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PubMedDate not supplied

Chronic lymphocytic leukemia (CLL) in younger adults: a retrospective study of 133 cases.

Clinical and biological data have been evaluated, using both univariate and Cox's multivariate statistical analyses, in a series of 133 Chronic Lymphocytic Leukemia (CLL) patients with a mean age of 46.6 years (range 31-50). In univariate analyses, anemia (Hb less than 13 g/dl), peripheral blood (PB) lymphocytosis (greater than 40 x 10(9)/l) and bone marrow (BM) lymphocytosis (greater than 80 per cent) were shown to be of significant prognostic value. Multivariate analysis, through a forward stepwise procedure, showed that the most important and independent variable is the BM lymphocytosis. These results are different from those obtained in previous studies and particularly in a recent identical study performed by the same Cooperative Group on 1777 patients with a mean age of 64.2 years (Mandelli et al., 1987). No significance can be demonstrated in stratifying this series of younger patients according to different staging methods (Rai et al., 1975; Binet et al., 1981b; Mandelli et al., 1987). Therefore this population of CLL patients, with less than 50 years of age, has risk factors quite different from classical CLL. The results of the present study show that the diagnostic approach to B-CLL in younger adults must be more complete: using the common diagnostic criteria, established staging systems appear to be inadequate in this series of younger patients.

Adult↗

PubMed2002

Significance of phosphotyrosine proteins, Bcl-2 and p53 for apoptosis in resting B-chronic lymphocytic leukemia (CLL) cells.

Signal transduction and apoptosis in B-cell chronic lymphocytic leukemia (CLL) cells with a post-germinal center (GC) phenotype were studied. Specific activation of the cells was induced by a combination of soluble anti-CD40 monoclonal antibody and interleukin-4 (CD40/IL-4) and nonspecific activation with a combination of phytohemagglutinin, phorbol-12-myristate-13-acetate and ionomycin (chemical mixture). Less than 5% of these leukemia cells entered the cell cycle after activation, as indicated by the number of cells in G0/G1 phase. The protein tyrosine phosphorylation pattern and expression of the Bcl-2 protein were specific in ex vivo CLL cells of each individual patient. Expression of the p53 protein was not detectable in these leukemia cells. Cross-linking of the CD40/IL-4 receptors on CLL cells significantly upregulated phosphotyrosine proteins and the p53 protein. In the presence of chemical mixture, downregulated phosphotyrosine proteins were detected. Alterations in Bcl-2 expression were independent of cross-linking with CD40/IL-4 or chemical mixture. A high frequency of apoptotic cells was detected in cells that had downregulated phosphotyrosine proteins and Bcl-2 protein. There was no correlation between induction of apoptosis and expression of p53 protein. Our results suggest that apoptosis in resting leukemia cells could occur prior to the cell cycle progression. Alterations in phosphotyrosine proteins and Bcl-2 but not p53 might play an important role in the regulation of apoptosis in resting G0/G1 memory post-GC B-CLL cells.

Antibodies, Monoclonal↗

PubMed1995

Epstein Barr virus (EBV)-carrying cells of a chronic lymphocytic leukemia (CLL) subpopulation express EBNA1 and LMPs but not EBNA2 in vivo.

We have previously described an exceptional CLL patient, P.G., whose leukemic cell population contained a small fraction of Epstein-Barr virus (EBV)-carrying cells. These cells grow directly into permanent cell lines in vitro. Using RT-PCR analysis, we now show that the in vivo EBV-carrying CLL cells expressed EBNAI, LMPI, LMP2a and 2b, but not EBNA2, in 4 of 4 blood samples obtained during the last 3 years of the patient's life. Our data also show that the CLL cells used a promoter in the F/Q, but not the W or C, region. This is consistent with the fact that CLL cells resemble resting lymphocytes rather than immunoblasts. Expression of LMP1 and LMP2b differs from the exclusive EBNAI and LMP2a expression of normal resting B cells, however, and corresponds to the state defined as latency II. This form of latency was until now detected only in EBV-carrying non-B cells in vivo. Our data show that a B-cell subtype can also show this expression pattern in vivo.

Antigens, Viral↗

PubMed1998

Assignment of 48 ESTs to chromosome 13 band q14.3 and expression pattern for ESTs located in the core region deleted in B-CLL.

