SOX10-Internal Tandem Duplications and PLAG1 or HMGA2 Fusions Segregate Eccrine-Type and Apocrine-Type Cutaneous Mixed Tumors.

Cutaneous mixed tumors exhibit a wide morphologic diversity and are currently classified into apocrine and eccrine types based on their morphologic differentiation. Some cases of apocrine-type cutaneous mixed tumors (ACMT), namely, hyaline cell-rich apocrine cutaneous mixed tumors (HCR-ACMT) show a prominent or exclusive plasmacytoid myoepithelial component. Although recurrent fusions of PLAG1 have been observed in ACMT, the oncogenic driver of eccrine-type cutaneous mixed tumors (ECMT) is still unknown. The aim of the study was to provide a comprehensive morphologic, immunohistochemical, and molecular characterization of these tumors. Forty-one cases were included in this study: 28 cases of ACMT/HCR-ACMT and 13 cases of ECMT. After morphologic and immunohistochemical characterization, all specimens were analyzed by RNA sequencing. By immunohistochemistry, all cases showed expression of SOX10, but only ACMT/HCR-ACMT showed expression of PLAG1 and HMGA2. RNA sequencing confirmed the presence of recurrent fusion of PLAG1 or HMGA2 in all cases of ACMT/HCR-ACMT, with a perfect correlation with PLAG1/HMGA2 immunohistochemical status, and revealed internal tandem duplications of SOX10 (SOX10-ITD) in all cases of ECMT. Although TRPS1::PLAG1 was the most frequent fusion, HMGA2::WIF1 and HMGA2::NFIB were detected in ACMT cases. Clustering analysis based on gene expression profiling of 110 tumors, including numerous histotypes, showed that ECMT formed a distinct group compared with all other tumors. ACMT, HCR-ACMT, and salivary gland pleomorphic adenoma clustered together, whereas myoepithelioma with fusions of EWSR1, FUS, PBX1, PBX3, POU5F1, and KLF17 formed another cluster. Follow-up showed no evidence of disease in 23 cases across all 3 tumor types. In conclusion, our study demonstrated for the first time SOX10-ITD in ECMT and HMGA2 fusions in ACMT and further refined the prevalence of PLAG1 fusions in ACMT. Clustering analyses revealed the transcriptomic distance between these different tumors, especially in the heterogenous group of myoepitheliomas.

Granulocyte microvesicles with a high plasmin generation capacity promote clot lysis and improve outcome in septic shock.

Microvesicles (MVs) have previously been shown to exert profibrinolytic capacity, which is increased in patients with septic shock (SS) with a favorable outcome. We, therefore, hypothesized that the plasmin generation capacity (PGC) could confer to MVs a protective effect supported by their capacity to lyse a thrombus, and we investigated the mechanisms involved. Using an MV-PGC kinetic assay, ELISA, and flow cytometry, we found that granulocyte MVs (Gran-MVs) from SS patients display a heterogeneous PGC profile driven by the uPA (urokinase)/uPAR system. In vitro, these MVs lyse a thrombus according to their MV-PGC levels in a uPA/uPAR-dependent manner, as shown in a fluorescent clot lysis test and a lysis front retraction assay. Fibrinolytic activators conveyed by MVs contribute to approximately 30% of the plasma plasminogenolytic capacity of SS patients. In a murine model of SS, the injection of high PGC Gran-MVs significantly improved mouse survival and reduced the number of thrombi in vital organs. This was associated with a modification of the mouse coagulation and fibrinolysis properties toward a more fibrinolytic profile. Interestingly, mouse survival was not improved when soluble uPA was injected. Finally, using a multiplex array on plasma from SS patients, we found that neutrophil elastase correlates with the effect of high-PGC-capacity plasma and modulates the Gran-MV plasmin generation capacity by cleaving uPA-PAI-1 complexes. In conclusion, we show that the high PGC level displayed by Gran-MVs reduces thrombus formation and improves survival, conferring to Gran-MVs a protective role in a murine model of sepsis.

Gene fusions in poroma, porocarcinoma and related adnexal skin tumours: An update.

