Search

The objective of this study was to evaluate the prognostic impact of genomic regions in a series of human immunodeficiency virus (HIV)-related diffuse large B-cell lymphomas (HIV-DLBCLs) and post-transplant DLBCLs (PT-DLBCLs) analyzed by genome-wide DNA profiling. Minimal common regions (MCRs) were estimated on genomic profiles obtained using Affymetrix Human Mapping 250k Nsp I arrays and tested for their impact on clinical outcome by univariate analysis on 36 PT-DLBCLs, 19 HIV-DLBCLs and, as a control group, 149 DLBCLs arising in immunocompetent individuals (IC-DLBCLs). PT-DLBCL and HIV-DLBCL presented a similar outcome. Immunodeficiency-related DLBCL (ID-DLBCL) had a worse overall survival (OS) than IC-DLBCL. Seven MCRs showed a statistical impact on OS in PT-DLBCL and four in HIV-DLBCL. Among these, the presence of gains at 1q or at 18q defined a group of patients with PT-DLBCL with a very poor outcome ( p < 0.0001). The presence of del(3p14.2) or of ++ 2p23.1 identified a group of HIV-DLBCLs with a very poor outcome ( p == 0.0072). It was concluded that genomic aberrations affecting outcome differ between ID-DLBCL and IC-DLBCL and are also dependent on the type of acquired immunodeficiency. [ABSTRACT FROM AUTHOR]

Novel N-glycosylation sites are introduced by somatic mutation into the V genes of the majority of follicular lymphomas. Sites are positively selected and rare in normal memory B cells, indicating a potential role in tumour survival in the germinal centre (GC). The incidence of c. 40% in diffuse large B-cell lymphomas (DLBCL) parallels the known heterogenity of the disease. Immunodeficiency-related non-Hodgkin's lymphomas (NHL) include post-transplant lymphoproliferative disorders (PTLD) and acquired immunodeficiency syndrome-related NHL (AIDS-NHL). Most PTLD derive from B cells that carry mutated VH genes and that have completed the GC reaction. All AIDS-NHL carry mutated VH genes and variable features of GC or post-GC cells. To determine if N-glycosylation is a feature of immunodeficiency-related lymphomas, we analysed the VH genes of 19 PTLD and 36 AIDS-NHL. Novel sites were rare in PTLD (4/19), similar to memory B cells ( P = 0·15). AIDS-NHL, including DLBCL and Burkitt's lymphomas (BL), showed heterogeneity with 16 of 36 (44%) having novel sites. The findings indicate no selection of N-glycosylation sites in PTLD, consistent with post-GC features. The variable incidence of N-glycosylation sites in AIDS-NHL mirrors that in DLBCL and sporadic BL of immunocompetent hosts, supporting the known heterogeneity of these disorders, and possibly pointing to distinct routes of tumour development. [ABSTRACT FROM AUTHOR]

Aberrant promoter hypermethylation is a mechanism of tumour suppressor gene inactivation. We explored aberrant promoter hypermethylation of multiple genes in 88 human immunodeficiency virus (HIV)-non Hodgkin lymphomas (NHL), 25 post-transplant lymphoproliferative disorders (PTLD) and five common variable immunodeficiency (CVI)-related NHL. Twenty-six of 79 (32·9%) HIV-NHL, eight of 14 (57·1%) PTLD and two of five (40·0%) CVI–NHL showed aberrant hypermethylation of O6-methylguanine-DNA methyltransferase ( MGMT). Aberrant hypermethylation of death-associated protein-kinase ( DAP-K) occurred in 70 of 84 (83·3%) HIV–NHL, 19 of 25 (72·0%) PTLD and three of five (60·0%) CVI–NHL. These data implicate MGMT and DAP-K hypermethylation in lymphomagenesis of immunodeficient hosts. In particular, promoter hypermethylation of DAP-K represents the most frequent molecular alteration yet identified in immunodeficiency-related lymphomas. [ABSTRACT FROM AUTHOR]

Examines the usability of 0&sup6;-methylguanine DNA methyltransferase (MGMT) promoter hypermethylation on predicting survival of patients with brain tumors. Association between MGMT methylation status and MGMT protein expression; Role of MGMT on predicting survival of patients treated with alkylating agents; Effectivity of MGMT on protecting against the toxicity of acrolein.

