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Background: The prognostic impact of variant allele frequency (VAF) on clinical outcome in BRAFV600 mutated metastatic melanoma patients (MMPs) receiving BRAF (BRAFi) and MEK inhibitors (MEKi) is unclear., Materials and Methods: A cohort of MMPs receiving first line BRAFi and MEKi was identified by inspecting dedicated databases of three Italian Melanoma Intergroup centres. VAF was determined by next generation sequencing in pre-treatment baseline tissue samples. Correlation between VAF and BRAF copy number variation was analysed in an ancillary study by using a training and a validation cohort of melanoma tissue samples and cell lines., Results: Overall, 107 MMPs were included in the study. The VAF cut-off determined by ROC curve was 41.3%. At multivariate analysis, progression-free survival (PFS) was significantly shorter in patients with M1c/M1d [HR 2.25 (95% CI 1.41-3.6, p < 0.01)], in those with VAF >41.3% [HR 1.62 (95% CI 1.04-2.54, p < 0.05)] and in those with ECOG PS ≥1 [HR 1.82 (95% CI 1.15-2.88, p < 0.05)]. Overall survival (OS) was significantly shorter in patients with M1c/M1d [HR 2.01 (95% CI 1.25-3.25, p < 0.01)]. Furthermore, OS was shorter in patients with VAF >41.3% [HR 1.46 (95% CI 0.93-2.29, p = 0.06)] and in patients with ECOG PS ≥1 [HR 1.52 (95% CI 0.94-2.87, p = 0.14)]. BRAF gene amplification was found in 11% and 7% of samples in the training and validation cohort, respectively., Conclusions: High VAF is an independent poor prognostic factor in MMP receiving BRAFi and MEKi. High VAF and BRAF amplification coexist in 7%-11% of patients., (© 2023 The Authors. Journal of the European Academy of Dermatology and Venereology published by John Wiley & Sons Ltd on behalf of European Academy of Dermatology and Venereology.)

Background: A polygenic inheritance involving high, medium and low penetrance genes has been suggested for melanoma susceptibility in adults, but genetic information is scarce for paediatric patients., Objective: We aim to analyse the major high and intermediate melanoma risk genes, CDKN2A, CDK4, POT1, MITF and MC1R, in a large multicentre cohort of Italian children and adolescents in order to explore the genetic context of paediatric melanoma and to reveal potential differences in heritability between children and adolescents., Methods: One-hundred-twenty-three patients (<21 years) from nine Italian centres were analysed for the CDKN2A, CDK4, POT1, MITF, and MC1R melanoma predisposing genes. The rate of gene variants was compared between sporadic, familial and multiple melanoma patients and between children and adolescents, and their association with clinico-pathological characteristics was evaluated., Results: Most patients carried MC1R variants (67%), while CDKN2A pathogenic variants were found in 9% of the cases, the MITF E318K in 2% of patients and none carried CDK4 or the POT1 S270N pathogenic variant. Sporadic melanoma patients significantly differed from familial and multiple cases for the young age at diagnosis, infrequent red hair colour, low number of nevi, low frequency of CDKN2A pathogenic variants and of the MC1R R160W variant. Melanoma in children (≤12 years) had more frequently spitzoid histotype, were located on the head/neck and upper limbs and had higher Breslow thickness. The MC1R V92M variant was more common in children than in adolescents. CDKN2A common polymorphisms and MC1R variants were associated with a high number of nevi., Conclusion: Our results confirm the scarce involvement of the major high-risk susceptibility genes in paediatric melanoma and suggest the implication of MC1R gene variants especially in the children population., (© 2021 European Academy of Dermatology and Venereology.)