An expression map containing 48 ESTs was constructed to identify a tumor-suppressor gene involved in B-cell chronic lymphocytic leukemia (B-CLL), which was previously assigned to chromosome band 13q14.3 close to genetic markers D13S25 and D13S319. Thirty-nine of these 48 ESTs, together with 11 additional ones listed in databases, were initially assigned to chromosome 13q14 between markers D13S168 and D13S176. Nine others have recently been located in the D13S319 region. Our results indicate that 48 of the 59 ESTs analyzed belong to a YAC contig of chromosome 13 band q14, and 22 are contained on YAC 933e9, which encompasses the B-CLL critical region. Ten of these 22 ESTs were accurately assigned on a PAC, BAC, and cosmid contig encompassing the smallest minimal deletion area described so far in B-CLL, and 20 were tested for their expression on 27 normal or tumor tissues. One EST appears to be a likely candidate for the tumor-suppressor gene involved in B-CLL.

Chromosome Mapping↗

PubMed1989

B cells from chronic lymphocytic leukemia (CLL) patients are strong inducers of proliferation and major histocompatibility complex (MHC)-unrestricted [natural killer (NK)-like] cytotoxicity in normal T-lymphocytes.

We studied the ability of chronic lymphocytic leukemia (CLL) B cells to stimulate proliferation and major histocompatibility complex (MHC)-unrestricted [natural killer (NK)-like] cytotoxicity in a mixed lymphocyte tumor-cell culture (MLTC). CLL-derived B cells induced a significantly higher proliferative response than did B cells from healthy normal donors. Comparable levels of interferon and interleukin-2 production were detected in the mixed lymphocyte cultures (MLC) set up with normal B cells from healthy donors and in MLTC. Higher levels of NK-like cytotoxic activity were induced after stimulation with CLL than with normal B cells in nonlytic precursors. Inhibition experiments with specific monoclonal antibodies indicate that the NK-like response in MLTC is attributable to HLA class II antigens, which are expressed at comparable levels on CLL and normal B cells. Percoll gradient centrifugation of the MLC and MLTC recovered cells demonstrated that the NK-like effectors in both types of cultures were blast-transformed, highly mitotic cells.

B-Lymphocytes↗

PubMed2003

In vitro evaluation of 213Bi-rituximab versus external gamma irradiation for the treatment of B-CLL patients: relative biological efficacy with respect to apoptosis induction and chromosomal damage.

External source radiotherapy and beta radioimmunotherapy (RIT) are effective treatments for lymphoid malignancies. The development of RIT with alpha emitters is attractive because of the high linear energy transfer (LET) and short path length, allowing higher tumour cell kill and lower toxicity to healthy tissues. We assessed the relative biological efficacy (RBE) of alpha RIT (in vitro) compared to external gamma irradiation with respect to induction of apoptosis in B chronic lymphocytic leukaemia (B-CLL) and induction of chromosomal damage in healthy donor B and T lymphocytes. The latter was measured by a micronucleus assay. 213Bi was eluted from a 225Ac generator and conjugated to CD20 antibody (rituximab) with CHX-A"-DTPA as a chelator. B-CLL cells from five patients were cultured for 24 h in RPMI/10% FCS while exposed to 213Bi conjugated to CD20 antibody or after external 60Co gamma irradiation. Binding assays were performed in samples of all patients to calculate the total absorbed dose. Apoptosis was scored by flow cytometric analyses of the cells stained with annexin V-FITC and 7-AAD. Apoptosis was expressed as % excess over spontaneous apoptosis in control. Full dose range experiments demonstrated 213Bi-conjugated CD20 antibody to be more effective than equivalent doses of external gamma irradiation, but showed that similar plateau values were reached at 10 Gy. The RBE for induction of apoptosis in B-CLL was 2 between 1.5 and 7 Gy. The micronucleus yield in lymphocytes of healthy volunteers was measured to assess the late toxicity caused by induction of chromosomal instability. While gamma radiation induced a steady increase in micronucleus yields in B and T cells, the damage induced by 213Bi was more dramatic, with RBE ranging from 5 to 2 between 0.1 Gy and 2 Gy respectively. In contrast to gamma irradiation, 213Bi inhibited mitogen-stimulated mitosis almost completely at 2 Gy. In conclusion, high-LET targeted alpha particle exposure killed B-CLL cells more effectively than did external gamma irradiation at a low dose (RBE=2), while a plateau was reached at a high dose. Long-term toxicity on healthy B and T lymphocytes was systematically higher for the alpha emitter (RBE=5 to 2).