Human herpesvirus type 8 (HHV-8) has been proposed as a pathogenetic factor for immunosuppression-associated primary central nervous system lymphoma (PCNSL). To verify this hypothesis, HHV-8 infection was investigated in 31 persons with PCNSL (16 AIDS-related, 15 AIDS-unrelated) and in 30 persons with systemic B cell non-Hodgkin's lymphomas (B-NHL; 15 AIDS-related, 15 AIDS-unrelated). All subjects with PCNSL scored negative by single-step polymerase chain reaction (PCR), suggesting a tumor virus load of < 100 viral copies/200,000 human haploid genome equivalents (HHGE). By applying Poisson assumptions to nested PCR, 16 of 31 persons with PCNSL were devoid of HHV-8 sequences: 1 subject with AIDS and PCNSL had 1–100 viral copies/200,000 HHGE, and 14 with PCNSL had <1 viral copy/200,000 HHGE. Similarly, 10 of 30 persons with systemic B-NHL were devoid of HHV-8 sequences; 20 had <1 viral copy/200,000 HHGE. The extremely low levels of infection rule out a role of HHV-8 in PCNSL pathogenesis and are consistent with HHV-8 infection of bystander cells contaminating the tumor clone. [ABSTRACT FROM PUBLISHER]

PKCζ and PKCι/λ form the atypical protein kinase C subgroup, characterised by a lack of regulation by calcium and the neutral lipid diacylglycerol. To better understand the regulation of these kinases, we systematically explored their interactions with various purified phospholipids using the lipid overlay assays, followed by kinase activity assays to evaluate the lipid effects on their enzymatic activity. We observed that both PKCζ and PKCι interact with phosphatidic acid and phosphatidylserine. Conversely, PKCι is unique in binding also to phosphatidylinositol-monophosphates (e.g., phosphatidylinositol 3-phosphate, 4-phosphate, and 5-phosphate). Moreover, we observed that phosphatidylinositol 4-phosphate specifically activates PKCι, while both isoforms are responsive to phosphatidic acid and phosphatidylserine. Overall, our results suggest that atypical Protein kinase C (PKC) localisation and activity are regulated by membrane lipids distinct from those involved in conventional PKCs and unveil a specific regulation of PKCι by phosphatidylinositol-monophosphates. [ABSTRACT FROM AUTHOR]

This article presents a letter to the editor concerning the risk of recurrent infections during watch and wait periods of early stage chronic lymphocytic leukemia.

Abstract: The frequent expression of the otherwise rare IgVH3-72 gene was demonstrated in highly stable B-cell chronic lymphocytic leukaemias (B-CLLs). Here we describe the immunophenotypic profiles of eleven IgVH3-72 B-CLLs, by investigating expression of ZAP-70 and of a set of surface antigens previously demonstrated to represent the signature of distinct B-CLL prognostic groups. All IgVH3-72 B-CLLs revealed a homogeneous immunophenotypic profile, expressing all the markers associated with good prognosis, but not those representing the signature of bad prognosis B-CLLs, including ZAP-70. Our results corroborate the notion that IgVH3-72 B-CLLs represent a molecularly and immunophenotypically homogeneous disease group with a good prognosis. [Copyright &y& Elsevier]

A letter to the editor discussing the transformation of philadelphia-negative chronic myeloproliferative disorders into leukaemic due to JAK2V617F mutation is presented.

