The role of alternative splicing of the adhesion molecule, CD44, in lymphoid malignancy.
AIM: To investigate the expression of CD44 isoforms containing variant exon 6 (v6) in a well characterised cohort of patients with non-Hodgkin's lymphoma (NHL) and chronic lymphocytic leukaemia (CLL), and to correlate this with phenotype and disease course. METHODS: Cryostat sections of OCT embedded diagnostic nodal material from NHL patients and cryopreserved mononuclear preparations from CLL patients were used as sources of RNA. After reverse transcription, PCR was carried out with amplimers positioned at either side of the variant exon insertion site to amplify all possible CD44 isoforms. Those isoforms containing v6 were identified after Southern blotting and hybridisation with a radiolabelled oligonucleotide. RESULTS: Of 32 NHL samples analysed, 16 did not express CD44 isoforms containing v6, six expressed an isoform containing exon v6 alone, and 10 expressed v6 long isoforms which contained exon v6 in addition to other variant exons. These data did not correlate with lymphoma classification, disease staging, or the presence or absence of extranodal disease. However, those patients expressing v6 long CD44 isoforms had a worse overall survival than those that did not. The plateau of the survival curves was 50% compared with 82%. No v6 long isoforms were detected in the 21 CLL samples investigated. CONCLUSIONS: The expression of v6 long CD44 isoforms is associated with aggressive disease in NHL, independent of grade, stage, or presence of extranodal disease. (+info)
Elevated serum thymidine kinase levels identify a subgroup at high risk of disease progression in early, nonsmoldering chronic lymphocytic leukemia.
Chronic lymphocytic leukemia (CLL) shows a remarkably heterogeneous clinical outcome; survival ranges from several months in advanced stages to more than 10 years in early stages. The Binet and Rai staging systems distinguish three major prognostic subgroups, but do not accurately predict the individual risk of disease progression in early CLL (Binet stage A or Rai stage 0 to II). Because most newly diagnosed CLL patients present with early disease, it seems desirable to search for additional prognostic factors to identify early CLL patients at high risk of rapid progression. It has been shown that elevated serum thymidine kinase (s-TK) levels predict disease progression in CLL. Therefore, this study aimed to assess the prognostic value of s-TK in 122 previously untreated patients with Binet stage A CLL (mean age +/- SD, 58.7 +/- 8.5 years). In univariate analyses, 18 of the 22 parameters investigated predicted progression-free survival (PFS). In a stepwise multiple regression analysis, only three parameters provided independent prognostic information on PFS: s-TK greater than 7.1 U/L; presence of lymphadenopathy; and white blood cell (WBC) count greater than 75, 000/microL. When added to the classification of smoldering versus nonsmoldering CLL, s-TK levels separated two groups within the group of nonsmoldering stage A patients: patients with s-TK values greater than 7.1 U/L had a median PFS of 8 months, whereas patients with s-TK values +info)
CD40-activated B-cell chronic lymphocytic leukemia cells for tumor immunotherapy: stimulation of allogeneic versus autologous T cells generates different types of effector cells.
Although spontaneous remissions may rarely occur in B-cell chronic lymphocytic leukemia (B-CLL), T cells do generally not develop a clinically significant response against B-CLL cells. Because this T-cell anergy against B-CLL cells may be caused by the inability of B-CLL cells to present tumor-antigens efficiently, we examined the possibility of upregulating critical costimulatory (B7-1 and B7-2) and adhesion molecules (ICAM-1 and LFA-3) on B-CLL cells to improve antigen presentation. The stimulation of B-CLL cells via CD40 by culture on CD40L expressing feeder cells induced a strong upregulation of costimulatory and adhesion molecules and turned the B-CLL cells into efficient antigen-presenting cells (APCs). CD40-activated B-CLL (CD40-CLL) cells stimulated the proliferation of both CD4(+) and CD8(+) T cells. Interestingly, stimulation of allogeneic versus autologous T cells resulted in the expansion of different effector populations. Allogeneic CD40-CLL cells allowed for the expansion of specific CD8(+) cytolytic T cells (CTL). In marked contrast, autologous CD40-CLL cells did not induce a relevant CTL response, but rather stimulated a CD4(+), Th1-like T-cell population that expressed high levels of CD40L and released interferon-gamma in response to stimulation by CD40-CLL cells. Together, these results support the view that CD40 activation of B-CLL cells might reverse T-cell anergy against the neoplastic cell clone, although the character of the immune response depends on the major histocompatibility complex (MHC) background on which the CLL or tumor antigens are presented. These findings may have important implications for the design of cellular immunotherapies for B-CLL. (+info)
Rituximab therapy in hematologic malignancy patients with circulating blood tumor cells: association with increased infusion-related side effects and rapid blood tumor clearance.
