Genetic lesions in MYC and STAT3 drive oncogenic transcription factor overexpression in plasmablastic lymphoma.

Plasmablastic lymphoma mutational profile is undescribed. Here we performed a targeted exonic NGS analysis of 30 plasmablastic lymphoma cases with a B cell lymphoma dedicated panel and FISH for the detection of MYC rearrangements. A complete phenotyping of the neoplastic and microenvironment cell populations was also performed. We have identified an enrichment in recurrent genetic events in MYC (69% with MYC translocation or amplification and 3 cases with missense point mutations), PRDM1/Blimp1 and STAT3 mutations. These gene mutations were more frequent in EBV positive disease. Other genetic events included mutations in BRAF, EP300, BCR (CD79A and CD79B), NOTCH pathway (NOTCH2, NOTCH1 and SGK1) and MYD88pL265P. Immunohistochemical analysis showed consistent MYC expression, higher in cases with MYC rearrangements together with phospho-STAT3 (Tyr705) overexpression in cases with STAT3 SH2 domain mutations. Microenvironment populations were heterogeneous and unrelated with EBV, with an enrichment of Tumor Associated Macrophages (TAM) and PD1 positive T cells. PD-L1 was expressed in all cases in the TAM population but only in 5 cases in the neoplastic cells (4 out of 14 EBV positive cases). HLA expression was absent in the majority of PBL cases. In summary, Plasmablastic lymphoma mutational profile is heterogeneous and related with EBV infection. Genetic events in MYC, STAT3 and PRDM1/Blimp1 are more frequent in EBV positive disease. An enrichment in TAM and PD1 reactive T lymphocytes is found in the microenvironment of PBL cases, that express PD-L1 in the neoplastic cells in a fraction of cases.

Diagnostic value of bone marrow core biopsy patterns in lymphoplasmacytic lymphoma/Waldenström macroglobulinaemia and description of its mutational profiles by targeted NGS.

AIMS: The aim of this study was to describe the characteristics of the bone marrow infiltration found in a series of clinically defined lymphoplasmacytic lymphoma (LPL)/Waldenström macroglobulinaemia (WM) and IgM-monoclonal gammopathy of undetermined significance (MGUS) and to perform a targeted next-generation sequencing (NGS) for the identification of additional somatic mutations to MYD88p.L265P in LPL/WM. METHODS: We have reviewed a series of 35 bone marrow biopsies from 28 patients with a clinical diagnosis of LPL/WM (24 cases) or MGUS (4 cases). Bone marrow infiltration characteristics by morphology, immunohistochemistry, flow cytometry (FCM), allele-specific real-time PCR for the detection of MYD88p.L265P mutation, targeted exonic amplicon-based NGS of 35 lymphoma-related genes and direct sequencing were analysed. RESULTS: Our findings show that bone marrow trephine biopsy evaluation is superior to FCM in the identification of significant lymphoid infiltrates. A combined paratrabecular and interstitial infiltration pattern is the most common feature in LPL/WM while a patchy interstitial pattern characterises IgM-MGUS cases. MYD88p.L265P mutation was found by allele-specific-PCR in 92% of the LPL cases (22 out of 24) and 25% of IgM-MGUS cases (1 out of 4 cases). In addition to MYD88p.L265P somatic mutations in CXCR4, KMT2D, PRDM1/Blimp1, MYC and ID3 were found by NGS and direct sequencing in 4 cases. CONCLUSIONS: In conclusion, bone marrow core biopsy evaluation is critical in the identification of unequivocal bone marrow infiltration by LPL/WM. In addition to MYD88p.L265P, somatic mutations in CXCR4, KMT2D, PRDM1/Blimp1, MYC and ID3 can appear in a fraction of LPL/WM.

Clonal Evolution in Primary Diffuse Large B-Cell Lymphoma of the Central Nervous System.