Biobanks are driving motors of precision and personalized medicine by providing high-quality biological material/data through the standardization and harmonization of their collection, preservation, and distribution. UPO Biobank was established in 2020 as an institutional, disease, and population biobank within the University of Piemonte Orientale (UPO) for the promotion and support of high-quality, multidisciplinary studies. UPO Biobank collaborates with UPO researchers, sustaining academic translational research, and supports the Novara Cohort Study, a longitudinal cohort study involving the population in the Novara area that will collect data and biological specimens that will be available for epidemiological, public health, and biological studies on aging. UPO Biobank has been developed by implementing the quality standards for the field and the ethical and legal issues and normative about privacy protection, data collection, and sharing. As a member of the "Biobanking and Biomolecular Resources Research Infrastructure" (BBMRI) network, UPO Biobank aims to expand its activity worldwide and launch cooperation with new national and international partners and researchers. The objective of this manuscript is to report an institutional and operational experience through the description of the technical and procedural solutions and ethical and scientific implications associated with the establishment of this university research biobank. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) isolated from plasma are increasingly recognized as promising circulating biomarkers for disease discovery and progression, as well as for therapeutic drug delivery. The scientific community underlined the necessity of standard operative procedures for the isolation and storage of the EVs to ensure robust results. The understanding of the impact of the pre-analytical variables is still limited and some considerations about plasma anticoagulants and isolation methods are necessary. Therefore, we performed a comparison study between EVs isolated by ultracentrifugation and by affinity substrate separation from plasma EDTA and sodium citrate. The EVs were characterized by Nano Tracking Analysis, Western Blot, cytofluorimetric analysis of surface markers, and lipidomic analysis. While anticoagulants did not significantly alter any of the analyzed parameters, the isolation methods influenced EVs size, purity, surface markers expression and lipidomic profile. Compared to ultracentrifugation, affinity substrate separation yielded bigger particles highly enriched in tetraspanins (CD9, CD63, CD81), fatty acids and glycerolipids, with a predominant LDL- and vLDL-like contamination. Herein, we highlighted that the isolation method should be carefully evaluated prior to study design and the need of standardized operative procedures for EVs isolation and application to biomarkers discovery. [ABSTRACT FROM AUTHOR]

A letter to the editor discussing prognostic marker in chronic lymphocytic leukaemia is presented.

A fraction of chronic lymphocytic leukaemia (CLL) cases carry highly homologous B-cell receptors (BCR), i.e. characterized by non-random combinations of immunoglobulin heavy-chain variable ( IGHV) genes and heavy-chain complementarity determining region-3 (HCDR3), often associated with a restricted selection of IGVK/L light chains. Such ‘stereotyped’ BCR occur more frequently in CLL with unmutated (UM) than mutated (M) IGHV genes. We analysed 1426 IG rearrangements (from 1398 CLL cases) by a clustering driven by HCDR3 similarities. Molecular findings were correlated to time-to-treatment (TTT) and presence of known prognosticators. Sixty-nine clusters (319 IG-rearrangements, 22·4%) with stereotyped BCR were identified. Among 30 confirmed clusters (≥3 IG-rearrangements/cluster), we found 14 novel clusters, of which 11 had M IG rearrangements (M clusters) and predominantly (8/11) used IGHV3 subgroup genes. Recurrent cluster-biased amino acid changes were found throughout IGHV sequences of these ‘M clusters’. Regarding clinical outcome: (i) UM CLL from the IGHV1-2/1-3/1-18/1-46/7-4-1/IGKV1-39 cluster had poorer prognosis than UM/M cases, or UM cases using the same IGHV genes but not in clusters; (ii) M CLL from the IGHV3-21/IGLV3-21 cluster had TTT similar to UM CLL, and shorter than M CLL expressing IGHV3-21 but not in cluster. Altogether, our analysis identified additional molecular and clinical features for CLL expressing stereotyped BCR. [ABSTRACT FROM AUTHOR]

Predictors of chronic lymphocytic leukaemia (CLL) transformation to Richter syndrome (RS) are not established and were investigated in 185 consecutive CLL cases. Actuarial incidence of RS ( n = 17; all diffuse large B-cell lymphomas) at 10 years was 16·2% (95% confidence interval: 8·0–24·4%). At CLL diagnosis, prognosticators of RS by univariate analysis were IGHV homology ≥98% ( P = 0·006), IGHV4-39 usage ( P < 0·001), del13q14 absence ( P = 0·004), expression of CD38 ( P < 0·001) and ZAP70 ( P = 0·004), size ( P < 0·001) and number ( P < 0·001) of lymph nodes, advanced Binet stage ( P = 0·002), and lactate dehydrogenase ( P < 0·001). Multivariate analysis, performed separately for biological and clinical variables, identified CD38 expression [Hazard ratio (HR) = 4·26; P = 0·018], IGHV4-39 usage (HR = 4·29; P = 0·018), and lymph node size ≥3 cm (HR = 9·07; P < 0·001) as independent RS prognosticators. A multivariate model simultaneously analysing biological and clinical variables identified lymph node size ≥3 cm (HR = 6·51; P = 0·001) and del13q14 absence (HR = 4·08; P = 0·031) as independent RS prognosticators. Risk factors of CLL transformation differed from risk factors of CLL progression. These results suggest that CD38 and del13q14 may identify biological subsets of CLL with different RS predisposition. Predominant nodal disease, CD38 expression, IGHV4-39 usage, and absence of del13q14 may help in predicting RS at CLL diagnosis. Close monitoring and a careful biopsy policy are needed in patients carrying transformation risk factors. [ABSTRACT FROM AUTHOR]