PURPOSE: Rituximab was recently approved for use in relapsed, low-grade non-Hodgkin's lymphoma; however, few data exist regarding the safety of this agent in patients with a high number of tumor cells in the blood. METHODS AND RESULTS: After the observation at our institution of a rapid reduction of peripheral-blood tumor cells with associated severe pulmonary infusion-related toxicity in two patients with refractory hematologic malignancies, data on three additional cases were collected from physician-submitted reports of adverse events related to rituximab treatment. Five patients with hematologic malignancies possessing a high number of blood tumor cells were treated with rituximab and developed rapid tumor clearance. The median age was 68 years (range, 26 to 78 years). Patients were diagnosed with B-cell prolymphocytic leukemia (n = 2), chronic lymphocytic leukemia (n = 2), or transformed non-Hodgkin's lymphoma (n = 1). All of these patients had bulky adenopathy or organomegaly. All five patients developed a unique syndrome of severe infusion-related reactions, thrombocytopenia, rapid decrement in circulating tumor cell load, and mild electrolyte evidence of tumor lysis, and all required hospitalization. In addition, one patient developed ascites. These events resolved, and four patients were subsequently treated with rituximab without significant complications. CONCLUSION: Rituximab administration in patients who have a high number of tumor cells in the blood may have an increased likelihood of severe initial infusion-related reactions. These data also suggest that rituximab may have activity in a variety of other lymphoid neoplasms, such as chronic lymphocytic leukemia and B-cell prolymphocytic leukemia. (+info)
Multicenter phase II study of fludarabine phosphate for patients with newly diagnosed lymphoplasmacytoid lymphoma, Waldenstrom's macroglobulinemia, and mantle-cell lymphoma.
PURPOSE: Fludarabine phosphate (F-AMP) has significant activity in follicular lymphoma and in B-cell chronic lymphatic leukemia, where it has demonstrated high complete response (CR) rates. Lymphoplasmacytoid (LPC) lymphoma, Waldenstrom's macroglobulinemia (WM), and mantle-cell lymphoma (MCL) also present with advanced-stage disease and are incurable with standard alkylator-based chemotherapy. A phase II trial was undertaken to determine the activity of F-AMP in patients newly diagnosed with these diseases. PATIENTS AND METHODS: Between 1992 and 1996, 78 patients (aged 18 to 75 years) received intravenous F-AMP (25 mg/m2/d for 5 days, every 4 weeks) until maximum response, plus two further cycles as consolidation. The primary end point was response rate; secondary end points included time to progression (TTP), duration of response, and overall survival (OS). RESULTS: Forty-four (62%) of 71 assessable patients had a response to F-AMP (LPC lymphoma, 63%; WM, 79%; MCL, 41%); the CR rate was 15%. At a median follow-up of 1.5 years, 19 of 44 responding patients have had progression of lymphoma; the median duration of response was 2.5 years. The median survival has not yet been reached. There was no significant difference in the duration of response or OS between patients with different histologies; TTP was shorter in patients with MCL (P = .015). Myelosuppression was relatively common, and the treatment-related mortality (TRM) was 5%, mostly associated with pancytopenia and infection. CONCLUSION: Single-agent fludarabine phosphate is active in previously untreated LPC lymphoma and WM, with only moderate activity in MCL. However, the CR rate is low, and the TRM is relatively high. Its role in combination chemotherapy remains to be demonstrated. (+info)
An alternatively spliced form of CD79b gene may account for altered B-cell receptor expression in B-chronic lymphocytic leukemia.