Poroma is a benign sweat gland tumour showing morphological features recapitulating the superficial portion of the eccrine sweat coil. A subset of poromas may transform into porocarcinoma, its malignant counterpart. Poroma and porocarcinoma are characterised by recurrent gene fusions involving YAP1, a transcriptional co-activator, which is controlled by the Hippo signalling pathway. The fusion genes frequently involve MAML2 and NUTM1, which are also rearranged in other cutaneous and extracutaneous neoplasms. We aimed to review the clinical, morphological and molecular features of this category of adnexal neoplasms with a special focus upon emerging differential diagnoses, and discuss how their systematic molecular characterisation may contribute to a standardisation of diagnosis, more accurate classification and, ultimately, refinement of their prognosis and therapeutic modalities.

Porocarcinomas with PAK1/2/3 fusions: a series of 12 cases.

AIMS: Porocarcinoma is a malignant sweat gland tumour differentiated toward the upper part of the sweat duct and may arise from the transformation of a preexisting benign poroma. In 2019, Sekine et al. demonstrated the presence of YAP1::MAML2 and YAP1::NUTM1 fusions in most poromas and porocarcinomas. Recently, our group identified PAK2-fusions in a subset of benign poromas. Herein we report a series of 12 porocarcinoma cases harbouring PAK1/2/3 fusions. METHODS AND RESULTS: Five patients were male and the median age was 79 years (ranges: 59-95). Tumours were located on the trunk (n = 7), on the thigh (n = 3), neck (n = 1), or groin area (n = 1). Four patients developed distant metastases. Microscopically, seven cases harboured a benign poroma component and a malignant invasive part. Ductal formations were observed in all, while infundibular/horn cysts and cells with vacuolated cytoplasm were detected in seven and six tumours, respectively. In three cases, the invasive component consisted of a proliferation of elongated cells, some of which formed pseudovascular spaces, whereas the others harboured a predominant solid or trabecular growth pattern. Immunohistochemical staining for CEA and EMA confirmed the presence of ducts. Focal androgen receptor expression was detected in three specimens. Whole RNA sequencing evidenced LAMTOR1::PAK1 (n = 2), ZDHHC5::PAK1 (n = 2), DLG1::PAK2, CTDSP1::PAK1, CTNND1::PAK1, SSR1::PAK3, CTNNA1::PAK2, RNF13::PAK2, ROBO1::PAK2, and CD47::PAK2. Activating mutation of HRAS (G13V, n = 3, G13R, n = 1, Q61L, n = 2) was present in six cases. CONCLUSION: Our study suggests that PAK1/2/3 fusions is the oncogenic driver of a subset of porocarcinomas lacking YAP1 rearrangement.

[Cutaneous adnexal tumours: Development and synthesis of diagnostic fusion genes]. / Tumeurs annexielles cutanées : mise au point et synthèse des gènes de fusion à connaître pour le diagnostic.

Cutaneous adnexal tumours are a heterogeneous group of epithelial lesions that includes tumours with follicular, sudoral and/or sebaceous differentiation, or even several combined lines of differentiation. Over the last few years, molecular analysis of these lesions has allowed to identify specific molecular events responsible for tumour development in an increasing number of tumour types. Like other rare neoplasms, such as soft tissue tumours, adnexal tumours display fusion genes resulting from chromosomal translocations that may be specific for the diagnosis if molecular data are properly integrated in the clinical and morphological setting. Molecular testing of adnexal tumours is valuable as it allows to strengthen the robustness of the diagnosis for a group of tumours displaying a wide morphological spectrum. It has allowed to refine the diagnostic criteria and to develop increasingly specific diagnostic immunostainings. Finally, molecular testing has been responsible for the identification of new entities or morphological subtypes of previously known entities. The aim of this review is to provide an update on cutaneous adnexal tumours associated with fusion genes and to evaluate the impact of molecular data on the diagnosis of these lesions.

Recurrent PAK2 rearrangements in poroma with folliculo-sebaceous differentiation.