We investigated the prevalence and prognostic role of CpG island methylation of the reduced folate carrier ( RFC) gene promoter region in primary central nervous system lymphoma (PCNSL) in immunocompetent patients. Genomic DNA from 40 PCNSL was used for methylation-specific polymerase chain reaction and bisulphite genomic sequencing of the RFC promoter region. Human immunodeficiency virus-negative systemic diffuse large B-cell lymphomas (DLBCL) were used as controls ( n = 50). The impact on outcome of RFC promoter methylation was assessed in 37 PCNSL patients treated with high-dose methotrexate (HD-MTX)-based chemotherapy ± radiotherapy. RFC promoter methylation occurred in 12 of 40 (30%) PCNSL and in four of 50 (8%) DLBCL ( P = 0·01). Of 37 PCNSL treated with HD-MTX-based chemotherapy, methylation occurred in nine cases (24%, M-PCNSL), while 28 cases (76%, U-PCNSL) were negative. Three M-PCNSL (33%) and 15 U-PCNSL (54%) achieved complete remission (CR) after primary chemotherapy. Logistic regression confirmed the independent association between CR rate and International Extranodal Lymphoma Study Group score ( P = 0·03), RFC promoter methylation ( P = 0·07) and use of cytarabine ( P = 0·08). The 3-year failure-free survival (FFS) and overall survival for M-PCNSL and U-PCNSL was 0% vs. 31 ± 9% ( P = 0·34) and 0% vs. 31 ± 9% ( P = 0·35) respectively. This is the first study to assess the methylation status of the RFC promoter in human tumour samples. RFC methylation is more common in PCNSL compared with systemic DLBCL, and is associated with a lower CR rate to HD-MTX-based chemotherapy. If confirmed in prospective trials on PCNSL treated with HD-MTX alone, these data may suggest the necessity for alternative strategies in M-PCNSL considering the increased risk of MTX resistance by tumour cells. [ABSTRACT FROM AUTHOR]

Summary. Plasmablastic lymphoma (PBL) of the oral cavity is an aggressive B-cell lymphoma associated with human immunodeficiency virus infection. Although the lymphoma phenotype is consistent with late B-cell maturation, the molecular histogenesis of PBL is unknown. We investigated PBL of the oral cavity (n = 12) for mutations of immunoglobulin variable heavy chain (IgVH ) and BCL -6 genes, which are acquired by B cells at the time of germinal centre (GC) transit, and for expression of BCL-6, MUM-1 and CD138, which distinguish GC B cells from post-GC B cells. Somatic IgVH hypermutation occurred in 4/10 PBL whereas 6/10 PBL displayed germline IgVH genes. Among PBL carrying hypermutated IgVH genes, the pattern of IgVH mutations was consistent with antigen stimulation in two cases. Mutations of the BCL -6 gene were restricted to 1/12 patients with PBL of the oral cavity. All cases of PBL of the oral cavity displayed the BCL-6– /MUM-1+ /CD138+ phenotype that is consistent with late stage of B-cell differentiation. Overall, these data indicate that, despite a common phenotype and an apparently similar degree of differentiation, PBL of the oral cavity are characterized by histogenetic heterogeneity. A subset of PBL of the oral cavity carried the molecular clues of GC transit and conceivably originated from a B-cell subset corresponding to post-GC B cells. Conversely, another fraction of these lymphomas were devoid of somatic IgVH mutations and appeared to originate from naive B cells that have undergone preterminal differentiation independent of GC transit. [ABSTRACT FROM AUTHOR]

