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Peripheral T-cell lymphoma (PTCL) is a clinically aggressive disease, with a poor response to therapy and a low overall survival rate of approximately 30% after 5 years. We have analyzed a series of 105 cases with a diagnosis of PTCL using a customized NanoString platform (NanoString Technologies, Seattle, WA) that includes 208 genes associated with T-cell differentiation, oncogenes and tumor suppressor genes, deregulated pathways, and stromal cell subpopulations. A comparative analysis of the various histological types of PTCL (angioimmunoblastic T-cell lymphoma [AITL]; PTCL with T follicular helper [TFH] phenotype; PTCL not otherwise specified [NOS]) showed that specific sets of genes were associated with each of the diagnoses. These included TFH markers, cytotoxic markers, and genes whose expression was a surrogate for specific cellular subpopulations, including follicular dendritic cells, mast cells, and genes belonging to precise survival (NF-κB) and other pathways. Furthermore, the mutational profile was analyzed using a custom panel that targeted 62 genes in 76 cases distributed in AITL, PTCL-TFH, and PTCL-NOS. The main differences among the 3 nodal PTCL classes involved the RHOAG17V mutations (P < .0001), which were approximately twice as frequent in AITL (34.09%) as in PTCL-TFH (16.66%) cases but were not detected in PTCL-NOS. A multivariate analysis identified gene sets that allowed the series of cases to be stratified into different risk groups. This study supports and validates the current division of PTCL into these 3 categories, identifies sets of markers that can be used for a more precise diagnosis, and recognizes the expression of B-cell genes as an IPI-independent prognostic factor for AITL.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive and heterogeneous disease with variable prognosis associated with clinical features, cell-of-origin and genetic aberrations. The main problems for DLBCL patients are that a substantial percentage of them (30-40%) are refractory to treatment or relapse and the lack of accurate predictive markers that adequately determine which patients will benefit from immunochemotherapy. Several recent deep-sequencing studies have proposed new genetic subtypes based on the DLBCL genomic profile (Wright et al., 2020; Lacy et al. 2020) and associated with clinical outcome. However, a consensus and validated classification is still needed. The aim was to determine whether genetic alterations of individual genes or genes clustered in pathways are associated with clinical outcome in immunochemotherapy-treated patients.

Introduction: Aberrant methylation of the 5′ gene promoter regions is an epigenetic phenomenon that is the one of the major mechanism of inactivation of tumor supressor genes. DNA methylation of the promoter region has been described for several genes in various malignant diseases, and each tumour type may have its own pattern of methylation. To determine the methylation status and expression of cell cycle inhibitors genes (p14, p15, p16), repair genes (MGMT and hMLH1) and the apoptosis regulator gene (DAPKinase) and the possible role of this epigenetic phenomenon in tumour progression of plasma cell disorders, we analyzed the methylation profile of MM, MGUS, and plasmacytomas comparing them with their protein expression. Patients and Methods: A total of 51 cases: 30 MM, 13 MGUS, and 8 plasmacytomas (3 Solitary Bone Plasmacytoma, 5 Extramedullary Plasmacytoma) were included in the study. Bone marrow plasma cells were purified using magnetic microbeads labeled with CD138 in samples with MM and MGUS. Methylation-specific polymerase chain reaction (MSP) for p14, p15, p16, MGMT, hMLH1 and DAPKinase was performed. MSP results were matched to protein expression studies by immunohistochemistry for p15, p16, MGMT and hMLH1. Results: The frequency of aberrant methylation among the MM samples was: 50% for p16, 16.7% for p15, 10% for hMLH1, 23.3% for MGMT, 30% for DAPK. In MGUS samples we found 38.5% for p16, 15.4% for p15, 0% for hMLH1, 7.7% for MGMT and 15.4% for DAPK methylation. The frequency of methylation in plasmacytomas was 62.5% for p16, 25% for p15, 0% for hMLH1, 62.5% for MGMT and 50% for DAPK. p14 was unmethylated in all cases (n=51). The correlation between gene methylation status and protein expression was assessed in 17 MM, 11 MGUS, and 8 plasmacytomas and we found that promoter methylation was strongly associated with gene silencing. All the samples methylated had lost the protein expression. In summary these findings demonstrate that aberrant methylation is an important mechanism of gene silencing in plasma cell disorders: 83.3% of MM, 46.1 of MGUS, and 75% of plasmacytomas have at least one hypermethylated gene (figure 1). We also show, that hypermethylation of p16 is a common phenomenon in plasma cell discrasias while there was no methylation of p14. Although the size of sample is small, we found that hMLH1 hypermethylation was found only in MM, while in plasmacytomas hypermethylation of MGMT and DAPK were frequent events. Moreover in survival studies of MM patients, a trend was observed between simultaneous aberrant methylation of hMLH1 and MGMT and poorer survival, but the number of cases studied limits the statistical analysis and the clinical implications of these results. To better define the clinical impact of methylation markers in plasma cell discrasias, it is therefore necessary to analyze a large number of patient samples. Figure 1. Gene methylation profiling of MM(A), MGUS(B), and plasmacytomas(C) using MSPCR Figure 1. Gene methylation profiling of MM(A), MGUS(B), and plasmacytomas(C) using MSPCR