Giemsa banding patterns in chronic lymphocytic leukaemia.

The banding patterns of chromosomes from 20 patients with chronic lymphocytic leukaemia (C.L.L.) have been analyzed. 97 of 100 metaphases examined had a normal banding pattern. The 3 remaining metaphases, all from one patient had bands similar to those seen after aging. It is concluded that the chromosomes in C.L.L. have normal banding patterns. The majority of cytogenetic studies in chronic lymphocytic leukaemia have reported normal chromosomes (Fitzgerald and Adams 1965; Oppenheim et al., 1965; Lawler et al., 1968). An inherited abnormality of G group chromosome (No. 22) has been reported in a family, three members of whom developed C.L.L. (Fitzgerald and Hamer, 1969), but further investigations of cases of familial leukaemia failed to reveal a similar abnormality (Fitzgerald et. al., 1966). The development of new techniques which allow the positive identification of individual chromosomes (Caspersson et al., 1969; Dutrillaux and Lejeune, 1971; Sumner et al., 1971; Seabright, 1971), has revolutionised human cutogenetics and revealed additional information regarding chromosome abnormalities and leukaemia (Rowley, 1973; Lobb et al., 1972; Milligan and Garson, 1974). The purpose of this investigation was to determine whether the chromosomes in C.L.L. have normal banding patterns.     

Karyotypes of 33 patients with clonal aberrations in chronic lymphocytic leukaemia. Review of 216 abnormal karyotypes in chronic lymphocytic leukaemia

We report on 33 unpublished patients with clonal anomalies in chronic lymphocytic leukaemia. The literature was thoroughly reviewed in order 1) to quantify the frequency of anomalies found in chronic lymphocytic leukaemia and to give new status to the rarest, 2) to determine whether a given anomaly was an additional anomaly and/or a primary anomaly, and 3) to find out whether strong associations between different anomalies exist in this disease.     

Development of chronic lymphocytic leukaemia after posttraumatic splenectomy

In the course of a post-splenectomy follow-up study, 2 out of 102 patients who had been splenectomized after an abdominal trauma were found to have developed chronic lymphocytic leukaemia 5 and 31 years after splenectomy, respectively. The possible association between splenectomy and secondary leukaemia is discussed.     

Different maturation capabilities of chronic lymphocytic leukaemia lymphocytes in vitro

Peripheral blood lymphocytes from five patients with B-derived chronic lymphocytic leukaemia were stimulated by Staphylococcus aureus strain Cowan together with T cell mitogen phytohaemagglutinin in 5-9 days suspension cultures. The responses of B lymphocytes were studied on a T cell depleted subpopulation, obtained from harvested lymphocyte cultures using the sheep red blood cell rosette technique. Proliferation tests were performed using a 3H-TdR blast cell index. The maturation process of B-lymphocytes was examined with cytoplasmic Ig studied by FITC-conjugated antisera. Results analysed indicate various degrees of maturation of B cells in different patients.     

Apoptosis induces Bcl-XS and cleaved Bcl-XL in chronic lymphocytic leukaemia

The Bcl-X gene has both pro-survival, Bcl-X_L, and pro-apoptotic, Bcl-X_S, gene products, which are produced by alternative splicing. The function of these proteins has previously been characterised in cell lines, often by transfecting expression constructs, and primary cell systems capable of dynamically regulating Bcl-X_L and Bcl-X_S have not been described. Such a system is potentially important to allow testing of agents that promote apoptosis by increasing the amount of Bcl-X_S at the expense of Bcl-X_L. In this report we characterise Bcl-X gene products in primary human leukaemic B-cells in culture conditions associated with survival and apoptosis. We found that Bcl-X_S was induced in spontaneous and drug-induced apoptosis and that apoptosis induced in cells cultured on mouse fibroblasts expressing CD40 ligand with IL-4 (CD154/IL-4), a condition mimicking the tissue microenvironment, additionally produced expression of cleavage products of Bcl-X_L. Both Bcl-X_S and Bcl-X_L were produced in a caspase dependent manner. We tested emetine, an agent previously reported to increase Bcl-X_S but found that it did not have this effect in primary human B-cells. Therefore, there are two mechanisms—cleavage of Bcl-X_L and production of Bcl-X_S—by which Bcl-X gene products could enhance apoptosis in CLL but neither appeared to have a primary role in inducing leukaemic cell death.     