Aged↗

PubMed2006

The expression of NK cell inhibitory receptors on cytotoxic T cells in B-cell chronic lymphocytic leukaemia (B-CLL).

Immune surveillance of tumours is mediated by cytotoxic T cells (CTL) that recognise tumour antigen. Reduced reactivity of CTL towards tumour cells could thus lead to disease progression and loss of tumour control. In B-cell chronic lymphocytic leukaemia (B-CLL), the function of tumour-reactive CTL seems to correlate inversely to disease stage. Inhibitory NK cell receptors are known to suppress the CTL response upon interaction with major histocompatibility complex (MHC) class I and increased expression of such receptors on CTL may inhibit the anti-tumour response. So, the aim of this study was to investigate the expression of NK cell inhibitory receptors on CTL in B-CLL patients and if such expression correlated to disease stage. CD8+ T cells from B-CLL patients in Binet stage A (n = 26) and stage C (n = 14) and healthy controls (n = 14) were analysed for the expression of killer immunoglobulin-like receptors (KIR) CD158a (KIR2DL1), CD158b (KIR2DL2), CD158e (KIR3DL1) and the C-type lectin receptor CD94, by flow cytometry analysis. Patients with advanced disease (Binet stage C) had a significantly greater percentage of CTL expressing CD158b, CD158e and CD94 than patients with non-progressive disease (Binet stage A) and healthy controls. Stage C patients also had a significantly higher percentage of CTL expressing CD158a than stage A patients. No statistically significant differences were found between Binet A patients and healthy controls. Our results suggest that increased expression of KIR and CD94 on CTL in advanced stage B-CLL may potentially contribute to the impaired anti-tumour immune response in these patients.

Adult↗

PubMed1997

Relationship between autoimmunity and immunodeficiency in CLL.

B-CLL patients have a high prevalence of autoimmune phenomena due to polyclonal autoantibodies (Abs) restricted to blood cell self-antigens (Ag) and progressively develop a severe hypogammaglobulinemia. Autoimmunity and immunodeficiency have been related to B-CLL biological properties and cellular origin. Three major issues have emerged: 1) the relevance of B cell receptor abnormalities. More specifically, the importance of the defective expression of CD79b which may explain the faint to virtually undetectable amounts of monoclonal sig, the anomalous signal transduction and the failure to present antigens (Ag). 2) the possibility that the relentless accumulation of B-CLL cells is also under microenvironmental influences. These are brought about by the interplay of cytokines produced through cell/cell contacts among malignant B cells, macrophages and T cells. 3) the view that the very properties of accumulating anergic self-reactive CD5+ B lymphocytes may provide a bridge between autoimmunity and immune incompetence. CD5+ malignant B cells influence the cytokine production by T cells, but are inefficient Ag presenting cells (APC). As such they are a hurdle to the production of normal Ab. However, by stimulating in vitro the CD40 molecule on the membrane of CLL cell with the CD40 ligand the inefficient APC is turned into an efficient APC. In vivo, such an activation may conceivably occur in specific environments like the spleen and favour the production by normal residual B-cells of polyclonal autoAb against red cell or platelet self Ag.

Animals↗

PubMed1999

Detection of minimal residual disease in B chronic lymphocytic leukemia (CLL).

UNLABELLED: In the absence of specific chromosomal translocations the best method for detecting minimal residual disease (MRD) in B cell malignancies is based on the uniqueness of immunoglobulin (Ig) genes rearrangement. We here report a very sensitive method for assessing MRD in complete hematological remission (CHR) chronic lymphocytic leukemia (CLL) patients as defined by the international workshop on CLL (IWCLL). PATIENTS: Twelve CLL patients in CHR and complete phenotypic remission (CPR) were included in the study. Eight of them received Fludarabine (FDR), one was treated by Chop regimen, and the remaining 3 were rescued by polychemotherapy followed by autologous bone marrow transplantation (ABMT). METHODS: DNA extracted from peripheral blood lymphocytes (PBL) of each patient was amplified with VH family specific and framework 3 primers in 5' and a consensus JH primer in 3', before treatment and sequentially after the CPR completion. When no clonal rearrangement could be detected by this assay, the CDR3 sequence specific probe of the clone was used as the 3' primer, associated to the VH family specific primer in 5'. PCR products were analyzed by classical procedures in agarose and/or acrylamide gels. RESULTS: Mixtures of leukemic cells and normal PBL showed detection of a single leukemic cell among more than 10(5) normal cells. Four out of the 12 patients achieved molecular remission (MR) when employing CDR3 amplification. All 3 autografted patients were in MR, whereas only one out of the 9 patients treated by chemotherapy alone achieved MR. When using a clone specific probe, a clonal signal was observed in all cases but one (ABMT). Results presented here confirm that MR may be achieved in a few cases of B-CLL. Further studies are needed to determine the exact relationship between MRD and clinical outcome.

