Laboratory of B Cell Neoplasia

Milano, Italy

Laboratory of B Cell Neoplasia

Milano, Italy
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Rawstron A.C.,Leeds Teaching Hospitals | Rawstron A.C.,University of York | Bottcher S.,University of Kiel | Letestu R.,AP HP | And 17 more authors.
Leukemia | Year: 2013

Detection of minimal residual disease (MRD) in chronic lymphocytic leukaemia (CLL) is becoming increasingly important as treatments improve. An internationally harmonised four-colour (CLR) flow cytometry MRD assay is widely used but has limitations. The aim of this study was to improve MRD analysis by identifying situations where a less time-consuming CD19/CD5/κ/λ analysis would be sufficient for detecting residual CLL, and develop a six-CLR antibody panel that is more efficient for cases requiring full MRD analysis. In 784 samples from CLL patients after treatment, it was possible to determine CD19/CD5/κ/λ thresholds that identified cases with detectable MRD with 100% positive predictive value (PPV). However, CD19/CD5/κ/λ analysis was unsuitable for predicting iwCLL/NCI response status or identifying cases with no detectable MRD. For the latter cases requiring a full MRD assessment, a six-CLR assay was designed comprising CD19/CD5/CD20 with (1) CD3/CD38/CD79b and (2) CD81/CD22/CD43. There was good correlation between four-CLR and six-CLR panels in dilution studies and clinical samples, with 100% concordance for detection of residual disease at the 0.01% (10-4) level (n=59) and good linearity even at the 0.001-0.01% (10-5-10-4) level. A six-CLR panel therefore provides equivalent results to the four-CLR panel but it requires fewer reagents, fewer cells and a much simpler analysis approach. © 2013 Macmillan Publishers Limited All rights reserved.

Bertilaccio M.T.S.,Laboratory of Lymphoid Malignancies | Scielzo C.,Laboratory of Lymphoid Malignancies | Scielzo C.,University of Milan | Simonetti G.,Laboratory of B Cell Neoplasia | And 14 more authors.
Blood | Year: 2010

Easily reproducible animal models that allow for study of the biology of chronic lymphocytic leukemia (CLL) and to test new therapeutic agents have been very difficult to establish. We have developed a novel transplantable xenograft murine model of CLL by engrafting the CLL cell line MEC1 into Rag2 -/-γc-/- mice. These mice lack B, T, and natural killer (NK) cells, and, in contrast to nude mice that retain NK cells, appear to be optimal recipient for MEC1 cells, which were successfully transplanted through either subcutaneous or intravenous routes. The result is a novel in vivo model that has systemic involvement, develops very rapidly, allows the measurement of tumor burden, and has 100% engraftment efficiency. This model closely resembles aggressive human CLL and could be very useful for evaluating both the biologic basis of CLL growth and dissemination as well as the efficacy of new therapeutic agents. © 2010 by The American Society of Hematology.

Fazi C.,Laboratory of B Cell Neoplasia | Scarfo L.,Laboratory of B Cell Neoplasia | Scarfo L.,Lymphoma Unit | Scarfo L.,University of Ferrara | And 17 more authors.
Blood | Year: 2011

Monoclonal B-cell lymphocytosis (MBL) is classified as chronic lymphocytic leukemia (CLL)-like, atypical CLL, and CD5 -MBL. The number of B cells per microliter divides CLL-like MBL into MBL associated with lymphocytosis (usually detected in a clinical setting) and low-count MBL detected in the general population (usually identified during population screening). After a median follow-up of 34 months we reevaluated 76 low-count MBLs with 5-color flow cytometry: 90% of CLL-like MBL but only 44.4% atypical CLL and 66.7% CD5 - MBL persisted over time. Population-screening CLL-like MBL had no relevant cell count change, and none developed an overt leukemia. In 50% of the cases FISH showed CLL-related chromosomal abnormalities, including monoallelic or biallelic 13q deletions (43.8%), trisomy 12 (1 case), and 17p deletions (2 cases). The analysis of the T-cell receptor β (TRBV) chains repertoire showed the presence of monoclonal T-cell clones, especially among CD4 highCD8 low, CD8 highCD4 low T cells. TRBV2 and TRBV8 were the most frequently expressed genes. This study indicates that (1) the risk of progression into CLL for low-count population-screening CLL-like MBL is exceedingly rare and definitely lower than that of clinical MBL and (2) chromosomal abnormalities occur early in the natural history and are possibly associated with the appearance of the typical phenotype. © 2011 by The American Society of Hematology.

Fazi C.,Laboratory of B Cell Neoplasia | Dagklis A.,Laboratory of B Cell Neoplasia | Cottini F.,Laboratory of B Cell Neoplasia | Scarfo L.,Laboratory of B Cell Neoplasia | And 5 more authors.
Cytometry Part B - Clinical Cytometry | Year: 2010

Background: Monoclonal B cell lymphocytosis (MBL) is a preclinical condition characterized by an expansion of clonal B cells in the absence of B lymphocytosis (BALC < 5 × 109/L) in the peripheral blood, without clinical signs, suggestive of a lymphoproliferative disorder. B cell clonal expansions are also associated with hepatitis C virus (HCV) infection and they can evolve into lymphoproliferative disorders such as mixed cryoglobulinemia and non-Hodgkin lymphomas (NHL). The relationship between MBL and HCV infection has not been established yet. Methods: By five-colour flow cytometry, we analyzed 123 HCV positive subjects with diagnosis of chronic hepatitis (94) or cirrhosis (29); 16 of those with cirrhosis had a diagnosis of hepatocellular carcinoma. Results: MBL were identified in 35/123 (28.5%), at significantly higher frequency than in the general population. Sixteen/thirty-five were atypical-chronic lymphocytic leukemia (CLL) MBL (CD5+, CD20bright), 13/35 were CLL-like MBL (CD5 bright, CD20dim), and 6/35 were CD5- MBL. Twenty-four/ninety-four (25.5%) patients affected by chronic hepatitis had MBL, whereas 11/29 (37.9%) patients with cirrhosis showed a B cell clone. A biased usage of IGHVgenes similar to HCV-associated NHL was evident. Conclusions: All three types of MBL can be identified in HCV-infected individuals at a higher frequency than in the general population, and their presence appears to correlate with a more advanced disease stage. The phenotypic heterogeneity is reminiscent of the diversity of NHL arising in the context of HCV infection. The persistence of HCV may be responsible for the dysregulation of the immune system and in particular of the B cell compartment. © 2010 International Clinical Cytometry Society.

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