The coexistence of chronic lymphocytic leukaemia and polycythaemia vera terminating in acute myeloid leukaemia

A 67-year-old man with the coexistence of CLL and PV converted after 4 years to AML is described. This rare simultaneous occurrence of both chronic lymphoid and myeloid proliferations as well as nonlymphoblastic leukaemia developing in a patient with CLL is discussed in the light of literature data.     

Immunohistochemical analysis of ZAP-70 expression in chronic lymphocytic leukemia

This paper shows a protocol for the detection of ZAP-70 expression in B-CLL (B cell chronic lymphocytic leukemia) tumor cells by common immunohistochemical methods. The study was conducted on bone marrow trephine biopsies from 62 B-CLL patients at the time of diagnosis. Immunohistochemical reactions based on peroxidase and alkaline phosphatase reactions were used, as well as double immunofluorescent labeling for ZAP-70 detection as an indirect marker of mutated and unmutated CLL. Clinical relevance of the ZAP-70 expression detection method was assessed using χ ² test between ZAP-70 positivity data and other known prognostic factors, i.e., clinical and cytogenetics data. ZAP-70 was detected in 13 out of 62 patients. Statistically significant results were obtained for ZAP-70 positive cases and known indicators of worse prognosis. Immunohistochemical analysis supported by double immunfluorescent labeling, as shown here, is an easy and reliable technique for the detection of ZAP-70 expression in B-CLL tumor cells applicable in every hematopathology laboratory.     

Cytokinesis arrest and multiple centrosomes in B cell chronic lymphocytic leukaemia

Cytokinesis failure leads to the emergence of tetraploid cells and multiple centrosomes. Chronic lymphocytic leukaemia (CLL) is the most common haematological malignancy in adults and is characterized by clonal B cell expansion. Here, we show that a significant number of peripheral blood CLL cells are arrested in cytokinesis and that this event occurred after nuclear envelope reformation and before cytoplasmic abscission. mRNA expression data showed that several genes known to be crucial for cell cycle regulation, checkpoint and centromere function, such as ING4, ING5, CDKN1A and CDK4, were significantly dysregulated in CLL samples. Our results demonstrate that CLL cells exhibit difficulties in completing mitosis, which is different from but may, at least in part, explain the previously reported accumulation of CLL cells in G0/1.     

Expression and distribution of aphidicolin-induced fragile sites in chronic myeloid leukaemia, acute lymphocytic leukaemia and acute myeloid leukaemia

New information is revealed concerning the frequency of expression and distribution of aphidicolin-induced fragile sites in eight leukaemic patients, namely, four chronic myeloid leukaemic patients (CML), three acute lymphocytic leukaemic (ALL) patients, and one acute myeloid leukaemic (AML) patient. The cytogenetic data demonstrate a statistically significant (p less than 10(-6] increase in the frequency of aphidicolin-induced fragile sites in seven of the eight leukaemic patients compared with healthy age-matched and sex-matched controls. The chromosomal band locations of the aphidicolin-induced fragile sites from 400 metaphase spreads of these leukaemic patients reveal a nonrandom distribution in the karyotype. Some aphidicolin-induced fragile sites in these leukaemic patients were located at chromosome bands known to be induced specifically by folic acid, distamycin A, bromodeoxyuridine or azacytidine. The cross-induction of fragile sites in the leukaemic patients may be indicative of shared molecular homology in the sequence composition of nonrandom chromosomal DNA.     

Apoptosis in B-chronic lymphocytic leukaemia

B-chronic lymphocytic leukaemia (B-CLL) is characterised by the progressive accumulation of monoclonal B cells, which may be the result of several factors leading to extended B-CLL cell lifespan, increased proliferative capacity and diminished cell death. Here we review the implications of several signals mediated by receptors, such as surface IgM, CD6 and CD40, for the B-CLL cell survival, together with data on gene modulation in relation to the apoptosis process in B-CLL cells. We also describe some features of the Fas/FasL system in B-CLL that hypothetically might contribute to the accumulation of leukaemic cells and the progressions of the disease, by downregulating the apoptotic response or avoiding the autologous immune response.     