Abstract As part of an effort to identify druggable diacylglycerol kinase alpha (DGKα) inhibitors, we used an in-silico approach based on chemical homology with the two commercially available DGKα inhibitors R59022 and R59949. Ritanserin and compound AMB639752 emerged from the screening of 127 compounds, showing an inhibitory activity superior to the two commercial inhibitors, being furthermore specific for the alpha isoform of diacylglycerol kinase. Interestingly, AMB639752 was also devoid of serotoninergic activity. The ability of both ritanserin and AMB639752, by inhibiting DGKα in intact cells, to restore restimulation induced cell death (RICD) in SAP deficient lymphocytes was also tested. Both compounds restored RICD at concentrations lower than the two previously available inhibitors, indicating their potential use for the treatment of X-linked lymphoproliferative disease 1 (XLP-1), a rare genetic disorder in which DGKα activity is deregulated. Graphical abstract Image 1 Highlights • A potent DGKα inhibitor (AMB639752) has been identified via virtual screening. • Available DGKα inhibitors (ritanserin) are serotonin receptors antagonist. • AMB639752, instead, does not perturb serotonin signaling. • AMB639752 restores restimulation induced cell death in SAP deficient lymphocytes. [ABSTRACT FROM AUTHOR]

Summary Changes in DNA copy number (CN) and DNA methylation represent important aberrations for lymphomas and other cancers. Here, for the first time, we show that the Illumina Infinium Methylation (IIM) assay, although not originally designed for CN profiling, is able to estimate CN changes. We compared the IIM CN profiles to those obtained with a standard technique in a series of diffuse large B-cell lymphomas: the profiles showed a high degree of consensus. The demonstration of CN profiling as an additional function of the IIM assay may impact the choice of platform for methylation profiling of haematological and solid tumours. [ABSTRACT FROM AUTHOR]

Analysis of the chronic lymphocytic leukemia (CLL) coding genome has recently disclosed that the NOTCH1 proto-oncogene is recurrently mutated at CLL presentation. Here we assessed the prognostic role of NOTCH1 mutations in CLL. Two series of newly diagnosed CLL were utilized as training (n=309) and validation (n=230) cohorts. NOTCH1 mutations occurred in 11.0% and 11.3% CLL of the training and validation series, respectively. In the training series, NOTCH1 mutations led to a 3.77 fold increase in the hazard of death and to shorter overall survival (OS) (p<.001). Multivariate analysis selected NOTCH1 mutations as an independent predictor of OS after controlling for confounding clinical and biological variables. The independent prognostic value of NOTCH1 mutations was externally confirmed in the validation series. The poor prognosis conferred by NOTCH1 mutations was attributable, at least in part, to shorter treatment-free survival and higher risk of Richter transformation. Although NOTCH1 mutated patients were devoid of TP53 disruption in >90% cases in both training and validation series, the OS predicted by NOTCH1 mutations was similar to that of TP53 mutated/deleted CLL. NOTCH1 mutations are an independent predictor of CLL OS, tend to be mutually exclusive with TP53 abnormalities and identify cases with a dismal prognosis. [ABSTRACT FROM AUTHOR]

The immunoglobulin heavy chain variable gene ( IGHV) mutational status has been recognized as an important predictor of prognosis in chronic lymphocytic leukaemia (CLL) since 1999. More recently, other features of the B-cell receptor, such as stereotypy, have been identified as capable of refining the prognostic potential of IGHV status in the clinical assessment of CLL patients. In this context, different genes belonging to the IGHV3 subgroup, the most frequently used subgroup in CLL, have been shown to denote disease subsets that either display a bad prognosis (i.e. IGHV3-21, IGHV3-23) or are associated with particularly good clinical outcomes, including a highly stable/indolent clinical course, even prone to spontaneous regression (i.e. IGHV3-72, IGHV3-30). The present review focuses on the molecular and biological features of CLL-expressing specific genes belonging to the IGHV3 subgroup that are known to mark disease subsets with completely different clinical courses, and may be possibly related to CLL pathogenesis via antigen and/or superantigen involvement. [ABSTRACT FROM AUTHOR]

