for KIT mutations on exons 11, 13, 17 and 18 by high-resolution melting analysis, direct sequencing and c-KIT expression by immunohistochemistry. Selected cases were also investigated for increased KIT gene copy number by fluorescent in situ hybridisation. Results Functional KIT mutations were demonstrated in 11/31 (35.5%) of AR melanomas and in 1/26 (3.8%) of GI melanoma metastases (p=0.004). A significant difference emerged between primary and metastatic MM with regards to KIT-positive immunostaining (p=0.002). Immunohistochemical c- KIT protein overexpression did not correlate with KIT mutational status. Increased KIT copy number was demonstrated in 5/20 AR primary cases. Conclusions The rate of functional mutations in KIT is significantly higher in AR MM than in GI metastatic melanoma. KIT protein overexpression does not correlate with KIT mutations and cannot be used for screening purposes. Recognising the molecular heterogeneity of MM helps to identify patients who require a different therapeutic approach.

Testing for NRAS is now integral part in the assessment of metastatic melanoma patients because there is evidence that NRAS-mutated patients may be sensitive to MEK inhibitors, and RAS mutation is a common mechanism of acquired resistance during treatment with BRAF inhibitors. This study evaluated the sensitivity and specificity of immunohistochemical analysis using an N-Ras (Q61R) antibody to detect the presence of the NRASQ61R mutation in melanoma patients. A total of 98 primary cutaneous melanomas that have undergone examination of NRAS mutation were retrieved from a multicentric database. Formalin-fixed and paraffin-embedded melanoma tissues were analyzed for BRAF and NRAS mutations by independent, blinded observers using both conventional DNA molecular techniques and immunohistochemistry with the novel anti-human N-Ras (Q61R) monoclonal antibody (clone SP174). The antibody showed a sensitivity of 100% (14/14) and a specificity of 100% (83/83) for detecting the presence of an NRASQ61R mutation. Of the NRAS-mutated cases, none of the non-Q61R cases stained positive with the antibody (0/7). There were three cases with discordant NRAS mutational results. Additional molecular analysis confirmed the immunohistochemically obtained NRAS result in all cases, suggesting that a multiple analytical approach can be required to reach the correct sample classification. The reported immunohistochemical method is an accurate, rapid, and cost-effective method for detecting NRASQ61R mutation in melanoma patients, and represents a valuable supplement to traditional mutation testing. If validated in further studies, genetic testing would only be required for immunohistochemistry-negative patients to detect non-Q61R mutations.

The distinction between Spitz nevus and melanoma is currently possible, applying a set of definite histological criteria. However, in certain lesions deviating from the stereotypical morphology of classic Spitz nevi ("atypical Spitz tumors"), the differentiation between benign and malignant cases appears problematic because objective criteria for a reliable diagnosis are lacking. We report the clinicopathologic findings of 12 patients with atypical Spitz tumors, who underwent sentinel node biopsy. All the tumors, composed of spindle and/or epithelioid cells, histologically showed features referable to Spitz nevi mixed to features generally found in malignant melanomas. Nine patients were females and three males, ranging in age from 2 to 48 years (mean, 23.2 years). The size of lesions ranged from 5 to 9 mm, the thickness from 1.12 to 5.70 mm. Nodal micrometastases were found in 4 (33.3%) patients. Among the patients with positive sentinel node, two showed minimal nodal involvements; one patient showed additional tumor deposits in one nonsentinel regional node. All patients are alive and free of disease with a follow-up of 2 to 90 months (mean, 26.3 months). Metastasizing and nonmetastasizing cases appeared clinically and histologically indistinguishable. The statistical analysis showed no significant difference between the two groups. Results suggested that all the reported cases may constitute a relatively homogeneous morphological group of lesions with a relevant metastatic potential that may be underdiagnosed.

