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Prolymphocytic leukaemias of B and T cell subtype are rare diseases. Despite recent advances in immunophenotyping and molecular cytogenetics, leading to a better understanding of the underlying cell biology of the prolymphocytic leukaemias, prognosis for these patients remains poor. Purine analogues and monoclonal antibodies have shown efficacy in B-cell prolymphocytic leukaemia although further studies are warranted. Monoclonal antibody therapy with alemtuzumab has significantly improved outcome in T-cell prolymphocytic leukaemia (T-PLL) but responses are still transient and further disease progression is inevitable. While allogeneic stem cell transplant is an attractive option, due to the older age group of T-PLL patients the morbidity and mortality associated with the procedure is significant. [ABSTRACT FROM AUTHOR]

The non-Hodgkin's lymphomas (NHLs) comprise a heterogeneous collection of lymphoproliferative malignancies, which are most common in people aged over 55 years. Diffuse large B-cell lymphoma (DLBCL) is the most common type of NHL, accounting for approximately 30% of all new patients. Follicular lymphoma (FL) is the second most common NHL sub-type, and accounts for a further 22% of cases. While the incidence of most other cancers is decreasing, that of NHL is increasing steadily. During the 1970's and 1980's, worldwide NHL incidence rose by 3-4% per year. This rise has slowed in the 1990's, but an annual increase of 1-2% is still being recorded. Over the last five years, the introduction of monoclonal antibodies, and specifically the anti-CD20 monoclonal antibody, rituximab, has radically changed treatment of B-cell NHL. Rituximab is a genetically engineered chimeric mouse/human monoclonal antibody which binds to the transmembrane antigen, CD20, a non-glycosylated phosphoprotein, located on pre-B and mature B lymphocytes. This antigen is expressed on over 95% of all B cell NHLs, and on normal B cells, but not on haematopoietic stem cells, normal or malignant plasma cells. The Fc domain of rituximab recruits immune effector functions to mediate B cell lysis. Possible mechanisms of cytotoxicity include complement-dependent cytotoxicity (CDC) resulting from C1q binding, and antibody-dependent cellular cytotoxicity (ADCC) mediated by one or more of the Fcγ receptors on the surface of granulocytes, macrophages and NK cells. It is also possible that the binding of rituximab to the CD20 antigen on the cell surface may directly induce apoptosis. For patients with both follicular and diffuse large B-cell NHL, several large scale prospective randomised trials have demonstrated prolongation of remission when rituximab is incorporated into first line treatment, and, in follicular lymphoma, as a component of salvage therapy. As a result of these studies, current European indications for rituximab include: • the treatment of previously untreated patients with stage III-IV follicular lymphoma in combination with cyclophosphamide, vincristine and prednisone (CVP) chemotherapy • as maintenance therapy in patients with relapsed follicular lymphoma responding to induction therapy with chemotherapy or immunochemotherapy • the treatment of patients with diffuse large B cell NHL in combination with cyclophosphamide, doxorubicin, vincristine and prednisone (CHOP) chemotherapy. This paper examines the evidence supporting the use of rituximab in these settings, and places its use into the context of standard clinical practice. [ABSTRACT FROM AUTHOR]

Objectives: Individual immunoglobulins expressed by B-cell lymphomas represent tumor-specific antigens (‘idiotypes’). Immunization with idiotype in follicular lymphoma patients may induce specific immune responses, sustained progression-free survival, and disappearance of minimal residual disease. Manufacturing of idiotype vaccines has mostly relied on heterohybridomas established from viable lymphoma cells. This paper describes the feasibility of production of GMP-grade idiotype vaccines as recombinant Fab fragments in Escherichia coli. Methods: IgH and IgL transcripts were analyzed by anchored PCR from 106 lymphoma and nine control biopsies. Lymphoma-derived V segments were inserted into prokaryotic expression plasmids. Recombinant idiotype Fab fragments were expressed in E. coli in a fermentation system. Results: Idiotype IgH and IgL transcripts were identified in 95% of 106 lymphoma biopsies according to stringent clonality criteria. Large-scale idiotype expression was successful in 69 of 78 cases (89%) and yielded a median of 17 mg (range: 1.2–250 mg) recombinant Fab protein. After affinity chromatography, median vaccine purity was 99% heterodimeric Fab protein (range: 72–100%). Bacterial protein contamination was detectable in one vaccine only. Fab proteins with IgL lambda chains had a tendency for inferior yield and lesser purity than κ-type Fabs. Among other structural idiotype features (isotype, V family usage, somatic hypermutation pattern, novel glycosylation sites, CDR III net charge), no consistent influences on Fab yield or purity were detected. Conclusions: Anchored PCR cloning and subsequent protein expression in E. coli provides a reliable technological basis for clinical idiotype vaccination trials. [ABSTRACT FROM AUTHOR]