Background: Many melanoma observational studies have been carried out across different countries and geographic areas using heterogeneous assessments of epidemiologic risk factors and clinical variables., Aim: To develop a consensus questionnaire to standardize epidemiologic and clinical data collection for melanoma risk assessment., Methods: We used a stepwise strategy that included: compilation of variables from case-control datasets collected at various centres of the MelaNostrum Consortium; integration of variables from published case-control studies; consensus discussion of the collected items by MelaNostrum members; revision by independent experts; addition of online tools and image-based charts; questionnaire testing across centres and generation of a final draft., Results: We developed a core consensus questionnaire (MelanoQ) that includes four separate sections: A. general and demographic data; B. phenotypic and ultraviolet radiation exposure risk factors and lifestyle habits; C. clinical examination, medical and family history; and D. diagnostic data on melanoma (cases only). Accompanying online tools, informative tables, and image-based charts aid standardization. Different subsections of the questionnaire are designed for self-administration, patient interviews performed by a physician or study nurse, and data collection from medical records., Conclusions: The MelanoQ questionnaire is a useful tool for the collection and standardization of epidemiologic and clinical data across different studies, centres, cultures and languages. This will expedite ongoing efforts to compile high-quality data for pooled analyses or meta-analyses and offer a solid base for the design of clinical, epidemiologic and translational studies on melanoma., (© 2018 European Academy of Dermatology and Venereology.)

Background: Histopathological diagnosis including selection of lesions, the determination of the best point of time for biopsy and workup is not trivial in cutaneous graft-versus-host disease (GvHD)., Objectives: To develop interdisciplinary recommendations on performing, the laboratory work up and reporting of the results of skin biopsies in patients with suspected cutaneous GvHD., Methods: A working group consisting of dermatopathologists, dermatologists, transplant-physicians and transplant-pathologists prepared recommendations for performing skin biopsies, laboratory workup and evaluation of tissue samples, and reporting of the results in patients with cutaneous GvHD. After achieving a consensus within the working group, a survey that comprised the core issues of the recommendations was electronically sent out to 72 alloHSCT centres within Germany, Austria, and Switzerland and their Departments of Pathology. The answers were discussed in a Consensus Conference and final recommendations were established., Results: Twenty-five centres responded to the clinical and 17 centres to the histopathological survey. Questions addressed to the clinicians comprised the indication for skin biopsy in chronic GvHD (cGvHD) and acute GvHD (aGvHD) and the appropriate point of time for skin biopsy. Eighty-eight per cent agreed that the skin biopsy is generally indicated in patients with suspected cGvHD lacking diagnostic features. In contrast, with suspected aGvHD, only 62% of respondents felt that skin biopsy was necessary even if GvHD had not been confirmed in another organ. Although restricted due to the fact that immunosuppression is often applied in an emergency setting most centres supported skin biopsies before initiation of topical or systemic immunosuppression. The majority of pathologists agreed that in non-sclerotic GvHD a punch biopsy is adequate, whereas in sclerotic GvHD a scalpel biopsy is preferred., Conclusion: While a consensus on the need for biopsies in cGvHD was reached the value of skin biopsies in aGvHD and subsequent biopsies during therapy requires further evaluation., (© 2014 European Academy of Dermatology and Venereology.)

Background: Patients who develop cutaneous melanoma are at increased risk of developing a second primary melanoma. There are many aetiological reasons by which the risk of a second melanoma increases. Among others, genetic factors may contribute to modulating this risk. The risk of identifying a CDKN2A germline mutation increases with the number of primary melanomas and with the presence of familial history of melanoma. Patients with melanoma are especially encouraged to have regular follow-up visits with their dermatologist to perform clinical and dermatoscopic examination. In particular, dermoscopy could be very useful in multiple primary melanoma (MPM) patients., Objectives: To analyse the clinical and dermatoscopic features of multiple melanomas, focusing on those features that are more frequently found in the same patient to recognize them earlier and understand whether they appear with the similar peculiar dermatoscopic features, especially in CDKN2A carriers., Methods: Medical records of MPM patients were selected from a database including 1065 patients with histopathologically proven melanoma diagnosis, all treated at the dermatology clinic of the University of Florence from 2000 to 2013. Pictures of melanoma were independently and blindly administered to three dermatologist experts in dermoscopy to evaluate the presence or absence of ABCD criteria for each clinical image, and the main pattern for the dermoscopic images. The results were then analyzed and crossed to rate the clinical and dermoscopic features of MPM., Results: Seventy five (7.0%) of 1065 patients included in our database were found to carry an MPM disease. Among them, we selected 12 (16%) patients with three or more MPMs. The presence of the CDKN2A melanoma susceptibility gene was observed in 4/12 (33.33%) patients; two patients presented the C500G and c.5 + 1delG polymorphisms in the CDKN2A gene. In CDKN2A carriers, each patient showed a similar and specific dermatoscopic pattern in their lesions., Conclusions: Even being aware of the limitations of this study, according to hereditary characters and their modes of transmissions, we could speculate that for each patient with a CDKN2A germline mutation, it is possible to find the same kind of dermoscopical pattern among their melanocytic tumours., (© 2014 European Academy of Dermatology and Venereology.)