Background: Melanocytic nevi in the genital, acral, and flexural sites often display clinical and histologic features that may simulate melanoma. We verified whether this is the case also for nevi of the breast. Methods: Eleven dermatopathologists, from nine Italian Institutions, collected the specimens of melanocytic lesions from the breast and other body sites, excluding the acral, genital, and flexural areas, as controls. Cases and controls were matched for sex and age. All nevi were observed ‘blindly’ and simultaneously by all participants. For each lesion, 10 histological parameters were analyzed: asymmetry, absence of lateral demarcation of melanocytes, lentiginous proliferation, nested and dyshesive pattern, intraepidermal melanocytes above the basal layer, involvement of the hair follicle, absence of maturation of dermal melanocytes, melanocytic atypia, fibroplasia of the papillary dermis, and lymphocytic dermal infiltrate. Each parameter was scored 2 when present and 1 when absent or not valuable. A total score was calculated for each lesion. Results were statistically analyzed by the chi-square test and the Mann–Whitney U-test. Results: One hundred and one nevi came from the breast area and 97 from elsewhere. Breast nevi exhibited significantly more atypical features than nevi from other sites. In particular, breast nevi with intraepidermal melanocytes, melanocytic atypia, and dermal fibroplasia were significantly more numerous. We did not find any sexual difference. Conclusions: To avoid undue concerns, dermatopathologists should be aware that melanocytic nevi of the breast may show a high degree of atypical features.

The status and relevance of repetitive nucleotide sequences or microsatellite alterations in sporadic cutaneous melanoma has not been fully clarified. In this study we evaluated the presence of microsatellite alterations in a series of sporadic primary and metastatic melanomas in order to discover which genetic events may have a pathogenetic role in the development of this disease. Tumour samples were obtained from 21 patients with sporadic cutaneous melanoma, and from eight corresponding positive sentinel lymph nodes and one corresponding in-transit metastasis. In each specimen, selected neoplastic cells were procured by laser-assisted microdissection. Polymerase chain reaction-based microsatellite analysis was performed using a panel of 11 microsatellite markers, located at chromosome 2p, 4q, 9p, 16q, 17p and 21q. Overall, we found microsatellite alterations in five (23.8%) melanomas. Of these, one case showed alteration at marker D2S2182 and one at marker D17S261, whereas in another case alterations at three loci, D2S2182, D2S2291 and D9S171, were found. The fourth patient demonstrated an alteration at locus D9S171 both in the primary tumour and in the histologically positive sentinel lymph node. The fifth case was characterized by alterations at D2S2182 and at D17S250, whereas the corresponding in-transit metastasis showed the same alterations as the primary tumour and an additional alteration at IFN alpha. In conclusion, our study confirms previous observations that cutaneous melanomas demonstrate microsatellite alterations, although such instability occurs at a lower frequency than specific mismatch repair defects. Genetic analysis of metastatic lesions revealed that the same microsatellite alterations as in the primary tumour are seen, but additional genetic changes may develop during disease progression.

background. CO2 laser is currently one of the most versatile and useful laser devices in dermatologic practice in the fields of both cosmetology and oncology. The CO2 laser in a superpulsed mode enables the operator to effect precise and adequate vaporization of the affected area, with the possibility to appreciate visually the depth achieved, and, when necessary, to perform histologic and cytologic examination of the surrounding skin. objective. To show superpulsed CO2 laser treatment of basal cell carcinoma. methods. One hundred forty patients presenting single, multiple, superficial, or nodular basal cell carcinoma (BCC) have been treated with the superpulsed CO2 laser. Before the laser treatment the lesion was subjected to cytologic examination by scraping; this examination was then repeated when the papillary dermis was clinically detectable, and again when the operator considered the BCC completely vaporized. In selected subjects, histopathologic examination was done three times (biopsies were obtained at the same time as the samples for the cytologic examination). results. Recovery time was fast, with good healing outcomes. After 3 years follow-up no recurrences were seen. The cytologic and histopathologic examinations showed BCC in the specimens obtained prior to and during laser therapy, but not in those obtained after laser therapy. conclusion. This technique causes minimal thermal cellular damage and no severe morphologic cellular alterations. Thus it permits an intraoperatory cytologic and histopathologic examination. Finally, this technique enables the operator to recognize the different skin levels removed by vaporization and to stop the vaporization as soon as unaffected dermis has been reached, as shown by intraoperatory cytologic and histopathologic examination.