Several functional anomalies of B-chronic lymphocytic leukemia (B-CLL) cells may be explained by abnormalities of the B-cell receptor (BCR), a multimeric complex formed by the sIg homodimer and the noncovalently bound heterodimer Igalpha/Igbeta (CD79a/CD79b). Because the expression of the extracellular Ig-like domain of CD79b has been reported to be absent in the cells of most CLL cases, we have investigated the molecular mechanisms that may account for this defect. Peripheral blood lymphocytes (PBL) from 50 patients and two cell lines (MEC1, MEC2) obtained from the PBL of one of them were studied. MEC1, MEC2, and 75% of CLL cases did not express detectable levels of the extracellular Ig-like domain of CD79b, which was nevertheless present in greater than 80% CD19(+) cells from normal donors. In healthy subjects the expression of CD79b was equally distributed in CD5(+) and CD5(-) B-cell subsets. Reverse transcription-polymerase chain reaction (RT-PCR) analysis of CD79b RNA from all patients and from MEC1 and MEC2 cell lines consistently yielded two fragments of different size (709 bp and 397 bp). The 709-bp band corresponds to CD79b entire transcript; the 397-bp band corresponds to an alternatively spliced form lacking exon 3 that encodes the extracellular Ig-like domain. Both fragments were also visible in normal PBL. The expression of the 397-bp fragment was increased in normal activated B cells, while no difference was seen between CD5(+) and CD5(-) B cells. To obtain a more accurate estimate of the relative proportions of the two spliced forms, a radioactive PCR was performed in 13 normal and 22 B-CLL samples and the results analyzed using a digital imager. The mean value of the CD79b to the CD79b internally deleted ratio was 0.64 +/- 0.20 SD in normal donors and 0.44 +/- 0.27 SD in B-CLL (P =.01). Direct sequencing of 397-bp RT-PCR products and of genomic DNA corresponding to exon 3 from MEC1, MEC2, their parental cells, and five fresh B-CLL samples did not show any causal mutation. Single-strand conformation polymorphism analysis of exon 3 performed in 18 additional B-CLL cases showed a single abnormal shift corresponding to a TGT --> TGC polymorphic change at amino acid 122. We propose a role for the alternative splicing of CD79b gene in causing the reduced expression of BCR on the surface of B-CLL cells. As normal B cells also present this variant, the mechanism of CD79b posttranscriptional regulation might reflect the activation stage of the normal B cell from which B-CLL derives. (+info)
Fludarabine and epirubicin in the treatment of chronic lymphocytic leukaemia: a German multicenter phase II study.
PURPOSE: Fludarabine has been reported to be the most effective single-agent in previously treated chronic lymphocytic leukaemia (CLL). Based on the in vitro synergism of fludarabine with anthracyclines and on results showing a higher efficacy of CHOP against COP we attempted to improve treatment results with a combination of fludarabine and an anthracycline. PATIENTS AND METHODS: The aim of the multicenter study was to evaluate the rate and duration of remissions and investigate the toxic and immunosuppressive effects of fludarabine and epirubicin in the treatment of CLL in Binet stages B and C as first-line therapy or in first relapse. Thirty-eight patients were treated with fludarabine 25 mg/m2 on days 1-5 and epirubicin 25 mg/m2 on days 4 and 5. RESULTS: The overall response rate (OR) was 82% (95% confidence interval (95% CI): 66%-92%) with a CR rate of 32% (95% CI: 18%-49%). For the 25 previously untreated patients the OR was 92% (95% CI: 74%-99%) including 40% CRs (95% CI: 21%-61%). Granulocytopenia grade 3 occurred in 23% of all evaluable cycles, and grade 4 in 17%. The median remission duration was 19 months (range 6-37 months). CONCLUSION: The results show that the combination of fludarabine and epirubicin is tolerable and highly effective in the treatment of CLL. With the addition of epirubicin to fludarabine, it appears possible to achieve a higher response rate and a more rapid response, especially of nodal manifestations. This regimen can be administered in an outpatient facility except for the first cycle because of the risk of a tumour lysis. The possible benefit of the combination presented here in the treatment of CLL in comparison to single-agent fludarabine treatment is presently under study in a prospective randomized multicenter study. (+info)
Constitutive expression of CD26/dipeptidylpeptidase IV on peripheral blood B lymphocytes of patients with B chronic lymphocytic leukaemia.
We have investigated the expression of the ectoenzyme dipeptidylpeptidase IV (DPP IV)/CD26 on lymphocytes obtained from patients with B chronic lymphocytic leukaemia (B-CLL) and compared it with healthy subjects. Using two-colour immunofluorescence analysis with CD26 and CD20 or CD23 monoclonal antibodies, CD26 was found undetectable on peripheral resting B-cells (CD20+ CD23-) from normal donors whereas it was expressed on B-cells activated in vitro with interleukin (IL)-4 and Staphylococcus aureus strain cowan I (CD20+ CD23+). The expression of CD26 on leukaemic B-cells (CD20+ CD23+) was clearly induced in 22 out of 25 patients examined. Consequently, induced levels of CD26 cell surface expression on either normal activated and malignant B-cells coincided with the enhancement of DPP IV activity detected on the surface of these cells. Reverse transcription polymerase chain reaction analyses showed that the transcript levels of the CD26 gene was higher in normal activated B-cells and B-CLL cells than in resting B-cells, suggesting that CD26 was expressed at the level of transcriptional activation. These observations provide evidence of the abnormal expression of DPPIV/CD26 in B-CLL which, therefore, may be considered as a novel marker for B-CLL. Further investigation in relation to CD26 expression and other B malignancies needs to be defined. (+info)