Aged↗

PubMed2005

EBV infection induces expression of the transcription factors ATF-2/c-Jun in B lymphocytes but not in B-CLL cells.

B cell type chronic lymphocytic leukaemia (B-CLL) cells carry the Epstein-Barr virus (EBV) receptor CD21 and can be infected in vitro with the virus. The infected cells exhibit an unusual EBV program, they express the nuclear proteins but not latent membrane protein 1 (LMP-1). Similar cells were encountered in lymphoid tissues of infectious mononucleosis (IM) patients and in lymphoproliferations of immunosuppressed patients. EBV infected B-CLL cells can be regarded as model for this viral program. In B cells the regulation of LMP-1 is executed mainly by EBV encoded nuclear antigen 2 (EBNA-2), interacting with several cellular proteins and these complexes bind to specific sequences in the LMP-1 promoter. ATF2 and c-Jun were shown to be among the interacting partners of EBNA-2. These molecules can be detected in experimentally infected B lymphocytes. We found c-Jun and/or phosphorylated ATF-2 (p-ATF-2) expression in some B-CLL ex vivo samples. They disappeared or their expression declined promptly in explanted cells, even if they were infected with EBV in vitro. Activation of the infected B-CLL cells by exposure to CD40L was accompanied by p-ATF-2 and c-Jun but not by LMP-1 expression. In one of three clones tested, subsequent treatment with histone deacetylase inhibitors (HDACi), TSA or n-butyrate, could induce LMP-1. Treatment with phorbol-12, 13-dibutyrate (PDB) induced LMP-1 expression in three of four clones. Neither the HDACi nor the PDB treated cells survived.

Activating Transcription Factor 2↗

PubMed1987

Acid hydrolases in B-chronic lymphocytic leukemia (B-CLL): a comparison with normal peripheral B lymphocytes and normal B-cell subset with the phenotype of B-leukaemic cells.

The acid phosphatase (AP), beta-glucuronidase (BG), and alpha-naphthyl-acetate esterase (ANAE) distribution and the morphology of stage 0 B-CLL lymphocytes were studied. The results were compared with the same hydrolase equipment and morphology of normal B-cell populations showing the B-CLL-like phenotype, and thus regardable as the possible counterpart of leukaemic cells. The functional and structural characters of the former were strikingly different from those of the latter. In fact the normal B-cell population with B-CLL-like phenotype showed an homogeneous enzyme pattern with predominant ANAE and a strictly lymphocytic cell morphology. In contrast, the leukaemic cells showed various and unrelated morphological and cytochemical features, thus forming apparent subgroups of B-CLL. The development of an irregular "switch on" mechanism in the blocked malignant cell clone, may account for these functional and structural maturation discrepancies. Moreover, the fact that in the leukaemic cells ANAE activity could be demonstrated only after the appearance of the other hydrolases, makes it a marker of functionally differentiated B lymphocytes.

Acid Phosphatase↗

PubMed2006

Monoclonal antibody combinations in CLL: evolving strategies.

Monoclonal antibodies have led to a profound shift in the therapeutic landscape of chronic lymphocytic leukemia (CLL). Alemtuzumab and rituximab remain the most active antibodies, and their single-agent activity has been established in previously untreated and relapsed patients with CLL. Higher response rates and a better quality of response through eradication of minimal residual disease have been reported with monoclonal antibody combinations. Chemoimmunotherapy regimens are being actively explored in frontline CLL therapy, and numerous combination regimens have been investigated in relapse. New and more effective therapies are shifting the focus from palliation to treatment algorithms with curative attempt. Challenges for monoclonal antibody combinations in the future include: (1) defining appropriate patient populations for combination therapies; (2) assessing the impact of pretreatment biologic prognostic factors; (3) enhancing eradication of minimal residual disease; and (4) reassessing response criteria in CLL.

Alemtuzumab↗

PubMed2006

Burkitt t(8;14)(q24;q32) and cryptic deletion in a CLL patient: report of a case and review of literature.

