Arnaud de la Fouchardière

ZEB1-mediated melanoma cell plasticity enhances resistance to MAPK inhibitors

Targeted therapies with MAPK inhibitors (MAPKi) are faced with severe problems of resistance in BRAF‐mutant melanoma. In parallel to the acquisition of genetic mutations, melanoma cells may also adapt to the drugs through phenotype switching. The ZEB1 transcription factor, a known inducer of EMT and invasiveness, is now considered as a genuine oncogenic factor required for tumor initiation, cancer cell plasticity, and drug resistance in carcinomas. Here, we show that high levels of ZEB1 expression are associated with inherent resistance to MAPKi in BRAFV600‐mutated cell lines and tumors. ZEB1 levels are also elevated in melanoma cells with acquired resistance and in biopsies from patients relapsing while under treatment. ZEB1 overexpression is sufficient to drive the emergence of resistance to MAPKi by promoting a reversible transition toward a MITFlow/p75high stem‐like and tumorigenic phenotype. ZEB1 inhibition promotes cell differentiation, prevents tumorigenic growth in vivo, sensitizes naive melanoma cells to MAPKi, and induces cell death in resistant cells. Overall, our results demonstrate that ZEB1 is a major driver of melanoma cell plasticity, driving drug adaptation and phenotypic resistance to MAPKi.

Melanomas Associated With Blue Nevi or Mimicking Cellular Blue Nevi: Clinical, Pathologic, and Molecular Study of 11 Cases Displaying a High Frequency of GNA11 Mutations, BAP1 Expression Loss, and a Predilection for the Scalp.

Melanomas associated with blue nevi (MABN) or mimicking cellular blue nevi (MMCBN) represent exceptional variants of malignant cutaneous melanocytic tumors. Uveal and leptomeningeal melanomas frequently have somatic mutations of GNAQ or GNA11, which are believed to be early driver mutations. In uveal melanomas, monosomy 3, linked to the BAP1 gene, is an adverse prognostic factor. We have studied the clinical, histologic, BAP1 expression profile, and molecular data of 11 cases of MABN/MMCBN and 24 cellular blue nevi. Most of the cases of MABN/MMCBN occurred on the scalps of adult patients and presented as rapidly growing nodules, typically >1 cm, often arising at the site of a preexisting melanocytic lesion. The MABN/MMCBN were composed of dense nests of large dermal atypical melanocytes, in some cases lying adjacent to a blue nevus. Four patients developed metastatic disease, and 2 died from their disease. A GNA11 mutation was found in 8/11 cases and a GNAQ mutation in 1 case. Seven of 11 cases showed loss of nuclear BAP1 immunohistochemical (IHC) expression in the malignant component, sparing the adjacent nevus. Array comparative genomic hybridization revealed recurrent deletions of chromosomes 1p, 3p, 4q, 6q, 8p, 16q, and 17q and recurrent gains of chromosomes 6p, 8q, and 21q. The 24 cases of cellular blue nevi frequently occurred on the sacrum, had GNAQ mutations, and showed normal positive IHC staining for BAP1. These results underscore overlapping features in all blue-like malignant melanocytic tumors. Loss of BAP1 IHC expression was restricted to melanomas, including all metastatic cases.

Primary leptomeningeal melanoma is part of the BAP1-related cancer syndrome.

In this case report, a 40-year-old woman, with a recent history of seizures, was diagnosed with a supratentorial brain mass. Surgical exploration revealed a heavily pigmented, 5-cm mass adhered to the leptomeninges. Pathological analysis and immunohistochemistry confirmed a malignant melanoma. Due to the absence of a cutaneous or ocular melanoma, a diagnosis of primary leptomeningeal melanoma was made. A few months after this diagnosis, the patient developed a papillary thyroid carcinoma. One year later, the melanoma relapsed. Excision with complementary radiotherapy was performed. Four months after this operation, the patient relapsed again, leading to a progressive, strictly intracranial, multifocal diffusion of the tumor. The patient died from intracranial hypertension related to tumor compression 26 months after the initial surgery. Familial anamnesis revealed a medical history of The BRCA1-associated Protein 1 (BAP1) gene encodes a ubiquitin hydrolase that plays a role in several key cellular processes, including transcriptional regulation, growth, DNA damage response, and chromatin dynamics [1]. Heterozygous germline BAP1 mutations were initially associated with a higher risk of mesothelioma and uveal melanoma, as well as multiple atypical cutaneous melanocytic tumors. However, recent reports have extended their clinical phenotype to others tumors such as clear cell renal cancer, lung adenocarcinoma, meningioma, paraganglioma, and multiple basal cell carcinomas [1, 4].

The sum of gains and losses of genes encoding the protein tyrosine kinase targets predicts response to multi-kinase inhibitor treatment: Characterization, validation, and prognostic value.

