BRAF activation by metabolic stress promotes glycolysis sensitizing NRASQ61-mutated melanomas to targeted therapy.

NRAS-mutated melanoma lacks a specific line of treatment. Metabolic reprogramming is considered a novel target to control cancer; however, NRAS-oncogene contribution to this cancer hallmark is mostly unknown. Here, we show that NRASQ61-mutated melanomas specific metabolic settings mediate cell sensitivity to sorafenib upon metabolic stress. Mechanistically, these cells are dependent on glucose metabolism, in which glucose deprivation promotes a switch from CRAF to BRAF signaling. This scenario contributes to cell survival and sustains glucose metabolism through BRAF-mediated phosphorylation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-2/3 (PFKFB2/PFKFB3). In turn, this favors the allosteric activation of phosphofructokinase-1 (PFK1), generating a feedback loop that couples glycolytic flux and the RAS signaling pathway. An in vivo treatment of NRASQ61 mutant melanomas, including patient-derived xenografts, with 2-deoxy-D-glucose (2-DG) and sorafenib effectively inhibits tumor growth. Thus, we provide evidence for NRAS-oncogene contributions to metabolic rewiring and a proof-of-principle for the treatment of NRASQ61-mutated melanoma combining metabolic stress (glycolysis inhibitors) and previously approved drugs, such as sorafenib.

Changes in the microbiological diagnosis and epidemiology of cutaneous leishmaniasis in real-time PCR era: A six-year experience in a referral center in Barcelona.

BACKGROUND: Leishmaniasis is a neglected disease caused by different species of the protozoa Leishmania spp. Cutaneous lesions are the most common clinical manifestation. This disease is prevalent in tropical and subtropical areas, including the Mediterranean basin. In Spain, Leishmania (L.) infantum is the only endemic species, but imported cases are often diagnosed. Different classical parasitological methods can be performed for cutaneous leishmaniasis (CL) diagnosis; but currently molecular techniques serve as a relevant tool for the detection and characterization of Leishmania parasites. We aimed to evaluate clinical and epidemiological characteristics of CL diagnosed patients by real-time PCR in a tertiary hospital over a six-year period. METHODOLOGY/PRINCIPAL FINDINGS: Clinical, epidemiological and microbiological data were retrospectively collected and analyzed. In our study, CL was confirmed in 59 (31.4%) out of 188 patients by real-time PCR, showing an increase over recent years: 11 cases of CL between 2014 and 2016 and 48 between 2017 and 2019. Real-time PCR was performed on skin swabs and/or biopsies samples, with a positivity of 38.5% and 26.5%, respectively. Results were 100% concordant when biopsy and skin swab were performed simultaneously. L. (L.) infantum was the most frequent species detected (50%), followed by L. (L.) major (45%) and Viannia subgenus (5%), which were detected only in imported cases. L. (L.) major was almost entirely detected in travelers/migrants from Morocco. Multiple and atypical skin lesions were more common in imported cases than in autochthonous cases (44.4% vs. 21.8%). CONCLUSIONS/SIGNIFICANCE: An increase in both autochthonous and imported CL cases has been observed in past years in our hospital. Molecular techniques assist in improving CL diagnosis and characterization of the Leishmania species, mainly in imported cases.

STK11 (LKB1) missense somatic mutant isoforms promote tumor growth, motility and inflammation.

Elucidating the contribution of somatic mutations to cancer is essential for personalized medicine. STK11 (LKB1) appears to be inactivated in human cancer. However, somatic missense mutations also occur, and the role/s of these alterations to this disease remain unknown. Here, we investigated the contribution of four missense LKB1 somatic mutations in tumor biology. Three out of the four mutants lost their tumor suppressor capabilities and showed deficient kinase activity. The remaining mutant retained the enzymatic activity of wild type LKB1, but induced increased cell motility. Mechanistically, LKB1 mutants resulted in differential gene expression of genes encoding vesicle trafficking regulating molecules, adhesion molecules and cytokines. The differentially regulated genes correlated with protein networks identified through comparative secretome analysis. Notably, three mutant isoforms promoted tumor growth, and one induced inflammation-like features together with dysregulated levels of cytokines. These findings uncover oncogenic roles of LKB1 somatic mutations, and will aid in further understanding their contributions to cancer development and progression.

