PRAME expression in 137 primary cutaneous melanomas and comparison with 38 related metastases.

Melanocytic lesions have always represented a diagnostic challenge for surgical pathologists. According to the literature, PRAME showed great promise as an immunohistochemical tool in the distinction between benign and malignant melanocytic lesions. In the present study, we retrospectively analyzed 137 thicker (Breslow > 1 mm) primary cutaneous melanomas with the aim to better understand the utility of PRAME immunohistochemistry in daily practice and also to investigate if PRAME could represent a prognostic biomarker for cutaneous melanomas. PRAME immunohistochemistry was performed in all melanomas and in the metastases with antibodies to PRAME (dilution 1:1000, clone Ab219650) on an automated immunostainer (Ventana Benchmark Ultra) using a brown chromogen (DAB). We found that melanomas (59.1%) show diffuse PRAME expression (score 4 +). 99 (72.3%) primary cutaneous melanoma had no relapse during the follow-up. Of this group of melanomas, 61/99 (61.6%) were diffusely positive for PRAME. 38 (27.7%) primary cutaneous melanoma had relapses. Of this group, 28/36 (77.7%) were diffusely positive. We did not find any statistical correlation between diffuse PRAME expression and the presence of driver mutation in BRAF gene (p = 0.927), NRAS gene (p = 0.496) or either of the two (p = 0.138). We did not find a prognostic significance of diffuse PRAME expression for relapse (p = 0.462) or survival rate (p = 0.245). The prognostic value of PRAME has been only reported in mucosal, uveal and cutaneous thin melanomas. Here, we show statistical analyses on PRAME expression for melanoma with Breslow > 1 mm based on survival rate and long-term follow-up. According to our results, PRAME is a useful immunohistochemical ancillary tool in daily practice diagnosis of melanocytic lesions.

GRM1 Gene Fusions as an Alternative Molecular Driver in Blue Nevi and Related Melanomas.

Activating mutations in GNAQ, GNA11, CYSLTR2, and PLCB4 genes are regarded as the main oncogenic drivers of blue nevi (BN) and blue malignant melanocytic tumors. Here we report 4 cases of blue melanocytic neoplasms devoid of these mutations but harboring GRM1 gene fusions. In this short series, there was no gender predominance (sex ratio, 1). The mean age at diagnosis was 40 years (range, 12-72). Tumors were located on the face (n = 2), forearm (n = 1), and dorsum of the foot (n = 1). Clinically, a plaque-like pre-existing BN was found in 2 cases, including a deep location; another case presented as an Ota nevus. Two cases were diagnosed as melanoma ex-BN, one as an atypical BN, and one as a plaque-like BN. Microscopic examination revealed a dermal proliferation of dendritic melanocytes in a sclerotic stroma. A dermal cellular nodule with atypia and mitotic activity was observed in 3 cases. Genetic investigation by whole exome RNA sequencing revealed MYO10::GRM1 (n = 2) and ZEB2::GRM1 (n = 1) fusions. A GRM1 rearrangement was identified by fluorescence in situ hybridization in the remaining case. SF3B1 comutations were present in the 2 melanomas, and both had a MYO10::GRM1 fusion. Array comparative genomic hybridization was feasible for 3 cases and displayed multiple copy number alterations in the 2 melanomas and limited copy number alterations in the atypical BN, all genomic profiles compatible with those of classical blue lesions. GRM1 was overexpressed in all cases compared with a control group of blue lesions with other typical mutations. Both melanomas rapidly developed visceral metastases following diagnosis, with a fatal outcome in one case and tumor progression under palliative care in the other. These data suggest that GRM1 gene fusions could represent an additional rare oncogenic driver in the setting of BN, mutually exclusive of classical canonical mutations, especially in plaque-type or Ota subtypes.

Granulysin, a novel marker for extranodal NK/T cell lymphoma, nasal type.

Granulysin is a cytolytic protein expressed in cytotoxic T and natural killer (NK) cells. Abnormal serum levels of granulysin in lymphomas with NK and cytotoxic phenotype have been shown to correlate with tumour progression. In this study, we investigated the expression pattern of granulysin in routine sections of normal and reactive lymphoid tissues as well as in a large series of lymphomas. In normal tissues, granulysin labelled a small population of cells that double immunostaining revealed to belong to the pool of cytotoxic T/NK cells. Among lymphoid neoplasms, the highest expression of granulysin (71%) was found in extranodal NK/T cell lymphomas of nasal type (ENKTL). To note is that 29% of ENKTLs, which were negative for one or more of classical cytotoxic markers strongly expressed granulysin. Furthermore, expression of granulysin was observed in rare cases of T cell lymphomas with a cytotoxic phenotype (i.e. ALK-negative anaplastic large cell lymphoma (26%), enteropathy-associated T cell lymphoma (12%) and peripheral T cell lymphoma, NOS (4%)). None of the investigated non-Hodgkin B cell lymphomas, Hodgkin lymphoma and plasma cell myeloma were granulysin positive. The results suggest granulysin as a novel marker for a subset of cytotoxic NK cell derived malignancies and its usefulness is highlighted in those ENKTLs that lack expression of other cytotoxic markers but retain granulysin expression.