Background: Histopathology is an important tool in diagnosing cutaneous graft-versus-host disease (GvHD). Minimum diagnostic criteria for active chronic GvHD have recently been defined. However, they are not specific and their interpretation is dependent on observer judgement., Aims of the Study: i) to explore interobserver variability in the interpretation of histopathological changes in GvHD, and ii) to analyse the impact of detailed clinical data on histopathological diagnosis of GvHD., Methods: Histopathological slides from 15 skin biopsies of GvHD and from dermatoses with histopathologically similar appearance were sent in two phases to four dermatopathologists experienced in cutaneous GvHD in France, Germany, Italy and Switzerland (first round of 'blind' review followed by a second round with complete clinical information provided)., Results: Interface dermatitis, especially vacuolar alteration, was the most inconsistently evaluated, particularly in cases with minor alterations. Interestingly, for vacuolar alteration and apoptotic keratinocytes, interobserver variability was lower in the adnexal epithelia than in the interfollicular epidermis. Complete clinical information resulted in increased diagnostic confidence and greater concordance on the final diagnosis, rising from 53% (first round, k = 0.345, fair agreement) to 80% (second round, k = 0.529, moderate agreement). The percentage of correct diagnoses increased from 33.3% to 80%., Conclusion: For the diagnosis of GvHD, histopathological analysis is of importance, but, for correct diagnosis, the correlation of pathological findings with clinical results is crucial. In cases of minor alteration, histopathologists should focus on the interpretation of vacuolar changes and apoptotic keratinocytes, possibly on the adnexal epithelia., (© 2013 European Academy of Dermatology and Venereology.)

Background: The differential diagnosis between Reed nevi and melanoma becomes more difficult if the lesion to analyse presents a small size, with a diameter of 6 mm or smaller. Many studies have reported various dermoscopic features of Reed nevi during their growth phases. In early stages of evolution, the lesions generally show a characteristic globular appearance typically found in childhood, followed by the so-called starburst pattern., Objective: The aim of the study was to identify the main dermoscopic features in small Reed nevi (<6 mm in size)., Methods: Using a computerized skin-imaging database for melanoma prevention surgery at the Department of Dermatology of the University of Florence, 15 Reed nevi were selected among 103 small (<6 mm) melanocytic lesions consecutively excised. Images of small Reed nevi, independently blinded to histopathological diagnosis, were administered to a dermatologist expert in dermoscopy, who separately examined the clinical and the dermatoscopic images of small Reed nevi and evaluated their clinical and dermoscopic parameters., Results: Analysis of the main dermoscopic patterns showed that 40% had a reticular pattern, 20% had a starburst pattern, 6.5% had a globular pattern, 6.5% had a homogeneous pattern and 27% had an atypical pattern., Conclusion: We propose that small, early-stage Reed nevus are not characterized by an evolution of growth patterns to a phenotype typical of larger lesions. We assume that the patterns are distributed in a linear manner between age groups, may all be present at the outset and thus are independent from the various stages of nevus development., (© 2012 The Authors. Journal of the European Academy of Dermatology and Venereology © 2012 European Academy of Dermatology and Venereology.)