Manifestaciones cutáneas en contexto del brote actual de enfermedad por coronavirus 2019 / Cutaneous manifestations in the current pandemic of coronavirus infection disease (COVID 2019)

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Expanding the Clinical and Genetic Spectra of Primary Immunodeficiency-Related Disorders With Clinical Exome Sequencing: Expected and Unexpected Findings.

Primary immunodeficiencies (PIDs) refer to a clinically, immunologically, and genetically heterogeneous group of over 350 disorders affecting development or function of the immune system. The increasing use of next-generation sequencing (NGS) technology has greatly facilitated identification of genetic defects in PID patients in daily clinical practice. Several NGS approaches are available, from the unbiased whole exome sequencing (WES) to specific gene panels. Here, we report on a 3-year experience with clinical exome sequencing (CES) for genetic diagnosis of PIDs. We used the TruSight One sequencing panel, which includes 4,813 disease-associated genes, in 61 unrelated patients (pediatric and adults). The analysis was done in 2 steps: first, we focused on a virtual PID panel and then, we expanded the analysis to the remaining genes. A molecular diagnosis was achieved in 19 (31%) patients: 12 (20%) with mutations in genes included in the virtual PID panel and 7 (11%) with mutations in other genes. These latter cases provided interesting and somewhat unexpected findings that expand the clinical and genetic spectra of PID-related disorders, and are useful to consider in the differential diagnosis. We also discuss 5 patients (8%) with incomplete genotypes or variants of uncertain significance. Finally, we address the limitations of CES exemplified by 7 patients (11%) with negative results on CES who were later diagnosed by other approaches (more specific PID panels, WES, and comparative genomic hybridization array). In summary, the genetic diagnosis rate using CES was 31% (including a description of 12 novel mutations), which rose to 42% after including diagnoses achieved by later use of other techniques. The description of patients with mutations in genes not included in the PID classification illustrates the heterogeneity and complexity of PID-related disorders.

PD-L1 Expression is Increased in Metastasizing Squamous Cell Carcinomas and Their Metastases.

Programmed cell death ligand 1 (PD-L1) expression by tumor cells plays an important role in the inhibition of T cell-mediated immune response in cancer. PD-L1 expression by tumor cells has been linked to poor prognosis in a wide variety of cancers. However, PD-L1 expression in cutaneous squamous cell carcinoma (cSCC) has been scarcely studied, and its role as a prognosis biomarker remains controversial. The association of PD-L1 expression and the metastatic risk in a series of cSCC was assessed. PD-L1 and CD8 immunostainings of full excision sections of 99 primary tumors and 24 lymphatic metastases were semiquantitatively evaluated. Primary cSCCs were grouped according to the development of lymphatic metastatic spread [metastasizing squamous cell carcinoma (MSCC)] (n = 48) or the absence of progression [nonmetastasizing squamous cell carcinoma (NMSCC)] (n = 51). PD-L1-positive expression (cut off ≥1%) was found in 26% NMSCCs and in 50% MSCCs (P = 0.02). PD-L1 association with an increased metastatic risk was confirmed in the multivariate analysis (P < 0.05), along with the following features: recurrence, poor differentiation, and perineural invasion. Ninety percent of the metastases of PD-L1-positive tumors were also positive for PD-L1, displaying a trend toward a higher PD-L1 expression when compared with their primary tumors (P = 0.058). No significant differences in the peritumoral inflammatory infiltrate or in the expression of CD8 were found between metastasizing and nonmetastasizing primary tumors. Our results suggest that PD-L1 may play a relevant role in metastatic spread and may be a candidate prognostic biomarker in cSCC.

TNFAIP3 haploinsufficiency is the cause of autoinflammatory manifestations in a patient with a deletion of 13Mb on chromosome 6.