A 53-year-old man diagnosed with chronic lymphocytic leukemia (CLL)-small lymphoma following splenectomy was found to have a t(8;14) with an apparent cryptic deletion of the MYC gene. This patient's spleen and bone marrow (BM) showed that 93% and approximately 70% of the viable cells, respectively, were lambda-monoclonal B-cells coexpressing CD5 with CD20, CD19, CD23, CD22, CD38, and low FMC-7. The smear showed a marked increase in small, mature lymphoid cells, with <2% prolymphocytes. The BM karyotype was 46,XY,t(8;14)(q24;q32),-18,+mar[3]/46,XY[27] and FISH analysis with an IGH/MYC green-red dual-fusion signal probe showed an atypical interphase result of one fusion, two green, and one red signal in 70% of the cells. The MYC dual red-green split-apart probe showed the expected t(8;14) pattern in 62% of the cells; however, sequential FISH on a t(8;14) GTG-metaphase showed the single fusion signal on derivative chromosome 8 and only a green signal on der(14) for the IGH/MYC dual-fusion probe and a green signal on der(14), red signal on der(8), and fusion signal on the normal chromosome 8 for the MYC split-apart probe. Thus, the apparently balanced t(8;14) had a cryptic deletion (approximately 1.6 Mb) between the red and the green regions flanking the MYC gene in the MYC split-apart probe, 128,585,631-130,226,339 bp from 8pter. The rarity of t(8;14) in CLL together with a cryptic deletion that probably includes the MYC gene in our CLL patient persuaded us to explore the clinicopathological role of MYC translocations by comparing disease progression in our patient and in other reported CLL cases.

Burkitt Lymphoma↗

PubMed2005

Surface and cytoplasmic immunoglobulin expression in B-cell chronic lymphocytic leukemia (CLL).

Flow cytometric analysis of abnormal lymphocyte populations in chronic lymphocytic leukemia (CLL) has been widely reported to show weak expression of surface immunoglobulin (sIg). The international scoring system to help discriminate between CLL and other B-cell lymphoproliferative disorders lists this as the first of 5 criteria worth 1 point each. In the present study, 30 cases of CLL were studied for surface and cytoplasmic Ig expression. 23 of these 30 (76.7%) cases were positive for sIg. Of these 23, 14 cases (60.9%) showed moderate to bright sIg expression. All of these 23 cases were positive for CD5 and CD23; all were negative for CD10 and only 6 (26.1%) were positive for FMC7. 27 of 30 (90.0%) cases expressed cytoplasmic immunoglobulin (cIg) compared to 5% reported by others. This shows that cytoplasmic Ig occurs in a much greater percentage of cases than reported previously. 23 of 30 (76.7%) cases showed positivity for both surface and cytoplasmic Ig, with 15 showing kappa light chain restriction and 8 showing lambda light chain restriction. Six expressed cytoplasmic Ig only, with 4 showing kappa light chain restriction and 2 showing lambda light chain restriction. One case expressed neither cytoplasmic nor surface Ig. CD38 positivity portends a worse prognosis. Of the 29 cases tested for CD38, 13 (44.8%) were positive. Of these 13 cases, 12 were in the surface Ig/cytoplasmic Ig+ group and 1 in the cytoplasmic Ig+ group only. Also, of the 23 cases tested for CD22 expression, 16 (69.6%) were positive. These data question the use of both "weak" surface Ig expression and lack of CD22 expression as valid scoring criteria for CLL.

Cell Membrane↗

PubMed2003

Influence of CD40 ligation on survival and apoptosis of B-CLL cells in vitro.

Modulation of signal transduction pathways represents a promising approach for altering the biological behavior of hematopoetic malignancies. The cells of chronic lymphocytic leukemia were treated in vitro with CD40-ligand or IL-4 to explore their effects on survival and sensitivity to apoptosis induced by Fluda. The expression of G1 cell cycle regulatory proteins was also measured. Stimulation via CD40-CD40L resulted in increased viability, as did stimulation with IL-4. A combination of the two stimulators (CD40L plus IL-4) induced increased expression of cyclins D3 and E, pRb phosphorylation and downregulated p27. Cdk2 and Cdk4 activities were not detected. It seems that this combination induced also some progression in the cell cycle. Furthermore, Fluda-induced apoptosis was not prevented by CD40L, IL-4, or a combination of both agents, although a delay in the onset of apoptosis was observed. Taken together, these results support the view that CD40L and IL-4 sustain B-cell chronic lymphocytic leukemia (B-CLL) survival by different pathways and their synergistic action might induce cell cycle progression in B-CLL. The exposure of B-CLL to CD40L, IL-4 or both did not impair the sensitivity of B-CLL to Fluda. CD40L and IL-4 postponed apoptosis induced by Fluda, depending on a fashion of administration.