AIMS: Poroma is a benign adnexal neoplasm with differentiation towards the upper portion of the sweat gland apparatus. In 2019, Sekine et al. demonstrated recurrent YAP1::MAML2 and YAP1::NUTM1 fusion in poroma and porocarcinoma. Follicular, sebaceous and/or apocrine differentiation has been reported in rare cases of poroma and whether these tumours constitute a variant of poroma or represent a distinctive tumour is a matter to debate. Herein we describe the clinical, immunophenotypic, and molecular features of 13 cases of poroma with folliculo-sebaceous differentiation. METHODS AND RESULTS: Most of the tumours were located on the head and neck region (n = 7), and on the thigh (n = 3). All presented were adults with a slight male predilection. The median tumour size was 10 mm (range: 4-25). Microscopically, lesions displayed features of poroma with nodules of monotonous basophilic cells associated with a second population of larger eosinophilic cells. In all cases, ducts and scattered sebocytes were identified. Infundibular cysts were present in 10 cases. In two cases high mitotic activity was noted, and in three cases cytologic atypia and areas of necrosis were identified. Whole transcriptome RNA sequencing demonstrated in-frame fusion transcripts involving RNF13::PAK2 (n = 4), EPHB3::PAK2 (n = 2), DLG1::PAK2 (n = 2), LRIG1::PAK2 (n = 1), ATP1B3::PAK2 (n = 1), TM9SF4::PAK2 (n = 1), and CTNNA1::PAK2 (n = 1). Moreover, fluorescence in situ hybridisation (FISH) analysis revealed PAK2 rearrangement in an additional case. No YAP1::MAML2 or YAP1::NUTM1 fusion was detected. CONCLUSION: Recurrent fusions involving the PAK2 gene in all analysed poroma with folliculo-sebaceous differentiation in this study confirms that this neoplasm represents a separate tumour entity distinct from YAP1::MAML2 or YAP1::NUTM1 rearranged poromas.

Distinct regulations driving YAP1 expression loss in poroma, porocarcinoma and RB1-deficient skin carcinoma.

AIMS: Recently, YAP1 fusion genes have been demonstrated in eccrine poroma and porocarcinoma, and the diagnostic use of YAP1 immunohistochemistry has been highlighted in this setting. In other organs, loss of YAP1 expression can reflect YAP1 rearrangement or transcriptional repression, notably through RB1 inactivation. In this context, our objective was to re-evaluate the performance of YAP1 immunohistochemistry for the diagnosis of poroma and porocarcinoma. METHODS AND RESULTS: The expression of the C-terminal part of the YAP1 protein was evaluated by immunohistochemistry in 543 cutaneous epithelial tumours, including 27 poromas, 14 porocarcinomas and 502 other cutaneous tumours. Tumours that showed a lack of expression of YAP1 were further investigated for Rb by immunohistochemistry and for fusion transcripts by real-time PCR (YAP1::MAML2 and YAP1::NUTM1). The absence of YAP1 expression was observed in 24 cases of poroma (89%), 10 porocarcinoma (72%), 162 Merkel cell carcinoma (98%), 14 squamous cell carcinoma (SCC) (15%), one trichoblastoma and one sebaceoma. Fusions of YAP1 were detected in only 16 cases of poroma (n = 66%), 10 porocarcinoma (71%) all lacking YAP1 expression, and in one sebaceoma. The loss of Rb expression was detected in all cases except one of YAP1-deficient SCC (n = 14), such tumours showing significant morphological overlap with porocarcinoma. In-vitro experiments in HaCat cells showed that RB1 knockdown resulted in repression of YAP1 protein expression. CONCLUSION: In addition to gene fusion, we report that transcriptional repression of YAP1 can be observed in skin tumours with RB1 inactivation, including MCC and a subset of SCC.

Recurrent FOXK1::GRHL and GPS2::GRHL fusions in trichogerminoma.