There is scarce literature about autoinflammation in syndromic patients. We describe a patient who, in addition to psychomotor and growth delay, presented with fevers, neutrophilic dermatosis, and recurrent orogenital ulcers. Comparative Genomic Hybridization (CGH) array permitted to identify a 13.13Mb deletion on chromosome 6, encompassing 53 genes, and including TNFAIP3 gene (A20). A20 is a potent inhibitor of the NF-kB signalling pathway and restricts inflammation via its deubiquitinase activity. Western blotting and immunoprecipitation assays showed decreased A20 expression and increased phosphorylation of p65 and IkBa. Patient's cells displayed increased levels of total K63-linked ubiquitin and increased levels of ubiquitinated RIP and NEMO after stimulation with TNF. We describe the molecular characterization of an autoinflammatory disease due to a large chromosomal deletion and review the phenotypes of patients with A20 haploinsufficiency. CGH arrays should be the first diagnostic method for comprehensive analysis of patients with syndromic features and immune dysregulation.

The Polycomb proteins RING1B and EZH2 repress the tumoral pro-inflammatory function in metastasizing primary cutaneous squamous cell carcinoma.

Cutaneous squamous cell carcinoma (cSCC) is the second most common malignancy in humans and approximately 5% metastasize, usually to regional lymph nodes. Epigenetic regulation of gene expression may allow tumoral cells to acquire new functions in order to escape from the primary tumor. The aim of this study was to investigate the expression and function of proteins of the Polycomb family of epigenetic regulators in the metastatic process of cSCC. A higher expression of RING1B and EZH2 was detected by immunohistochemistry in a series of primary cSCC tumors that metastasized (MSCCs) when compared with non-metastasizing cSCCs (non-MSCCs). Stable downregulation of RING1B and EZH2 in cSCC cells results in enhanced expression of inflammatory cytokines and activation of the NF-κB signaling pathway. Accordingly, non-MSCCs display higher levels of membranous pS176-inhibitor of NF-kB kinase, and their stroma is enriched in neutrophils and eosinophils when compared with MSCCs. In vitro, hematopoietic cells exhibit a substantial migratory response to supernatants from Polycomb-depleted cSCC cells. Altogether, these data indicate that RING1B and EZH2 repress the innate inflammatory cSCC function and impair tumor immunosurveillance and suggest that patients with high-risk cSCCs could benefit from clinical therapies addressed to harness the immune response.

Epithelial-to-mesenchymal transition in penile squamous cell carcinoma.

PURPOSE: Epithelial-to-mesenchymal transition is a phenomenon in epithelial tumors that involves loss of intercellular adhesion, mesenchymal phenotype acquisition and enhanced migratory potential. While the epithelial-to-mesenchymal transition process has been extensively linked to metastatic progression of squamous cell carcinoma, studies of the role of epithelial-to-mesenchymal transition in squamous cell carcinoma containing high risk human papillomaviruses are scarce. Moreover, to our knowledge epithelial-to-mesenchymal transition involvement in human penile squamous cell carcinoma, which can arise through transforming HPV infections or independently of HPV, has not been investigated. We evaluated the presence of epithelial-to-mesenchymal transition markers and their relationship to HPV in penile squamous cell carcinoma. MATERIALS AND METHODS: We assessed the expression of E-cadherin, vimentin and the epithelial-to-mesenchymal transition related transcription factors Twist, Zeb1 and Snail by immunohistochemical staining in 64 penile squamous cell carcinoma cases. HPV was detected by polymerase chain reaction amplification. RESULTS: Simultaneous loss of membranous E-cadherin expression and vimentin over expression were noted in 43.5% of penile squamous cell carcinoma cases. HPV was significantly associated with loss of membranous E-cadherin but not with epithelial-to-mesenchymal transition. Recurrence and mortality rates were significantly higher in cases showing epithelial-to-mesenchymal transition. CONCLUSIONS: Our findings indicate that in penile squamous cell carcinoma epithelial-to-mesenchymal transition is associated with poor prognosis but not with the presence of HPV.

A mouse model uncovers LKB1 as an UVB-induced DNA damage sensor mediating CDKN1A (p21WAF1/CIP1) degradation.