Validated predictive biomarkers for multi-tyrosine kinase inhibitors (MTKI) efficacy are lacking. We hypothesized that interindividual response variability is partially dependent on somatic DNA copy number alterations (SCNAs), particularly those of genes encoding the protein tyrosines targeted by MTKI (called target genes). Genomic alterations were investigated in MTKI responsive and non responsive patients with different histological subtypes included in the ProfiLER protocol (NCT 01774409). From March 2013 to August 2014, 58 patients with advanced cancer treated with one of 7 MTKIs were included in the ProfiLER trial and split into one discovery cohort (n = 13), and 2 validation cohorts (n = 12 and 33). An analysis of the copy number alterations of kinase-coding genes for each of 7 MTKIs was conducted. A prediction algorithm (SUMSCAN) based on the presence of specific gene gains (Tumor Target Charge, TTC) and losses (Tumor Target Losses, TTL) was conceived and validated in 2 independent validation cohorts. MTKI sensitive tumors present a characteristic SCNA profile including a global gain profile, and specific gains for target genes while MTKI resistant tumors present the opposite. SUMSCAN favorable patients achieved longer progression-free and overall survival. This work shows that the copy number sum of kinase-coding genes enables the prediction of response of cancer patients to MTKI, opening a novel paradigm for the treatment selection of these patients.

Mutated and amplified NRAS in a subset of cutaneous melanocytic lesions with dermal spitzoid morphology: report of two pediatric cases located on the ear

Extensive cytogenetic testing is slowly unveiling the complexity of the genomics of melanocytic tumors. NRAS mutations have been the first genetic abnormality described in malignant melanomas. We report the cases of two children, presenting a melanocytic lesion located on the ear. One appeared as a combined dermal clone inside a congenital nevus and the other as a centimetric purely dermal tumor. Both tumors were composed of spindled spitzoid melanocytes with atypical histologic features. aCGH and FISH revealed an amplification of the NRAS gene. Sequencing showed an exon 3 NRAS mutation. In the combined case, the amplification was limited to the spitzoid component, underscoring a possible phenotypic shift induced by the alteration. Similarly an overexpression of CyclinD1 and elevation of ki‐67 was found in the spitzoid component confirming a raise in proliferation.

Lymphomes B cutanés primitifs : corrélations anatomo-cliniques dans une série de 44 cas

Resume Objectifs etude des relations histologiques et cliniques de 44 cas de lymphome B cutane primitif, classes selon les criteres de la classification OMS de 2001. Materiel et methodes l’analyse histologique, immunologique et moleculaire a ete correlee aux donnees cliniques. Resultats 33 cas (75%) etaient des lymphomes B de la zone marginale (LZM), localises principalement au tronc ou la face (âge median 54 ans). Une recidive dans un autre site muqueux a ete observee dans 3 cas. L’evolution a ete le plus souvent favorable. Une transformation histologique a necessite un traitement agressif dans 3 cas. 9 (20%) lymphomes diffus a grandes cellules B (LDGCB) se presentaient comme un nodule unique, avec une predominance feminine (âge median 74 ans). Quatre patients âges (> 70 ans) sont decedes de leur lymphome (un seul localise au membre inferieur). Des aspects morphologiques agressifs etaient presents dans 3 cas mortels. 2 (5 %) lymphomes folliculaires (LF) de grade 3 de l’OMS avaient un phenotype heterogene, une localisation faciale, des rechutes cutanees et un bon pronostic. Conclusion le LZM, type predominant, possede un bon pronostic bien qu’une transformation puisse survenir, necessitant un traitement plus agressif. Le LDGCB survient chez des sujets âges et possede des facteurs pronostiques cliniques et histologiques identiques aux lymphomes extra-cutanes. Le LF est rare, son phenotype heterogene, et ses relations avec son equivalent ganglionnaire restent obscures.

Primary Melanoma of the Leptomeninges with BAP1 Expression-Loss in the Setting of a Nevus of Ota: A Clinical, Morphological and Genetic Study of 2 Cases.

1 Department of Biopathology, University Hospital, 80 av Augustin Fliche, 34295 Montpellier Cedex 5, France. 2 Department of Biopathology, La Source Hospital, 1 rue Porte Madeleine 45000 Orl eans, France. 3 Department of Dermatology, La Source Hospital, 1 rue Porte Madeleine 45000 Orl eans, France. 4 Department of Neurosurgery, La Source Hospital, 1 rue Porte Madeleine 45000 Orl eans, France. 5 Department of Biopathology, Dupuytren University Hospital, 2 Avenue Martin Luther King, 87000 Limoges, France. 6 Department of Biopathology, Centre L eon B erard, 28 rue Laennec 69008 Lyon, France.