Objective: The mutational status of the immunoglobulin (Ig) VH gene in B-cell chronic lymphocytic leukaemia (B-CLL) identifies two subgroups of patients with significantly different outcomes. We investigated the association of ZAP-70 expression with IgVH mutational status in B-CLL by quantifying ZAP-70 mRNA, to evaluate its use as a surrogate marker for mutational status. The aim of this study was to develop a quantitative reverse transcriptase-polymerase chain reaction (RQ-PCR) assay for the detection of ZAP-70 expression in a group of patients whose mutational status and cytogenetics had been determined previously. Methods: RQ-PCR was used to analyse ZAP-70 expression from 42 B-CLL patients. B cells were purified using CD19 magnetic bead system and total RNA was isolated. RQ-PCR was performed using Taqman PCR. Results: Twenty-five patients (60%) had mutated and 17 (40%) had unmutated IgVH genes; 94% (16/17) of patients with unmutated IgVH gene were ZAP-70 positive as assessed by RQ-PCR and 92% (23/25) of patients with mutated IgVH gene were ZAP-70 negative. In three patients, ZAP-70 expression and IgVH mutational status were discordant. Conclusion: This paper describes an RQ-PCR assay for the detection of ZAP-70 expression and confirms that IgVH unmutated CLL cells have a high expression of ZAP-70 in comparison with IgVH mutated CLL. This robust method acts as a surrogate marker for IgVH mutational status albeit with <100% concordance. However, it does provide better concordance with mutational status than that reported using flow cytometry. [ABSTRACT FROM AUTHOR]

Hematopoietic stem cell (HSC) transplantation's success lies in its ability to induce immune reconstitution. To date, there is no review published to compare the immune reconstitution among the three sources of HSC: umbilical cord blood (UCB), bone marrow (BM), and peripheral blood (PB). The review aims to analyze the kinetic of immune reconstitution among UCB, PB, and BM in HSC transplant patients by focusing on natural killer (NK) cells, B and T lymphocytes, and neutrophils. A systematic review was conducted through five databases, searching for clinical trials and randomized control trials (RCTs) which analyze the kinetics of immune reconstitution in at least two sources. Selected studies were assessed with Cochrane RoB 2.0. This review included 14 studies, with a total of 2539 subjects. The PB group achieved the fastest time to neutrophil recovery, while the B‐cell count was the highest in the UCB group. The T‐cell count is the lowest in the BM group, and the NK‐cell count does not differ significantly among the three HSC sources. Among the three sources of HSC, there is no superior HSC source for any immune reconstitution parameter. More studies must be conducted to compare the immune reconstitution and clinical outcomes of all HSC sources in specific diseases. [ABSTRACT FROM AUTHOR]