Comprehensive characterization of immunoglobulin gene rearrangements in patients with chronic lymphocytic leukaemia

Previous studies have suggested a geographical pattern of immunoglobulin rearrangement in chronic lymphocytic leukaemia (CLL), which could be as a result of a genetic background or an environmental antigen. However, the characteristics of Ig rearrangements in the population from the South of France have not yet been established. Here, we studied CLL B‐cell repertoire and mutational pattern in a Southern French cohort of patients using an in‐house protocol for whole sequencing of the rearranged immunoglobulin heavy‐chain genes. Described biased usage of variable, diversity and joining genes between the mutated and unmutated groups was found in our population. However, variable gene frequencies are more in accordance with those observed in the Mediterranean patients. We found that the third complementary‐determining region (CDR) length was higher in unmutated sequences, because of bias in the diversity and joining genes usage and not due to the N diversity. Mutations found in CLL followed the features of canonical somatic hypermutation mechanism: preference of targeting for activation‐induced cytidine deaminase and polymerase motifs, base change bias for transitions and more replacement mutations occurring in CDRs than in framework regions. Surprisingly, localization of activation‐induced cytidine deaminase motifs onto the variable gene showed a preference for framework regions. The study of the characteristics at the age of diagnosis showed no difference in clinical outcome, but suggested a tendency of increased replacement and transition‐over‐transversion mutations and a longer third CDR length in older patients.     

Bone marrow histopathologic patterns and immunologic phenotype in B-cell chronic lymphocytic leukaemia

The present work analyzes the clinicobiological and immunological characteristics - the latter hitherto unexplored - of the different bone marrow histopathological patterns of the B-cell chronic lymphocytic leukaemia (B-CLL). In addition, we studied whether any or some of these parameters were able to predict the probability of a particular pattern of bone marrow involvement appearing. Of the 100 B-CLL cases studied 41 had a diffuse pattern and 59 were non-diffuse - interstitial 27, nodular 11 and mixed 21 -. Neither clinical nor immunological differences were observed among the distinct non-diffuse patterns. The patients in the diffuse group displayed an increased incidence of mu+ isotype and a higher proportion of HLA-DR and HAN-PC 1 positive cells while, conversely, reactivity with the FMC 8 McAb was lower. In addition, patients with a diffuse pattern of BM involvement displayed features of a more extensive disease: a higher incidence of adenopathies (p less than 0.05), hepatomegaly (p less than 0.01), splenomegaly (p less than 0.01), anaemia (p less than 0.01) and thrombopenia (p less than 0.01) as well as higher levels of peripheral blood lymphocytosis (p less than 0.05) and a higher percentage of BM lymphocytic infiltration (p less than 0.001). Multiple regression analysis showed that thrombopenia and splenomegaly were the two most important features in predicting the probability of a diffuse pattern.     

Stromal control of cystine metabolism promotes cancer cell survival in chronic lymphocytic leukaemia

Tissue stromal cells interact with leukaemia cells and profoundly affect their viability and drug sensitivity. Here we show a biochemical mechanism by which bone marrow stromal cells modulate the redox status of chronic lymphocytic leukaemia (CLL) cells and promote cellular survival and drug resistance. Primary CLL cells from patients exhibit a limited ability to transport cystine for glutathione (GSH) synthesis owing to a low expression level of Xc-transporter. In contrast, bone marrow stromal cells effectively import cystine and convert it to cysteine, which is then released into the microenvironment for uptake by CLL cells to promote GSH synthesis. The elevated level of GSH enhances leukaemia cell survival and protects them from drug-induced cytotoxicity. Furthermore, disabling this protective mechanism significantly sensitizes CLL cells to drug treatment in the stromal environment. This stromal-leukaemia interaction is critical for CLL cell survival and represents a key biochemical pathway for effectively targeting leukaemia cells to overcome drug resistance in vivo.