Exposure to ultraviolet (UV) radiation from sunlight accounts for 90% of the symptoms of premature skin aging and skin cancer. The tumor suppressor serine-threonine kinase LKB1 is mutated in Peutz-Jeghers syndrome and in a spectrum of epithelial cancers whose etiology suggests a cooperation with environmental insults. Here we analyzed the role of LKB1 in a UV-dependent mouse skin cancer model and show that LKB1 haploinsufficiency is enough to impede UVB-induced DNA damage repair, contributing to tumor development driven by aberrant growth factor signaling. We demonstrate that LKB1 and its downstream kinase NUAK1 bind to CDKN1A. In response to UVB irradiation, LKB1 together with NUAK1 phosphorylates CDKN1A regulating the DNA damage response. Upon UVB treatment, LKB1 or NUAK1 deficiency results in CDKN1A accumulation, impaired DNA repair and resistance to apoptosis. Importantly, analysis of human tumor samples suggests that LKB1 mutational status could be a prognostic risk factor for UV-induced skin cancer. Altogether, our results identify LKB1 as a DNA damage sensor protein regulating skin UV-induced DNA damage response.

Epithelial to mesenchymal transition markers are associated with an increased metastatic risk in primary cutaneous squamous cell carcinomas but are attenuated in lymph node metastases.

BACKGROUND: Cutaneous squamous cell carcinoma (cSCC) is the second most common malignancy in humans and approximately 5% metastasize, usually to regional lymph nodes. Epithelial to mesenchymal transition (EMT) is a process involving loss of intercellular adhesion, acquisition of a mesenchymal phenotype and enhanced migratory potential; epithelial markers, such as E-cadherin, are down-regulated and mesenchymal proteins (Vimentin), increased. OBJECTIVE: To investigate the expression of EMT markers in metastatic SCC (MSCC) and their corresponding metastases, and to correlate them with clinico-pathological factors associated with an increased risk of metastasis. METHODS: We performed a retrospective study that included 146 cSCC samples (51 primary non-metastatic, 56 primary metastatic, 39 lymphatic metastases). Immunohistochemistry for E-cadherin, Vimentin, Snail, beta-catenin, Twist, Zeb1 and Podoplanin was performed. RESULTS: Loss of membranous E-cadherin was observed in 77% cSCCs, with no differences between MSCC and non-MSCC. Among the transcriptional factors controlling EMT, no significant Snail1 expression was detected. Twist, Zeb1, Vimentin, beta-catenin and Podoplanin were significantly overexpressed in MSCCs. Twist ectopic expression in SCC13 cells induced Zeb1, Vimentin and Podoplanin expression and E-cadherin delocalization. These changes resulted in a scattered migration pattern in vitro. Expression of EMT markers was decreased in the metastases when compared with the corresponding primary tumors. CONCLUSION: These results suggest that a partial EMT, characterized by the expression of Twist but without a total E-cadherin depletion, is involved in the acquisition of invasive traits by cSCC, but the process is downregulated in lymph node metastases.

MYC copy number gains are associated with poor outcome in penile squamous cell carcinoma.

PURPOSE: We determined MYC gene numerical aberrations and protein expression at different stages of penile squamous cell carcinoma carcinogenesis. We correlated these findings with clinicopathological parameters and HPV infection. MATERIALS AND METHODS: We evaluated 79 cases of penile squamous cell carcinoma, including 11 in situ and 68 invasive carcinomas. The MYC cytogenetic profile was evaluated by fluorescence in situ hybridization. HPV was detected by polymerase chain reaction amplification. RESULTS: MYC gains were identified in 4 of 11 in situ carcinomas (36%) and 50 of 68 invasive penile squamous cell carcinomas (73%). A significant association between MYC gains, and tumor progression and poor outcome was demonstrated (p <0.05). HPV DNA was detected in 32 of 79 penile squamous cell carcinomas (39%). High risk type 16 was the most prevalent type. MYC numerical aberrations did not correlate with HPV status. A significant association between HPV and MYC protein over expression was noted. In HPV negative cases MYC gains correlated with MYC over expression. CONCLUSIONS: MYC gains progressively increased during penile squamous cell carcinoma progression from in situ samples to metastases. MYC gains were an independent factor for poor prognosis. These findings were independent of HPV infection. MYC expression was increased in samples with HPV infection, probably reflecting direct activation of MYC.