Objective: Given a proposed role for PD‐L1+ and IL‐10‐producing B‐cell subsets in promoting certain cancers, we sought to characterize the frequency and phenotype of B cells in patients with chronic myeloproliferative neoplasms (MPNs) and the influence of ruxolitinib and interferon‐α2 therapy. Methods: We analyzed B‐cell frequencies and phenotype in patients with MPNs (n = 107), before and during treatment with ruxolitinib (n = 29), interferon‐α2 (n = 21), or the two drugs in combination (COMBI; n = 42) and healthy donors (HDs; n = 52) using flow cytometry. Results: Myelofibrosis patients had lower lymphocyte counts and proportions of B cells than patients with essential thrombocythemia or polycythemia vera and HDs. The B‐cell count correlated inversely with JAK2‐V617F allele burden and spleen size and increased after ruxolitinib or COMBI treatment. The proportions of PD‐L1+ B cells and PD‐1+ B cells were significantly higher in patients with myelofibrosis or polycythemia vera than in HDs and decreased during ruxolitinib and COMBI treatment. The proportions of TNF‐α+ and IL‐6+ B cells were elevated in myelofibrosis patients. The proportion of IL‐6+ B cells decreased, and the proportion of IL‐10+ B cells increased during ruxolitinib treatment. Conclusion: B‐cell frequency and phenotype were altered in MPN patients. Ruxolitinib therapy had marked effects on both frequency and phenotype. [ABSTRACT FROM AUTHOR]

The management of patients with chronic lymphocytic leukaemia ( CLL) has improved with the utilisation of ofatumumab as a novel anti- CD20 monoclonal antibody. However, as half of the patients fail to respond to the treatment, the aim of this study was to evaluate circulating CLL cell depletion and clinical response according to the context of complement activation and Fcγ RIIIA polymorphism in ten CLL patients with relapsed/refractory disease. At the end of the treatment, results indicated that circulating CD5+ CD19+ CLL cell depletion was major (<0.01 × 109/L) in 4 of 10 patients, partial (>50% decrease) in 4 of 10 patients and ineffective for the two other patients. No clinical modifications were observed following ofatumumab introduction. Ofatumumab administration leads to a rapid and important exhaustion of complement C4 levels in patients with initial lymphocytosis. C4 exhaustion was accelerated in a non-responder patient, and incomplete in two patients with partial circulating depletion. Moreover, delaying weekly to monthly ofatumumab injections improved CLL cell depletion in two patients. Fcγ RIIIA 158 polymorphism ( FF n = 6 and VF n = 4) was not associated with major and/or partial circulating CLL cell depletion. In conclusion, ofatumumab induces an important C4 exhaustion that needs to be taken into account when treating CLL patients with ofatumumab. [ABSTRACT FROM AUTHOR]

The physiology of B cells is intimately connected with the function of their B-cell receptor ( BCR). B-cell lymphomas frequently (dys)regulate BCR signalling and thus take advantage of this pre-existing pathway for B-cell proliferation and survival. This has recently been underscored by clinical trials demonstrating that small molecules (fosfamatinib, ibrutinib, idelalisib) inhibiting BCR-associated kinases ( SYK, BTK, PI3K) have an encouraging clinical effect. Here we describe the current knowledge of the specific aspects of BCR signalling in diffuse large B-cell lymphoma ( DLBCL), follicular lymphoma, chronic lymphocytic leukaemia ( CLL) and normal B cells. Multiple factors can contribute to BCR pathway (dys)regulation in these malignancies and the activation of 'chronic' or 'tonic' BCR signalling. In lymphoma B cells, the balance of initiation, amplitude and duration of BCR activation can be influenced by a specific immunoglobulin structure, the expression and mutations of adaptor molecules (like GAB1, BLNK, GRB2, CARD11), the activity of kinases (like LYN, SYK, PI3K) or phosphatases (like SHIP-1, SHP-1 and PTEN) and levels of micro RNAs. We also discuss the crosstalk of BCR with other signalling pathways ( NF-κB, adhesion through integrins, migration and chemokine signalling) to emphasise that the ' BCR inhibitors' target multiple pathways interconnected with BCR, which might explain some of their clinical activity. [ABSTRACT FROM AUTHOR]