Background: Two-photon excitation (TPE) fluorescence microscopy is a high-resolution laser-scanning imaging technique enabling deep imaging inside biological tissues. TPE microscopy has the triple advantage of offering high spatial resolution (250 nm radially, 800 nm axially), high penetration depth inside skin (200 mm ), and low photodamage effects. Further, cells and extracellular matrix intrinsically contain a variety of fluorescent molecules (NADH, tryptophan, keratins, melanin, elastin, cholecalciferol and others), so that biological tissues can be imaged by TPE microscopy without any exogenous probe. The time-resolved analysis of the fluorescence signal, known as fluorescence lifetime imaging microscopy (FLIM), is an additional non-invasive microscopy technique useful to characterize endogenous fluorescence species and their surrounding medium by measuring the mean lifetime of fluorescent emission. Finally, multispectral (MTPE) tissue imaging can also be used to identify different endogenous fluorescent species by measuring their two photon emission spectra. Those techniques offer functional information about the relative quantities of fluorescent molecules, which are correlated with tissue structure in physiological and pathological states., Objective: We have decided to apply these three methods at the same time for cutaneous tumors in order to evaluate their possible future use., Method: We have analyzed a melanoma and a basal cell carcinoma, with their surrounding healthy skin, to evaluate any difference in healthy skin and neoplasia. The samples were excised during dermatological surgery, then cut, saving some healthy skin in both, to obtain a regular shape, allowing its positioning either with the skin surface parallel to the optical axis (horizontal optical sectioning), or perpendicular (vertical optical sectioning)., Conclusion: This first result demonstrates that FLIM is effective in discriminating healthy skin from MM, while MTPE is effective in discriminating healthy skin from BCC.

We report two cases of cutaneous B-cell pseudolymphoma (PCBCL) induced by intradermal antigen injections for specific immune therapy (SIT). In both cases, the lesions had first developed on the area of injection; years later, new lesions appeared far from the original site. The histological, immunohistochemical, and molecular findings of the lesions showed features consistent with the diagnosis of PCBCL in both cases. In particular, the staining for sIg light chains showed a polyclonal pattern, and the molecular analysis by reverse transcriptase-polymerase chain reaction (RT-PCR) and Southern blot showed a germ-line configuration of the Ig heavy chain genes. While the development of PCBCL related to a specific stimulus is well known and widely reported, the development of histologically and immunohistologically identical lesions far from the injection site is definitely worthy of note. This behaviour might be due to the presence of retained antigens in the injection site. This chronic antigenic stimulation could induce the progressive selection--and subsequent dissemination--of antigen-specific B cell clones.

Basal cell carcinomas may attain giant proportions due primarily to recurrence or because the tumour is neglected. We report the case of a 66-year-old man who presented with a bleeding, polypoid, cutaneous tumour located on the left shoulder region of 13 years duration. The man had not received any previous treatment. The lesion was biopsied and histopathologically diagnosed as a solid type basal cell carcinoma with focal areas of squamous differentiation and keratinization. The man refused complete surgical removal and therefore was treated with roentgentherapy, with satisfactory results and no complications in the irradiated area. No recurrences had manifested after 1 year follow-up.

We report on a 14-year-old boy who presented with a 4-year history of acral pains and febrile episodes. On physical examination, numerous small reddish papules were present on his abdomen, located predominantly on the periumbelical region. Renal function was within normal limits. Ophthalmological examination revealed whorled opacities of the cornea (cornea verticillata) and dilated tortuous conjunctival vessels. Histopathological examination of one of the cutaneous papules showed several dilated blood vessels in the superficial dermis surrounded by collarettes of thickened rete ridges, consistent with a diagnosis of angiokeratoma. The electron-microscopic study of a skin specimen demonstrated the presence of dilated lysosomes with deposition of electron-dense bodies, some of which with laminated structure, in endothelial cells and fibroblasts. These findings were regarded as indicative of Fabry's disease. Subsequent biochemical analysis confirmed the presence of a alpha-galactosidase A deficiency in leukocytes. In conclusion, we described the clinical, histopathological and submicroscopic findings of a case of Fabry's disease, in which the combination of electron microscopic and biochemical approaches allowed the correct diagnosis.

We report on a patient developing simultaneous occurrence of lentigo maligna lesions, solar lentigines and an extensive melanosis of the oral mucosa. Diagnostically, epiluminescence microscopy had a relevant role in the preoperative assessment and selection of suspicious pigmented lesions, as the lesions histologically labelled as lentigo maligna and solar lentigo were clinically indistinguishable. We review the clinical, dermoscopic and histopathologic differential diagnosis of solar lentigo, malignant lentigo and mucosal melanosis with other melanocytic and keratinocytic lesions and discuss the possible relationship between these entities.