We expanded and extended our studies of Pediatric-type follicular lymphoma (PTFL), which were reported last year. Recently, recurrent genetic alterations of potential importance for its pathogenesis that disrupt pathways associated with the germinal center reaction (TNFRSF14, IRF8), immune escape (TNFRSF14) and anti-apoptosis (MAP2K1) have been described. In an attempt to shed more light onto the pathogenesis of PTFL, an integrative analysis of these mutations was undertaken in a large cohort of 43 cases previously characterized by targeted next generation sequencing and copy number array. Mutations in MAP2K1 were found in 49% (20/41) of the cases, second in frequency to TNFRSF14 alterations (22/41; 54%), and all together were present in 81% of the cases. Immunohistochemical analysis of the MAP2K1 downstream target ERK demonstrated its phosphorylation in the evaluable cases and revealed a good correlation with the allelic frequency of the MAP2K1 mutation. The IRF8 p.K66R mutation was present in 15% (6/39) of the cases and was concomitant with TNFRSF14 mutations in four cases. This hot-spot seems to be highly characteristic for PTFL. In conclusion, TNFRSF14 and MAP2K1 mutations are the most frequent genetic alterations found in PTFL and occur independently in most cases, suggesting that both mutations might play an important role in PTFL lymphomagenesis. We have had a long-standing interest in clarifying the classification of T-cell neoplasms, incorporating traditional pathological and genomic approaches. Expanding on prior work on hepatosplenic T-cell lymphomas, in a large collaborative effort, the genetic basis of this disease was more fully elucidated. Through whole-exome sequencing of 68 HSTLs, recurrently mutated driver genes and copy-number alterations in the disease were identified. Chromatin-modifying genes, including SETD2, INO80, and ARID1B, were commonly mutated in HSTL, affecting 62% of cases. HSTLs manifest frequent mutations in STAT5B (31%), STAT3 (9%), and PIK3CD (9%), for which there currently exist potential targeted therapies. In addition, less frequent events in EZH2, KRAS, and TP53 were described. SETD2 was the most frequently silenced gene in HSTL. SETD2 was shown to act as a tumor suppressor gene. In addition, mutations in STAT5B and PIK3CD activate critical signaling pathways important to cell survival in HSTL.The work thus defines the genetic landscape of HSTL and implicates gene mutations linked to HSTL pathogenesis and potential treatment targets. Primary intestinal T-cell lymphomas (ITCL) comprise mainly enteropathy-associated T-cell lymphomas (EATL). EATL is an aggressive, rare, lymphoma, which represents approximately 5% of mature T-cell lymphomas. Two subtypes are recognized based on distinct morphology, immunophenotype and epidemiology. EATL type I (EATL I) is more common in Western countries, is highly associated with celiac disease (CD), and shows a phenotype akin to that of the majority normal alpha-beta intraepithelial lymphocytes (IEL). EATL type II (EATL II), is more frequent in Asia, is uncommon in patients with CD, and is usually derived from mature activated cytotoxic gamma-delta T-cells. The mechanisms and genetic aberrations responsible for malignant transformation are largely unknown, due to the rarity of these lymphomas. Thirty-four ITCL with formalin-fixed paraffin-embedded tissue were analyzed using a targeted next generation sequencing strategy for mutations in 38 genes. These included genes previously reported to be mutated in T-cell lymphomas, components of the JAK/STAT pathway, and selected genes involved in T-cell receptor signaling and proliferation. Thirty-one and thirty-three samples, respectively, were also tested for mutations within JAK1 codon 1097, and GNAI2 codons 179 and 182 by targeted pyrosequencing, as these recently described mutational hotspots were not covered in the NGS panel. A total of 49 mutations were identified in the 34 ITCL cases, including 46 nonsynonymous single nucleotide variants and 3 deletions. All mutations were predicted to be deleterious based on computational algorithms. The most common alterations involved members of JAK/STAT pathway found in 67.6% of cases, followed by RAS pathway gene alterations in 24.2% of cases. Less common mutations included TET2 (12.1%), EZH2, FYN, NOTCH1 and CD247 (3% each). Other mutations previously reported in T-cell lymphoma subtypes or in other JAK/STAT pathway genes including IDH2, DNMT3A, RHOA, GNB1, PLCG1, CCR4, JAK2, IL7R, and CD130 (IL6ST) were not detected. GNAI2 mutations were not detected in 33 cases studied, including 20 EATL II cases. Within the JAK/STAT cascade, STAT5B and JAK3 were the most frequently mutated genes present in 26.5% and 27.3% of cases, respectively. JAK/STAT pathway mutations were found in all ITCL subtypes, regardless of alpha-beta or gamma-delta origin. These were present in 50% of EATL I (5/10), 80% of EATL II (16/20) and 50% (2/4) of PTCL-NOS cases. The occurrence of such a high frequency of JAK1/3, STAT3/5 mutations and of RAS/RAF mutations might suggest that all intestinal T-cell lymphomas arise, in part, through subversion of cytokine signaling pathways, which are critical for the development and homeostasis of normal alpha-beta and gamma-delta intestinal T-cells. Further studies examined another rare tumor, with relevance ot the classification of T-cell and NK-cell neolasms, aggressive NK-cell leukemia/ lymphoma. Aggressive natural killer cell leukemia (ANKL) is a systemic NK-cell neoplasm, almost always associated with Epstein-Barr virus (EBV). Rare cases of EBV-negative ANKL have been described, and some reports suggested more indolent behavior. We reported the clinicopathologic, immunophenotypic, and molecular characteristics of 7 EBV-negative ANKL. All patients were adults, with a median age of 63 years (range 22 to 83 y) and an M:F ratio of 2.5:1. Five patients were White, 1 Black, and 1 Asian. All patients presented acutely, with fever (6/7), cytopenias (6/7), and splenomegaly (4/7). Four patients had lymphadenopathy, 4 had extranodal disease. Bone marrow involvement was present in 5, with hemophagocytosis in 3. Peripheral blood was involved in 5 with the neoplastic cells containing prominent azurophilic granules. By immunohistochemistry and/or flow cytometry, the tumor cells lacked surface CD3 and were positive for CD56 (7/7), CD2 (5/5), CD8 (3/7), CD30 (4/5), and granzyme-B (6/6). They were negative for CD4, CD5, betaF1, TCRgamma, LMP1, and EBV-encoded RNA. Polymerase chain reaction for TCR gamma gene clonality was polyclonal. Mutational analysis revealed missense mutations in the STAT3 gene in both cases studied. Median survival was 8 weeks from the onset of disease. One patient received allogeneic bone marrow transplant and is alive with no disease (follow-up 15 mo). EBV-negative aggressive natural killer cell leukemia exists but is rare. It tends to occur in older patients and is indistinguishable clinically and pathologically from EBV-positive ANKL, with a similar fulminant clinical course. The high prevalence of Asian patients seen with EBV-positive disease seems less evident with EBV-negative cases.
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