Combined cutaneous tumors with a melanoma component: A clinical, histologic, and molecular study

BACKGROUND The histogenesis and clinical behavior of combined cutaneous tumors (CCTs) in which the mesenchymal component consists of melanoma remain unclear. OBJECTIVE We sought to characterize the clinical, histologic, and molecular findings in CCTs with an epithelial and a melanoma component. METHODS We retrospectively reviewed the records from 2 institutions for CCTs. Fluorescence in situ hybridization was performed to assess chromosomal copy number alterations in both components. RESULTS Sixteen CCTs were included. The most common subtype was the squamomelanocytic tumor (11), followed by the basomelanocytic tumor (3) and the trichoblastomelanoma (2). CCTs were more common in men (87%), on the head and neck (57%), and had extensive solar elastosis (81%). The median follow-up was 25 months (range, 8-167 months). One case had an adverse outcome. Fluorescence in situ hybridization revealed chromosomal alterations in approximately 55% of the cases. Five cases showed chromosomal gains only in the melanocytic component. One case showed 11q13 gains in both the epithelial and melanocytic components. LIMITATIONS Our study is retrospective and the sample is small. CONCLUSIONS The low incidence of adverse outcomes suggests that CCT may be more indolent than noncombined tumors. 11q13 amplification in both components supports the theory of dual differentiation from a common progenitor cell.

Cutaneous Melanocytoma With : Report of 5 Cases Resembling Clear Cell Sarcoma crtc1-trim11 : Report of 5 Cases Resembling Clear Cell Sarcoma Fusion: Report of 5 Cases Resembling Clear Cell Sarcoma

We report 5 cases of primary intradermal nodular unpigmented tumors with a melanocytic immunophenotype associated with a novel CRTC1-TRIM11 fusion. Clinically, the cutaneous nodules were slowly growing in 3 women and 2 men (25 to 82 y old, median, 28 y) with no specific topography. Lesion size ranged from 4 to 12 mm (median, 5 mm). The tumors were strictly located in the dermis with a nodular pattern. The cells were arranged in confluent nests and fascicules. Central fibronecrotic areas were present in 2 cases. Cells were medium to large, sometimes multinucleated, and presented a spindled and epithelioid cytology with prominent nucleoli. Cytonuclear atypia was constant, and mitotic activity in hotspot areas ranged from 1 to 5/mm². Immunohistochemistry found a constant positivity with S100, MiTF, and Sox10, and a heterogenous staining by MelanA or HMB45. NTRK1 was strongly positive in 3 cases. In all cases, RNA sequencing found an invariable CRTC1(e1)-TRIM11(e2) fusion, confirmed by fluorescent in situ hybridization techniques with a TRIM11 break-apart probe. In 4/4 cases, nuclear TRIM11 expression was positive by immunohistochemistry. Fluorescent in situ hybridization techniques showed no rearrangement of NTRK1 or EWSR1, and array-comparative genomic hybridization displayed no alteration (1 case) or only a whole chromosome 7 gain (2 cases) when performed. No relapse or metastatic event was observed during follow-up [3 to 72 months (median, 14 mo)]. Cutaneous clear cell sarcoma was the main differential diagnosis. Overlapping morphologic features previously described in primary dermal melanomas and paraganglioma-like melanocytic tumors were present. The CRTC1-TRIM11 fusion appears to be specific of an unpigmented nodular tumor combining a melanocytic phenotype and low-grade tumor behavior.

Tetraspanin 8 is a novel regulator of ILK-driven β1 integrin adhesion and signaling in invasive melanoma cells

Melanoma is well known for its propensity for lethal metastasis and resistance to most current therapies. Tumor progression and drug resistance depend to a large extent on the interplay between tumor cells and the surrounding matrix. We previously identified Tetraspanin 8 (Tspan8) as a critical mediator of melanoma invasion, whose expression is absent in healthy skin. The present study investigated whether Tspan8 may influence cell-matrix anchorage and regulate downstream molecular pathways leading to an aggressive behavior. Using silencing and ectopic expression strategies, we showed that Tspan8-mediated invasion of melanoma cells resulted from defects in cell-matrix anchorage by interacting with β1 integrins and by interfering with their clustering, without affecting their surface or global expression levels. These effects were associated with impaired phosphorylation of integrin-linked kinase (ILK) and its downstream target Akt-S473, but not FAK. Specific blockade of Akt or ILK activity strongly affected cell-matrix adhesion. Moreover, expression of a dominant-negative form of ILK reduced β1 integrin clustering and cell-matrix adhesion. Finally, we observed a tumor-promoting effect of Tspan8 in vivo and a mutually exclusive expression pattern between Tspan8 and phosphorylated ILK in melanoma xenografts and human melanocytic lesions. Altogether, the in vitro, in vivo and in situ data highlight a novel regulatory role for Tspan8 in melanoma progression by modulating cell-matrix interactions through β1 integrin-ILK axis and establish Tspan8 as a negative regulator of ILK activity. These findings emphasize the importance of targeting Tspan8 as a means of switching from low- to firm-adhesive states, mandatory to prevent tumor dissemination.