We report 21 cases of trichogerminoma harbouring previously undescribed FOXK1::GRHL1/2 or GPS2::GRHL1/2/3 in-frame fusion transcripts. Microscopic examination of a preliminary set of five cases revealed well-delimitated tumours located in the dermis with frequent extension to the subcutaneous tissue. Tumours presented a massive and nodular architecture and consisted of a proliferation of basaloid cells. A biphasic pattern sometime resulting in tumour cell nests ('cell balls') was present. Immunohistochemistry demonstrated the expression of cytokeratins (CKs) 15, 17, and PHLDA1. In addition, numerous CK20-positive Merkel cells were detected. RNA sequencing (RNA-seq) revealed a FOXK1::GRHL1 chimeric transcript in three cases and a FOXK1::GRHL2 fusion in two cases. In a second series for validation (n = 88), FOXK1::GRHL1/2 fusion transcripts were detected by RT-qPCR or FISH in an additional 12 trichogerminomas and not in any other follicular tumour entities or basal cell carcinoma cases (n = 66). Additional RNA-seq analysis in trichogerminoma cases without detected FOXK1::GRHL1/2 rearrangements revealed GPS2::GRHL1 fusion transcripts in two cases, GPS2::GRHL2 in one case, and GPS2::GRHL3 fusion transcript in one case. Therefore, our study strongly suggests that GRHL1/2/3 gene rearrangements might represent the oncogenic driver in trichogerminoma, a subset of follicular tumours characterized by immature features and numerous Merkel cells. © 2022 The Pathological Society of Great Britain and Ireland.

Wholistic approach: Transcriptomic analysis and beyond using archival material for molecular diagnosis.

Many neoplasms remain unclassified after histopathological examination, which requires further molecular analysis. To this regard, mesenchymal neoplasms are particularly challenging due to the combination of their rarity and the large number of subtypes, and many entities still lack robust diagnostic hallmarks. RNA transcriptomic profiles have proven to be a reliable basis for the classification of previously unclassified tumors and notably for mesenchymal neoplasms. Using exome-based RNA capture sequencing on more than 5000 samples of archival material (formalin-fixed, paraffin-embedded), the combination of expression profiles analyzes (including several clustering methods), fusion genes, and small nucleotide variations has been developed at the Centre Léon Bérard (CLB) in Lyon for the molecular diagnosis of challenging neoplasms and the discovery of new entities. The molecular basis of the technique, the protocol, and the bioinformatics algorithms used are described herein, as well as its advantages and limitations.

CIC-DUX4 sarcomas.

PURPOSE OF REVIEW: CIC-DUX4 sarcoma (CDS) is a high-grade undifferentiated round cells sarcoma that belongs to the undifferentiated round cell sarcomas family. It represents less than one percent of sarcomas, defining a rarest among rare malignancies. It affects young adults, displaying soft tissue mass. Considered very aggressive, a high proportion of cases display an advanced disease with lung metastasis at diagnosis. Here we discuss recent progress in molecular characterization of CDS, the main tracks of CDS biology and the current and future prospects of therapeutic approaches. RECENT FINDINGS: CDS is characterized by a specific oncogenic translocation CIC::DUX4 that induce ETV4 overexpression. Patients with CDS show an aggressive clinical course and have a significantly unfavorable outcome compared to Ewing sarcoma. As of today, there is a lack of consensus on whether they should be treated with an Ewing-like approach, as currently done by most sites, or regarded as high-grade soft tissue sarcoma (STS). Anyway, when feasible, combination regimens including anthracycline and alkylating agents should be favored and patients should not benefit from a therapeutic de-escalation. Overall, registration within clinical trials and prospective registries is recommended. SUMMARY: Overall, CDS showed a poor prognosis regardless of the patterns of treatment that warrant biological studies to better understand the disease.

Molecular characterization of fast-growing melanomas.

BACKGROUND: The rate of growth of primary melanoma is a robust predictor of aggressiveness, but the mutational profile of fast-growing melanomas (FGMM) and the potential to stratify patients at high risk of death has not been comprehensively studied. OBJECTIVE: To investigate the epidemiologic, clinical, and mutational profile of primary cutaneous melanomas with a thickness ≥ 1 mm, stratified by rate of growth. METHODS: Observational prospective study. Deep-targeted sequencing of 40 melanoma driver genes on formalin fixed, paraffin-embedded primary melanoma samples. Comparison of FGMM (rate of growth > 0.5 mm/month) and nonFGMM (rate of growth ≤ 0.5 mm/month). RESULTS: Two hundred patients were enrolled, among wom 70 had FGMM. The relapse-free survival was lower in the FGMM group (P = .014). FGMM had a higher number of predicted deleterious mutations within the 40 genes than nonFGMM (P = .033). Ulceration (P = .032), thickness (P = .006), lower sun exposure (P = .049), and fibroblast growth factor receptor 2 (FGFR2) mutations (P = .037) were significantly associated with fast growth. LIMITATIONS: Single-center study, cohort size, potential memory bias, number of investigated genes. CONCLUSION: Fast growth is linked to specific tumor biology and environmental factors. Ulceration, thickness, and FGFR2 mutations are associated with fast growth. Screening for FGFR2 mutations might provide an additional tool to better identify FGMM, which are probably good candidates for adjuvant therapies.