D2-40 immunohistochemical overexpression in cutaneous squamous cell carcinomas: a marker of metastatic risk.

BACKGROUND: Approximately 4% of cutaneous squamous cell carcinomas (cSCCs) develop lymphatic metastases. The value of lymphatic endothelial markers to enhance the detection of lymphatic tumor invasion in cSCC has not been assessed previously. OBJECTIVE: We sought to evaluate the use of the antibody D2-40, a podoplanin immunohistochemical marker, to identify tumor lymph vessel invasion in cSCC and to assess its expression in tumor cells. METHODS: This was a retrospective case-control study. A series of 101 cSCC, including 51 cases that developed lymphatic metastatic spread (metastasizing cSCC [MSCC]) and 50 cases that resolved definitely after surgical excision (non-MSCC) were included in the study. Lymph vessel invasion using D2-40 was evaluated on all primary biopsy specimens. The percentage of tumor cells showing D2-40 positivity and intensity scoring were recorded. All the immunohistochemical findings were correlated with the clinicopathological features. RESULTS: Lymph vessel invasion was observed in 8% of non-MSCCs and in 25.5% of MSCCs (P = .031). D2-40 expression was significantly increased, both in intensity (odds ratio 4.42 for intensity ++/+++) and in area (odds ratio 2.29 for area >10%), in MSCC when compared with non-MSCC. Interestingly, almost half (49%) of the MSCC had moderate to intense D2-40 positivity compared with 16% of non-MSCC. D2-40 immunohistochemical expression was increased in tumors with an infiltrative pattern of extension. In the multivariate analysis, histologically poorly differentiated tumors, recurrent lesions, and cSCC showing D2-40 overexpression (in intensity) were significantly associated with lymphatic metastases development (odds ratios 15.67, 14.72, and 6.07, respectively). LIMITATIONS: This was a retrospective study. CONCLUSION: The expression of podoplanin associates with high metastatic risk in cSCC.

Infant de 17 mesos amb plaques eritematodescamatives generalitzades i onicodistròfia / No disponible

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Panniculitis: a cutaneous manifestation of dermatomyositis.

Specific and nonspecific panniculitis sometimes occurs during the course of connective tissue diseases. However, the association of panniculitis and dermatomyositis (DM) has only rarely been described. Previous reports have suggested that panniculitis may be an inherent part of DM but, because the cases reported are few, it is difficult to know the significance of this unusual association. We describe 2 patients with DM in whom panniculitis developed during the course of the disease, and discuss its clinical and diagnostic significance.

PTEN mutations in eight Spanish families and one Brazilian family with Cowden syndrome.

Cowden syndrome is an autosomal dominant genodermatosis, characterized by the presence of multiple hamartomas in the skin, breast, thyroid, gastrointestinal tract, central nervous system, and an increased risk in developing breast and thyroid carcinomas. Over 80 germline mutations of the tumor suppressor gene PTEN, on chromosome 10q23, have been reported in more than 100 unrelated patients and families; however, questions regarding distribution of the mutations in populations from different geographic areas, and phenotypic expression are still unclear. In this study the results are reported of mutation analysis of PTEN in 13 families from Spain and one family of Brazilian origin with Cowden syndrome. PTEN germline mutations were detected in nine of them (64%). Five mutations were located in exon 5, one in exon 6, two in exon 7, and one in exon 8. Four of the mutations were novel. In another case, an identical change had been previously reported as a somatic mutation in an endometrial carcinoma. In one family, the patient presented a de novo mutation, which was not detected in his parents. In five patients, the detection of the PTEN germline mutation confirmed their condition, even in the absence of sufficient criteria to make the clinical diagnosis of Cowden syndrome.