Aim We evaluated the prognostic significance of standardized uptake value ( SUVmax), metabolic tumor volume ( MTV), and total lesion glycolysis ( TLG) obtained by F-18 FDG PET/ CT ( PET/ CT) in patients with diffuse large B-cell Lymphomas ( DLBCL) presenting intermediate IPI score. Material and Methods Fifty-two patients (61 ± 13 yr) underwent PET/ CT before the first-line chemotherapy. The mean SUVmax value, the summed MTV (cm3; 42% threshold), and the cumulative TLG (g) were registered. The patients were followed up 18 months thereafter (range 3-41 months). The PET/ CT results were compared to the event-free survival ( EFS). Results At univariate analysis, SUVmax and lactate dehydrogenase ( LDH) levels were predictive, but discordantly. The Kaplan- Meier survival analysis for SUVmax showed a significant better EFS in patients presenting higher values as compared to those having lesser ( P = 0.0002, HR 0.13). Summed MTV and cumulative TLG were not suitable for predicting EFS. Conclusion Despite the availability of new tools for the quantitative assessment of disease activity on PET/ CT, the SUVmax rather than MTV and TLG remains the only predictor for EFS in DLBCL patients. The magnitude of glycolytic activity rather than the amount of metabolically active burden holds a predominant value for determining the response to chemotherapy in DLBCL. [ABSTRACT FROM AUTHOR]

Objectives The tumor microenvironment, including tumor-infiltrating lymphocytes and myeloid-derived cells, is an important factor in the pathogenesis and clinical behavior of malignant lymphoma. However, the prognostic significance of peripheral lymphocytes and monocytes in lymphoma remains unclear. Methods We evaluated the prognostic impact of the absolute lymphocyte count ( ALC), absolute monocyte count ( AMC), and lymphocyte/monocyte ratio ( LMR) in 359 diffuse large B-cell lymphoma ( DLBCL) patients treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone ( R- CHOP). Results The median follow-up time of the surviving patients was 58 months. Low ALC and an elevated AMC were both associated with poor survival rates. Receiver operating characteristic curve analysis showed that LMR was the best predictor of survival, with 4.0 as the cutoff point. Patients with LMR ≤4.0 were more likely to have an aggressive tumor, and this was associated with poor treatment responses. Patients with LMR ≤4.0 at diagnosis had significantly poorer overall survival ( OS) and progression-free survival ( PFS) than those with LMR >4.0. Multivariate analysis, which included prognostic factors of the International Prognostic Index, showed LMR ≤4.0 to be an independent predictor for the OS (hazard ratio [ HR], 2.507; 95% confidence interval [ CI], 1.255-5.007; P = 0.009) and PFS ( HR, 2.063; 95% CI, 1.249-3.408; P = 0.005). Conclusions The LMR at diagnosis, as a simple index which reflects host systemic immunity, predicts clinical outcomes in DLBCL patients treated with R- CHOP. [ABSTRACT FROM AUTHOR]

The peripheral cannabinoid receptor (CB2) is mainly detected on B cells in the germinal centers (GCs) of the immune system, using an antibody directed against the extra cellular N-terminal domain of the receptor. We retrospectively investigated the CB2 receptor expression in diffuse large B-cell lymphomas (DLBCL) and its clinical relevance for treatment outcome. We have constructed a tissue micro-array (TMA) using lymphoma tissue of a large cohort of patients with DLBCL ( N = 104) who were treated with CHOP. Forty-five out of 79 evaluable cases (57%) were CB2 positive. The expression of CB2 receptors was variably present in both the germinal center B cell (GCB) ( n = 31) and the non-GCB/activated B-cell (ABC) ( n = 43) DLBCL subtypes. CB2 positivity was not associated with a different outcome in this patient cohort (CR; P = 0.87, EFS; P = 0.32, DFS; P = 0.06 and OS; P = 0.18). Implementation of CB2 expression in the Hans algorithm using the markers CD10, BCL6, and MUM1 did not result in added prognostic value (all P-values >0.1). We hypothesize that although CB2 is normally expressed in GCs, the expression in one of the malignant counterparts such as DLBCL is aberrant. This may be an explanation for the absence of prognostic relevance for the expression of this protein. [ABSTRACT FROM AUTHOR]