Context.—Several aspects of sweat gland carcinomas (incidence, classification, diagnosis, and behavior) have not been definitively clarified and need to be studied further. Objective.—The clinicopathologic findings of a large series of sweat gland carcinomas, collected during a period of 15 years, are presented. Methods.—Sixty sweat gland carcinomas (41 porocarcinomas, 3 syringomatous carcinomas, 8 ductal carcinomas, 5 adenoid cystic carcinomas, and 3 mucinous carcinomas) were analyzed histologically and immunohistochemically. Results.—Porocarcinomas were composed of eosinophilic and clear atypical cells arranged in solid-cystic lobular masses. These tumors were divided into 2 subgroups: horizontal porocarcinomas, showing a prominent intraepidermal component, and nodular porocarcinomas, which demonstrated predominant nodular growth. Syringomatous carcinomas presented keratinizing and nonkeratinizing cysts, dilated tubules (sometimes with a “tadpole” appearance), small neoplastic ducts, solid islands, and cellular cords. Ductal carcinomas were characterized by a prominent formation of tubules, solid islands, and cellular cords. Adenoid cystic carcinomas presented a characteristic pattern, showing basaloid monomorphous cells with moderately atypical nuclei, arranged in cribriform or solid islands and in tubular structures. Mucinous carcinomas were composed of moderately atypical cells with eosinophilic vacuolated cytoplasm, forming solid and cystic islands floating in large mucin pools. Immunohistochemically, cytokeratin was found in neoplastic cells in all cases, carcinoembryonic antigen was detected in 73% of cases, and actin-positive (myoepithelial) cells were not found. Conclusions.—Although numerous studies have been published in recent years, the histologic features, histogenesis, and classification of sweat gland carcinomas still remain controversial and need to be clarified by further studies.

The atypical histologic features considered to be specific to dysplastic (atypical) nevi have been reported to occur in nevi that are common nevi by all other clinical and histologic features. The distribution and mutual relations among such features in nevi need to be further studied. Six histologic features (dimension > 5 mm, lentiginous proliferation, disordered nested pattern, melanocytic dyskaryosis, dermal lymphocytic infiltrate, suprabasal melanocytes) were analyzed in 253 melanocytic nevi with different clinical appearances. Atypical histologic features, found in 72% of nevi, occurred singly or formed numerous and highly variable combinations. Nevi formed a complex histologic spectrum comprising lesions showing a progressively increasing incidence of atypical features rather than two classes (common and dysplastic nevi). To divide the investigated lesions in objectively defined groups, we used a scoring system. In each nevus, a numeric value of 1 was assigned when each of the studied parameters was present and a value of 0 was assigned when each of these parameters was absent; on the basis of the final scores, nevi were divided in six different classes (classes 0-5). Diagnostic categories such as dysplastic nevi and common nevi seem to be inappropriate, as they do not reflect the real histologic complexity of such lesions.

BACKGROUND The c-erbB-2 gene codes for a putative transmembrane protein, similar in structure to the epidermal growth factor receptor. Amplification and/or overexpression of the gene has been recently described with a prognostic significance in a variety of human adenocarcinomas. METHODS A monoclonal antibody against the c-erbB-2 oncoprotein has been used immunocytochemically in a retrospective study of formalin-fixed, paraffin-embedded samples from 28 consecutive intestinal-type adenocarcinomas (ITACs) of the nose and paranasal sinuses. RESULTS Nine out of 28 primary adenocarcinomas (32%) showed positive staining. Clinical follow-up data, available for all patients, suggested in a univariate analysis a correlation between c-erbB-2 expression and poor prognosis, as measured by 5-year disease-free (p = .02) and overall survival curves (p = .07) as well as by recurrence of disease and the appearance of regional and distant metastases (p = .08). In multivariate analysis, c-erbB-2 expression was statistically significant in terms of disease-free survival (p = .046) but not of overall survival (p = .091) in our series. CONCLUSIONS These data indicate that c-erbB-2 oncogene activation could be involved in sinonasal tract oncogenesis, with possible prognostic implications.

A 96-year-old man presented with a polypoid melanoma, which showed a prominent clear stroma, similar to that of myxoid melanoma, but stained negative for Alcian blue at pH 2.5 and for colloidal iron. It is not clear whether melanoma displaying pseudomyxoid features (pseudomyxoid melanoma) may represent a distinct histological variant of melanoma. However, it must be differentiated from true myxoid melanomas, in which Alcian blue-positive acid mucopolysaccharides are present. Urso C. Cutaneous melanoma with pseudomyxoid features.