[Soft tissue tumours with FN1 (Fibronectin 1) fusion gene]. / Tumeurs des tissus mous avec fusion du gène FN1 (Fibronectine 1).

Translocations involving FN1 gene have been described in several tumours, which share the presence of a cartilaginous matrix with or without calcifications and a good prognosis. They encompass: soft tissue chondroma, synovial chondromatosis, calcifying aponeurotic fibroma, phosphaturic mesenchymal tumour and a new spectrum of tumours: "the calcified chondroid mesenchymal neoplasms". We review all the clinical, histopathological and molecular data of these tumours and discuss the differential diagnoses.

[BRAF mutation evolution in melanoma: Myth or reality?] / Évolution du statut braf dans le melanome : mythe ou Réalité ?

Knowledge of the BRAF mutational status has become essential for melanoma therapeutic management. B-Raf inhibitors are associated with significant overall survival in patients with BRAFV600-mutated metastatic melanoma. Although the BRAF mutation appears to be an early and driver mutation, some authors hypothesized that its expression was not stable during melanoma progression, suggesting a molecular heterogeneity. This argument is often used to explain discrepancy in molecular status among patients with melanoma, discrepancies that we occasionally met during our practice. We retrospectively compared BRAF mutational status on matched melanoma samples (primary & metastatic lesions), thus 150 samples from 56 patients were analysed through immunohistochemistry anti-BRAF, PCR-HRM and Sanger sequencing, Next Generation Sequencing (NGS) and digital PCR. Seven cases presented an apparent tumor heterogeneity. The analysis of these discrepancies by a technique of increasing sensitivity made it possible to identify 1 false-negative result for the immunohistochemistry, 1 false-negative result for the NGS sequencing and 5 (3%) false-negative results by PCR-HRM SANGER. Our results are consistent with the most recent data, demonstrating the stability of the BRAF mutation during the course of melanoma. Immunohistochemistry shows excellent sensitivity for detecting the main BRAF mutation. In our study, the mutational heterogeneity was actually misleading, a result of imperfect sensitivity of some older molecular approaches.

Acral FibroChondroMyxoid tumor: imaging features of a new entity.

"Acral FibroChondroMyxoid tumor" (AFCMT) is a recently described distinctive subtype of acral soft tissue tumor that typically arises on the fingers and toes. We herein present the unreported imaging features of AFCMT in a 44-year-old woman. This otherwise healthy patient was referred for a painful, slow-growing, soft tissue mass in the middle finger of her right hand. Initial radiographs and computed tomography showed a small lesion centered in the soft tissue of the ulnar aspect of the proximal phalanx, associated with scalloping of the underlying bone. Magnetic resonance imaging confirmed the presence of a well-circumscribed soft tissue tumor that exhibited relatively high T2-weighted signal intensity and marked enhancement after contrast administration. Subsequent excisional biopsy was performed. Histologically, the tumor was characterized by an abundant stroma displaying fibrous, chondroid, and myxoid areas. By immunohistochemistry, tumor cells stained for CD34, ERG, and focally S100 protein. RNA-sequencing allowed detection of THBS1-ADGFR5 gene fusion which confirmed the diagnosis of AFCMT. At 2-year follow-up, the patient remains free of recurrence. AFCMT is a previously unrecognized entity that may mimic chondroma and should be considered in the differential diagnosis of soft tissue tumors with cartilaginous or myxoid stroma in the extremities.

[Point of view: A lesson from lockdown, histopathology through videoconferencing]. / Point de vue : une leçon tirée du confinement, l'histopathologie en visioconférence.

The use of videoconferencing had increased significantly during lockdown. During this period, videoconferencing has been used in the pathological department of pathology (Timone university hospital, Marseille, France) for academic, diagnosis and referral. We provide our point of view regarding the use of this tool. As discussing slides through videoconferencing is a new and specific activity, we have also summarised specific recommendations for practical remote histopathology meetings.