Prognostic impact of peripheral blood white blood cell count (WBC) at the diagnosis of childhood acute lymphoblastic leukaemia (ALL) was evaluated in a population-based consecutive series of 2666 children aged 1-15 treated for ALL between 1992 and 2008 in the five Nordic countries (Denmark, Finland, Iceland, Norway and Sweden). Ten-year event-free (pEFS) survival and overall (pOS) survival were 0.75 ± 0.01 and 0.85 ± 0.01, respectively. Although treatment intensity was determined by WBC, non-remission and relapsed patients still had significantly higher WBC than those in remission for B-cell precursor (BCP) (median WBC: 24.8 vs. 14.0 vs. 8.3 × 10/L, P < 0.001), but not for T-lineage (T-ALL) (median WBC: 127.8 vs. 113.0 vs. 86.8 × 10/L, P = 0.22). pEFS was inversely related to WBC for BCP ( P < 0.001), but not for T-ALL. WBC was not associated with risk of event for BCP or T-ALL for patients with minimal residual disease at the end of induction (MRD) <10. In contrast, for MRD ≥ 10 and <5% leukaemic blasts in bone marrow at day 29, the pEFS for WBC < 100.0 ( N = 152) vs. ≥100.0 ( N = 19) was 0.76 vs. 0.50 ( P = 0.001). That was the case both for BCP (pEFS 0.76 vs. 0.58) and for T-ALL (pEFS 0.71 vs. 0.38). Whether the inferior EFS for the subset of patients with high WBC and slow initial response to treatment reflects rare or overlooked cytogenetic aberrations as well as the factors that determine WBC levels at diagnosis awaits exploration. [ABSTRACT FROM AUTHOR]

BLyS and APRIL share two receptors – transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) and B-cell maturation antigen (BCMA) – and BLyS binds to a third receptor, BAFF-R. We previously reported that TACI gene expression is a good indicator of a BLyS-binding receptor in human multiple myeloma cell lines (HMCLs), unlike BCMA, which is expressed by all HMCLs or BAFF-R which is typically not expressed by late-stage B cells. We hypothesised a link between APRIL and TACI through syndecan-1, similar to the situation reported for FGF and FGFR. We observed very strong binding of APRIL, but not BLyS, at the surface of all syndecan-1+ HMCLs and primary multiple myeloma cells (MMC). All syndecan-1+ HMCLs and MMC could also bind TACI-Fc, but not BCMA-Fc or BAFF-R-Fc molecules. Binding of APRIL or TACI-Fc was abrogated by heparin or cell pretreatment with heparitinase, which cleaves heparan sulfate chains. The growth factor activity of APRIL on MMC was also inhibited by heparin. Our data identify syndecan-1 as a co-receptor for APRIL and TACI at the cell surface of MMC, promoting the activation of an APRIL/TACI pathway that induces survival and proliferation in MMC. [ABSTRACT FROM AUTHOR]

B-cell lymphomas/leukemias with simultaneous t(14;18)(q32;q21) and MYC rearrangements have recently been shown to constitute a separate diagnostic entity, presenting with a rapid clinical course and a very poor prognosis. We describe the establishment of an Epstein–Barr virus negative cell line, designated U-2973, from a male patient with a de novo aggressive B-cell lymphoma/leukemia and very high peripheral blast cell count. Flow cytometry of bone marrow cells and U-2973 displayed a mature B-cell phenotype, and immunostaining showed expression of MYC and BCL2. IG gene rearrangement data were consistent with a lymphoid neoplasm of germinal centre derivation. Cytogenetic studies using conventional G-banding, fluorescent in situ hybridization, spectral karyotyping and single nucleotide polymorphism array demonstrated a complex karyotype with both a t(14;18) and double translocations between MYC and a non- IG gene partner located at chromosome 12p12.1. [ABSTRACT FROM AUTHOR]

Burkitt lymphoma/leukaemia (BL/L) is a heterogeneous disease with respect to epidemiological patterns and cell origin. The occurrence of BL/L with an immature phenotype raises the question whether this phenotype might be a consequence of early B-cell transformation or, alternatively, a secondary feature of transformed, mature B cells. It also poses important clinical questions regarding diagnosis and therapeutic procedures. Here we describe the case of a 4-yr-old child with BL/L and FAB L3 morphology, with phenotypic and genotypic characteristics of a CD10+ precursor B-cell acute lymphoid leukaemia (ALL) associated with t(8;14)(q24;q32). Molecular analysis showed expression of RAG1 and RAG2 and an unmutated VDJCμ immunoglobulin rearrangement coinciding with a lack of AICDA expression, indicating an immature B-cell origin. His clinical response suggested that FAB L3 ALL with MYC rearrangement and an aberrant precursor B-cell phenotype is clinically similar to BL/L. Moreover, short, intensive chemotherapeutic protocols seemed to be beneficial. This case also allowed us to refine the description of cellular and molecular variants of BL/L regarding the cell origin and pathogenesis of this biologically heterogeneous disease. [ABSTRACT FROM AUTHOR]