Apoptosis↗

PubMed2003

Simvastatin induces apoptosis of B-CLL cells by activation of mitochondrial caspase 9.

BACKGROUND AND OBJECTIVES: Chronic lymphocytic leukemia (CLL) is the most common leukemia in the western world. Despite several advances in therapeutic options, the disease remains incurable. Recently, it was repeatedly demonstrated that statins, competitive inhibitors of 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase, have antineoplastic effects. Therefore we aimed to study the effects of simvastatin (Sim) on malignant B cells derived from patients with CLL and mechanisms of action of the drug. METHODS AND RESULTS: Purified B-CLL cells from 15 patients were cultured either alone or with Sim at concentrations of 10, 50, and 100 microM. Viability, measured by the activity of mitochondrial dehydrogenases, was reduced significantly in the cells treated with Sim at 50 and 100 microM for 24 hours (p<0.005). The level of apoptosis, as measured by annexin binding to exposed phosphatidylserine moieties, increased significantly in the treated cells at concentrations higher than 50 microM for 24 hours (p<0.003). The level of necrosis, as measured by propidium iodide internalization, increased significantly after 24 hours exposure to Sim at 50 microM (p<0.01). The apoptotic cascade was studied by immunoblot analysis of caspases following Sim treatment. These showed cleavage of caspases 9, 8, and 3. Addition of the caspase inhibitor Z-VAD.fmk inhibited caspase 8 and 3 significantly but did not affect caspase 9. CONCLUSION: Exposure of clonal B lymphocytes from patients with CLL to simvastatin decreases viability significantly by the induction of apoptosis. The apoptosis induced by Sim is probably initiated by the mitochondrial caspase 9, which indirectly leads to activation of caspase 3 and 8.

Apoptosis↗

PubMed2002

Factor(s) released from irradiated B-CLL cells induce apoptosis in leukemic lymphocytes.

Photon irradiation of peripheral blood lymphocytes from 25 patients with untreated B-chronic lymphocytic leukemia (B-CLL) induced an increase in apoptotic response by 270%. No significant increase in apoptosis was observed after irradiation of peripheral blood mononuclear cells from 15 healthy volunteers. Supernatants (sups) derived from irradiated leukemic cells incubated with non-irradiated autologous cells induced a 75% enhancement in number of apoptotic cells, as compared with sups from non-irradiated CLL cells. The level of tumor necrosis factor alpha, a cytokine known to prevent apoptosis, was reduced in the sups of irradiated CLL cells in comparison to that of non-irradiated lymphocytes. The interleukin (IL)-10 level, an IL reported to induce apoptosis, was similar in the sups of irradiated and non-irradiated lymphocytes from B-CLL patients. No change in IL-2 levels was observed. The significance of these findings and the role of factor(s) in the sups of irradiated leukemic lymphocytes as inducers of apoptosis are discussed.

Aged↗

PubMed1997

Relationship between nitrogen mustard drug resistance in B-cell chronic lymphocytic leukemia (B-CLL) and protein expression of Bcl-2, Bax, Bcl-X and p53.

Several genes have been implicated in the regulation of apoptosis including bcl-2, bax, bcl-X and p53. These genes may be important in the development of nitrogen mustard (NM) drug resistance in B-cell chronic lymphocytic leukemia (B-CLL). Using Western blot analysis, we examined the levels of Bcl-2, Bax, Bcl-X and p53 protein expression and determined whether the levels of these proteins correlated with in vitro drug resistance in CLL patients' lymphocyte samples. Our investigations suggest that in CLL, NM drug resistance develops without any detectable alteration of Bcl-2, Bax or Bcl-X. In addition, we determined the presence of p53 mutations in 14 samples in order to assess if there is an association between in vitro drug resistance and the presence of p53 mutations. Using single-stranded conformational polymorphism (SSCP) and sequencing analysis, we observed a p53 mutation in two out of seven resistant samples. The mutation occurring in both cases was a G:C --> A:T transition at codon 273 (exon 8). One of these cases was de novo resistant to the nitrogen mustards. Only one of six samples with acquired resistance to the nitrogen mustards had a p53 mutation suggesting that p53 mutations are not a prominent feature of acquired NM resistance in CLL.

Aged↗