Recurrent novel THBS1-ADGRF5 gene fusion in a new tumor subtype "Acral FibroChondroMyxoid Tumors".

Acral soft tissue tumors are common neoplasms, a subset of which pose a diagnostic challenge. We report 10 cases of a previously unrecognized acral benign soft tissue tumor. These tumors arose on the fingers and toes and involved bone in half of cases. Histologically, the tumors were lobulated and displayed an abundant stroma made of variable fibrous, chondroid and myxoid material reminiscent of cartilaginous or myoepithelial differentiation. Tumor cells harbored small round to reniform nuclei with clear chromatin and inconspicuous nucleoli along with scant eosinophilic cytoplasm. The cells were mostly arranged haphazardly in the stroma but also in small clusters. No mitotic activity was detected. No specific feature was identified in recurrent cases. By immunohistochemistry, the cells consistently stained for CD34 (10/10), ERG (9/10), and SOX9 (7/10). Whole RNA sequencing identified a previously undescribed recurrent in frame THBS1-ADGRF5 gene fusion in all cases. The transcript was confirmed by RT-PCR and was not found in the control group of mimickers including soft tissue chondromas. We propose the name of Acral FibroChondroMyxoid Tumors for this new entity.

Early Phase of Primary Melanoma Growth from the Patient Point of view: A Prospective Cross Sectional Study on Melanoma over 1 mm in Thickness.

A Nodular Pink Lesion with an Uncommon Diagnosis: A QuizMost melanomas are diagnosed by the patients themselves or by their partners or relatives; they alone can describe its history. We designed a prospective cross-sectional study to describe patients' perception of morphology, growth pattern and kinetics of their primary melanoma over 1 mm in thickness before resection. Patients were interviewed with a questionnaire, a grid representing 9 possible scenarios of melanoma growth, and a set of 87 photographs of potential aspects of melanomas and precursors. Most patients were able to describe the growth of their melanoma and select pictures representative of its successive aspects before resection. Among 453 patients, 60% reported a preexisting lesion present for years. Growth pattern scenarios concurred with tumor kinetics but with no statistical difference between nodular and superficial spreading subtypes. These subjective patient-reported indicators about melanoma growth over time could dynamically complement its objective pathological analysis otherwise static at a single time point.

[Glomerulopathy associated with lecithin-cholesterol-acyltransferase deficiency: A case report and literature review]. / Glomérulopathie associée à un déficit en lécithine-cholestérol-acyltransférase : rapport de cas et revue de la littérature.

Glomerulopathy associated with lecithin-cholesterol-acyltransferase deficiency (LCAT) is a rare automosal recessive disease. Acquired LCAT deficiency due to inhibitory autoantibodies against LCAT are also described. This disease is induced by systemic deposits related to a lipid metabolism disorder and lead to multi-organ involvement including renal involvement. Lipid profile usually shows variable cholesterol levels but very low HDL levels. Here we describe the case of a 33-year-old man presenting a nephrotic syndrome associated with moderate renal insufficiency for which the pathological analysis allowed to guide towards the diagnosis of LCAT deficiency. Laboratory and genetic data confirmed this diagnosis. Familial history and lipid profile abnormalities are important in the identification of this disease.

[Diffuse esophageal leiomyomatosis and Alport's syndrome: A case report and review of the literature]. / Léiomyomatose œsophagienne diffuse et syndrome d'Alport : rapport d'un cas.

Diffuse esophageal leiomyomatosis is a rare esophageal tumor characterized by circumferential thickening of smooth muscle layers. Diffuse esophageal leiomyomatosis can be associated with Alport's syndrome and therefore diagnosed by skin biopsy. Alport syndrome is a hereditary disease usually defined by the association of glomerular nephropathy and perceptual deafness. Here we describe the management of a young women with a diffuse esophageal leiomyomatosis and a past history of uterine leiomyoma. The surgical treatment depends on the esophageal extent of the disease. Association between diffuse esophageal leiomyomatosis and early uterine leiomyomas could be also observed and leading to Alport's syndrome diagnosis despite the absence of renal abnormalities.