Primary diffuse large B-cell lymphoma (DLBCL) of the central nervous system (CNS) is an aggressive subtype of DLBCL with characteristic clinicopathologic features. Relapse outside the CNS involving extranodal locations has been found in a fraction of cases (16%). Here we describe a case of DLBCL arising in the CNS that relapsed 18 months after the initial diagnosis in the testis and bilateral adrenal glands. Both tumors showed equivalent morphology, phenotype, cytogenetic features, and clonal relationship. Somatic mutation analysis by next generation sequencing demonstrated MYD88L265P mutation in both tumors and de novo CD79B Y196S mutation exclusive to the relapse. The pattern of mutations suggest that the 2 tumors might have evolved from a common progenitor clone with MYD88L265P being the founder mutation. A meta-analysis of the literature shows a significantly high frequency of concurrent MYD88L265P and CD79B ITAM mutations in primary CNS lymphoma and testicular DLBCL, underscoring the role of B cell receptor and nuclear factor kB activation by somatic mutations in these lymphomas that colonize immune-privileged sites. In summary, here we illustrate that targeted next generation sequencing for the detection of hot spot somatic mutations in relapsed DLBCL is useful to confirm ABC phenotype and discovers relevant information that might influence therapeutic decision.

Mycosis fungoides progression could be regulated by microRNAs.

Differentiating early mycosis fungoides (MF) from inflammatory dermatitis is a challenge. We compare the differential expression profile of early-stage MF samples and benign inflammatory dermatoses using microRNA (miRNA) arrays. 114 miRNAs were found to be dysregulated between these entities. The seven most differentially expressed miRNAs between these two conditions were further analyzed using RT-PCR in two series comprising 38 samples of early MFs and 18 samples of inflammatory dermatitis. A series of 51 paraffin-embedded samples belonging to paired stages of 16 MF patients was also analyzed. MiRNAs 26a, 222, 181a and 146a were differentially expressed between tumoral and inflammatory conditions. Two of these miRNAs (miRNA-181a and miRNA-146a) were significantly deregulated between early and advanced MF stages. Bioinformatic analysis showed FOXP3 expression to be regulated by these miRNAs. Immunohistochemistry revealed the level of FOXP3 expression to be lower in tumoral MFs than in plaque lesions in paraffin-embedded tissue. A functional study confirmed that both miRNAs diminished FOXP3 expression when overexpressed in CTCL cells. The data presented here suggest that the analysis of a restricted number of miRNAs (26a, 222, 181a and 146a) could be sufficient to differentiate tumoral from reactive conditions. Moreover, these miRNAs seem to be involved in MF progression.

Plasmablastic lymphoma phenotype is determined by genetic alterations in MYC and PRDM1.

Plasmablastic lymphoma is an uncommon aggressive non-Hodgkin B-cell lymphoma type defined as a high-grade large B-cell neoplasm with plasma cell phenotype. Genetic alterations in MYC have been found in a proportion (~60%) of plasmablastic lymphoma cases and lead to MYC-protein overexpression. Here, we performed a genetic and expression profile of 36 plasmablastic lymphoma cases and demonstrate that MYC overexpression is not restricted to MYC-translocated (46%) or MYC-amplified cases (11%). Furthermore, we demonstrate that recurrent somatic mutations in PRDM1 are found in 50% of plasmablastic lymphoma cases (8 of 16 cases evaluated). These mutations target critical functional domains (PR motif, proline rich domain, acidic region, and DNA-binding Zn-finger domain) involved in the regulation of different targets such as MYC. Furthermore, these mutations are found frequently in association with MYC translocations (5 out of 9, 56% of cases with MYC translocations were PRDM1-mutated), but not restricted to those cases, and lead to expression of an impaired PRDM1/Blimp1α protein. Our data suggest that PRDM1 mutations in plasmablastic lymphoma do not impair terminal B-cell differentiation, but contribute to the oncogenicity of MYC, usually disregulated by MYC translocation or MYC amplification. In conclusion, aberrant coexpression of MYC and PRDM1/Blimp1α owing to genetic changes is responsible for the phenotype of plasmablastic lymphoma cases.