Deletion of 17p ( TP53) identifies a rare subset of chronic lymphocytic leukaemia (17p- CLL) with aggressive behaviour. Genome-wide DNA-profiling was performed to investigate 18 patients with 17p- CLL. All cases had multiple copy-number (CN) changes. Among the several recurrent CN changes identified, 8q24.13-q24.1-gain ( MYC), 8p-loss ( TNFRSF10A/B, also known as TRAIL1/2) and 2p16.1-p14-gain ( REL/BCL11A) appeared frequently represented. 8p-loss and 2p16.1-p14-gain also appeared clinically relevant and predicted significant shorter time from diagnosis to treatment (8p-loss) and overall survival (8p-loss and 2p16.1-p14-gain, P < 0·05). These observations document a highly unstable genome in 17p- CLL and suggest that additional genes outside the TP53 locus may be important for tumour behaviour. [ABSTRACT FROM AUTHOR]

Hairy cell leukaemia (HCL) is a rare B-cell neoplasm for which the molecular mechanisms are largely unknown. High-density genome-wide DNA profiling was performed with Affymetrix 250K arrays to analyse copy number (CN) changes and loss of heterozygosity (LOH) in 16 cases of HCL. Four of 16 cases (25%) demonstrated gross non-recurrent CN deletions. Within the affected regions, we identified genes involved in bone marrow fibrosis ( FGF12) and response to treatment ( TP53) in individual cases. Large regions (>5 Mb) of LOH without any concomitant DNA CN changes were identified in 5/16 (31%) HCL and were indicative of uniparental disomy UD. The germline origin of UD was demonstrated in one case for which a matched normal sample was available. Overall analysis of LOH showed that identical loci were recurrently targeted in chromosomes 1, 2 and 6. As a whole, however, HCL showed a remarkably stable genome. This finding adds to several other features that are unique to HCL among mature B-cell tumours. [ABSTRACT FROM AUTHOR]

Monoclonal gammopathy of uncertain significance (MGUS) may become symptomatic for autoimmune manifestations. We report on the prevalence and clinical course of immune thrombocytopenic purpura (ITP) observed in a consecutive series of 228 MGUS patients. At MGUS diagnosis, ITP was determined in 6/228 cases, accounting for a prevalence of 2630/100 000 [95% confidence interval (CI): 1210–5620]. One incidental ITP case occurred after 21 months of follow-up. After a follow-up of 681·3 patient-years, the crude incidence of ITP in MGUS was 146·8 per 100 000 patient-year (95% CI: 3·7–817·8). Overall, these observations point to an association between MGUS and ITP. [ABSTRACT FROM AUTHOR]

Post-transplant lymphoproliferative disorders (PTLD) are a major complication of solid organ transplantation, representing a cause of severe morbidity and mortality. Apart from Epstein–Barr virus infection, knowledge of the pathogenesis of monoclonal PTLD is limited. Powerful analysis techniques, such as whole genomic DNA profiling (array comparative genomic hybridisation), can improve our understanding of PTLD pathogenesis. Whole genome profiling using the Affymetrix GeneChip Human Mapping 10 k 2·0 was performed on 20 PTLD cases and 25 cases of diffuse large B-cell lymphoma (DLBCL) from immunocompetent patients as a control group. Recurrent lesions were detected among all the samples. Chromosome 18q, 7q, 3q and 12 were the most common gains in the control group. Chromosomes 5p and 11p were commonly gained in PTLD–DLBCL. The latter had frequent losses of 6q, 17p, 1p and 9p. Chromosome 12p was the most frequent target of deletions among PTLD–DLBCL cases. Loss of heterozygosity (LOH) did not always match DNA loss: chromosome 10 seemed to be targeted by uniparental disomy in PTLD. Small deletions and gains, involving both known ( BCL2 and PAX5) and unknown genes ( ZDHHC14), were identified. These data suggest that PTLD share, at a lower frequency, common genetic aberrations with DLBCL from immunocompetent patients. The demonstration of 9p13 amplification emphasises the importance of PAX5 in PTLD. The combination of DNA copy number and LOH assessment lead to the hypothesis that uniparental disomy may be a potential mechanism in B-cell lymphomagenesis. [ABSTRACT FROM AUTHOR]