Lymphomas are currently categorized according to their origin from a B or T lymphocyte. Immature and less commonly mature (peripheral) lymphomas may harbor rearrangements of both the B- and T-cell antigen receptor genes (dual genotype or bigenotype). Rarely, cells in lymphoma with a single genotype simultaneously express both B- and T-cell markers (biphenotypic lymphomas). We discuss the diagnostic and clinical implications in the case of a 42-yr-old female with a peripheral CD30+ lymphoma that displayed both characteristic B- and T-cell surface antigens and clonal rearrangement of B- and T-cell antigen receptor gene loci. Simultaneous nuclear expression of the transcription factor gene PAX5 suggested that this major driver of B-cell differentiation did not preclude expression of CD3ε, generally assumed to be a T-cell associated antigen. [ABSTRACT FROM AUTHOR]

Molica S, Vitelli G, Levato D, Giannarelli D, Vacca A, Cuneo A, Ribatti D, Digiesi G. Serum angiogenin is not elevated in patients with early B-cell chronic lymphocytic leukemia but is prognostic factor for disease progression. Eur J Haematol 2004: 73: 36–42. © Blackwell Munksgaard 2004. The association between angiogenin and cancer progression and poor outcome in solid tumors has been documented, but its significance in leukemias has not been evaluated. Using an ELISA technique (Quantikine Human Angiogenin Immunoassay; R&D Systems), we measured serum angiogenin levels in 77 previously untreated Binet stage A B-cell chronic lymphocytic leukemia (CLL) patients. No difference in angiogenin serum levels could be found between patients (median: 295 ng/mL; range: 74–1700) and 15 age- and sex-matched healthy controls (median: 264 ng/mL; range: 29–1835) ( P = NS; Mann–Whitney test). Increased angiogenin serum level was associated with higher LDH ( P = 0.03) and β2-m ( P = 0.007) concentrations. However, angiogenin did not reflect the extent of bone marrow (BM) angiogenesis as evaluated by microvessel area ( P = 0.611), circulating levels of vascular endothelial growth factor (VEGF) ( P = 0.873) and basic fibroblastic growth factor (FGF-2) ( P = 0.421). When the 25 patients with available data were stratified into the four major cytogenetic categories (normal karyotype, 13q as a sole aberration, 12q trisomy, 11q or 17p deletion) and aberrations were compared with angiogenin serum levels, no correlation was found ( P = 0.651; Kruskall–Wallis test). A cut-off of angiogenin serum level corresponding to median (i.e. 330 ng/mL) or higher identified later upstaging and longer progression-free survival (PFS). The 5-yr PFS was 51.5% for patients with angiogenin levels lower than median and 85% for patients with higher values [ P = 0.03; hazard ratio (HR) = 2.86; 95% CI: 1.08–6.72]. Although in multivariate analysis only Rai substages ( P = 0.00001) and peripheral blood lymphocytosis ( P = 0.009) retained their prognostic significance, angiogenin could be incorporated into the Rai substages thus leading to the identification of the following risk categories: (i) stage 0 (angionenin >330 ng/mL); (ii) stage 0 (angiogenin <330 ng/mL) + stage I–II (angiogenin >330 ng/mL); and (iii) stage I–II (angiogenin <330 ng/mL). The 40-month PFS were as follows: 85%, 65%, 25% ( χ2 for trend = 6.33; d.f. = 1; P = 0.01). In conclusion, serum angiogenin levels although not increased in comparison with healthy controls, may predict clinical outcome of patients with early CLL and help to refine Rai's stratification. [ABSTRACT FROM AUTHOR]