Unveiling Another Missing Piece in EBV-Driven Lymphomagenesis: EBV-Encoded MicroRNAs Expression in EBER-Negative Burkitt Lymphoma Cases.

Epstein-Barr virus (EBV) is a gammaherpesvirus linked to a number of lymphoid and epithelial malignancies, including Burkitt lymphoma (BL) in which its frequency ranges from 30% in sporadic cases to 100% in the endemic ones. The possible contribution of EBV to BL pathogenesis is largely unknown. It has been suggested that EBV may be associated with all of the cases, including those diagnosed as EBV negative by a mechanism of hit-and-run. Early during oncogenesis, viral genes are essential for initiating disease. Progressively, viral genome is lost to escape the immune system and host mutations accumulate in proto-oncogenic cell. The main problem with the hit-and-run hypothesis is the lack of evidence in primary tumors. The routine methods applied to detect the virus [i.e., immunohistochemistry and EBV-encoded RNAs (EBER) in situ hybridization (ISH)] have a low specificity and accuracy. The aim of this study was to identify the most suitable method to detect EBV infection in pathology samples by applying conventional and non-conventional methods (i.e., EBV-microRNAs detection and EBV viral load measurement). We investigated a total of 10 cases and we found that all the samples (n = 6) diagnosed as EBV negative by immunohistochemistry and EBER-ISH demonstrated the presence of EBV-microRNAs and EBV genome. This points at the possibility that EBV might have contributed to lymphomagenesis in all our patients, and propose microRNAs detection as the most specific and sensitive tool to recognize EBV vestiges. It is worth noting that our data would have considerable implications for EBV-related diseases control. By using anti-EBV vaccines, one could potentially prevent also some cancers less suspected of a viral origin because of viral genome loss.

MicroRNAs sequencing unveils distinct molecular subgroups of plasmablastic lymphoma.

Plasmablastic lymphoma (PBL) is an aggressive lymphoma, often arising in the context of immunodeficiency and associated with Epstein-Barr virus (EBV) infection. The most frequently detected genetic alteration is the deregulation of MYC gene through the translocation - t(8;14)(q24;q32). The diagnosis of PBL is often challenging because it has an overlap in morphology, immunophenotype, cytogenetics and virus association with other lymphomas and plasma cell neoplasms; further, its molecular basis remains elusive. In the present study we aimed to better define the possible contribution of EBV infection as well as miRNA deregulation in PBL pathogenesis. We studied 23 cases of PBL, 19 Burkitt lymphomas (BL), and 17 extra-medullary plasmacytoma (EMPC). We used qPCR and immunohistochemistry to assess EBV latency patterns, while micro-RNA (miRNA) profiling was performed by next generation sequencing (Illumina) and validated by qPCR. Our analysis revealed a non-canonical EBV latency program with the partial expression of some proteins characterizing latency II and the activation of an abortive lytic cycle. Moreover, we identified miRNA signatures discriminating PBL from BL and EMPC. Interestingly, based on the miRNA profile, PBL appeared constituted by two discrete subgroups more similar to either BL or EMPC, respectively. This pattern was confirmed in an independent set of cases studied by qPCR and corresponded to different clinico-pathological features in the two groups, including HIV infection, MYC rearrangement and disease localization. In conclusion, we uncovered for the first time 1) an atypical EBV latency program in PBL; 2) a miRNA signature distinguishing PBL from the closest malignant counterparts; 3) the molecular basis of PBL heterogeneity.

Amphicrine carcinoma of the ampullary region.

Amphicrine carcinoma is a peculiar tumor in which the cells have both exocrine and neuroendocrine differentiation, with mucus and neuroendocrine granules within the cytoplasm. In the 2010 WHO classification of tumors of the digestive tract, they have been included in the intermediate-grade malignant category of mixed adenoneuroendocrine carcinomas (MANECs). These tumors are extremely rare in the gastrointestinal tract. Four cases have been reported in the stomach, three in the pancreas, and one in the liver. To the best of our knowledge, this report describes the first case of amphicrine carcinoma in the ampullary region.