Monoclonal antibody studies in B-cell chronic lymphocytic leukemia and allied disorders

Lymphocytes from 48 patients with B-cell chronic lymphocytic leukemia (B-CLL) and allied disorders were examined with a panel of monoclonal antibodies and conventional surface marker techniques. Surface immunoglobulin (SIg) and Ia-like antigen were regularly present on B-CLL cells. In addition, 24 of 32 specimens reacted with OKT1, a monoclonal antibody which detects both peripheral and thymic T cells, but reactivity was not observed with anti-T-cell monoclonal antibodies of more restricted specificity (OKT3, OKT4, OKT6, OKT8 or OKT11). Eighteen of 20 samples in which only SIgM was detected were OKT1-positive, while all 4 with only SIgG were OKT1-negative. Cells from patients with hairy cell leukemia were unreactive with OKT1, but resembled B-CLL lymphocytes in the presence of clonal SIg and Ia-like antigen. Neoplastic plasma cells lacked Ia-like antigen and frequently SIg (2 of 5), but cytoplasmic immunoglobulin was present, cells reacted with OKT10 (replicating lymphoid cells) and 2 of 5 with OKT9 (transferrin receptor) as well. Cutaneous T-cell lymphoma samples were reactive with the anti-T-cell monoclonal antibodies OKT1, OKT3 and OKT4 even when the sheep cell receptor could not be detected. Monoclonal antibodies can provide more certain diagnosis and superior resolution of cell lineage in these disorders than is possible by morphology alone.     

Pattern of subtypes of acute lymphoblastic leukemia in India

Leukemic cells from 124 acute lymphoblastic leukemia (ALL) and 31 chronic lymphatic leukemia (CLL) were examined for sheep erythrocyte receptor (E), surface immunoglobulin (SIg) and their reactivity with a panel of monoclonal antibodies recognizing specific surface antigens including pan-T, Common ALL and Ia antigens. In acute lymphatic leukemia, 33% of patients reveal T-cell receptor associated with higher age group, mediastinal mass and high WBC count. Common ALL was predominant between 2 and 9-yr age group. Among chronic lymphatic leukemia, 2 patients were found to be T-CLL while 29 revealed presence of SIg. Ia antigen was detected in 44.4% of ALL and 64% fo CLL patients. The pattern of surface marker observed in our series may be related to our life style, socio-economic and environmental factors.     

Cross-linking of cell surface immunoglobulins induces Epstein-Barr virus in Burkitt lymphoma lines

Anti-human immunoglobulin (Ig) antibodies with single-chain specificities induced Epstein-Barr virus (EBV) in various Burkitt lymphoma lines when the corresponding Ig chain was expressed on the cell surface. The F(ab')2 fragments of IgG antibody were as potent as intact Ig, while the Fab and Fc fragments gave no induction, indicating that cross-linking of surface Ig was required for the induction. Simultaneously with EBV induction, anti-Ig inhibited the uptake of 3H-thymidine. This inhibition was also seen in EBV-genome-negative Burkitt lymphoma lines. In contrast, no effect on virus induction and cell growth was noted in lymphoblastoid lines of non-neoplastic origin. The possible relationship between cell differentiation and latency of EBV-carrier state is discussed.     

Characterization of lymphoma-derived cell lines: comparison of cell lines positive and negative for Epstein-Barr virus nuclear antigen. I. Physical, cytogenetic, and growth characteristics

Sixteen lymphoid cell lines were derived from patients with undifferentiated lymphoma of Burkitt's or non-Burkitt's type. They were obtained directly from tumor biopsies, from serous effusions, or from bone marrow. In 10 of the cell lines, the Epstein-Barr virus (EBV) nuclear antigen (EBNA) was undetectable; the remaining 6 lines were EBNA-positive (EB-pos). Of the 16 lines, 15 were aneuploid, with detectable chromosome "14q+ markers (11 had +8;14 translocations). These 15 lines, which included the EBNA-negative (EB-neg) lines, were believed to be of tumor cell origin. The remaining line consisted predominantly of diploid cells derived from normal lymphocytes, but some cells of tumor origin were present. Four EB-pos cell lines derived from EB-neg tumors had an aneuploid karyotype consistent with an origin from tumor cells (including no.8;14 translocation in two), which suggested that either tumor cells were infected with EBV in vitro or a tiny fraction of EB-pos tumor cells (or potential tumor cells) present in vivo gave rise to the predominant cell of the line. EB-neg B-cell lines and EB-pos cell lines established from undifferentiated lymphomas differed greatly. EB-neg lines had consistently smaller electronic mean cell volumes and narrow-angle light scatter than did EB-pos lines. This finding correlated with a lower nuclear:cytoplasmic ratio in EB-pos lines. EB-neg lines also had higher saturation cell densities than did EB-pos lines under standard culture conditions. The data indicate either that EBV influences the morphologic and physiologic characteristics of lymphoid cell lines or that EB-neg B-cell lines and EB-pos cell lines are derived ultimately from different lymphocyte subpopulations or that both may apply.     

In-vitro infection of chronic lymphocytic leukemia cells by Epstein-Barr virus (EBV)

We sought to determine the potential of infecting lymphoid cells from patients with chronic leukemia (CLL) with Epstein-Barr virus (EBV) by testing for EBV receptors (EBVR) by flow cytometry, assessing for infectability of these cells by culturing with B95-8-derived virus, and staining for EB nuclear-associated antigens (EBNA) at various times post-infection. EBVR were present on 54-91% of lymphoid cells in seven cases of CLL and on 46% of prolymphocytic leukemia cells. Dynamic changes regarding EBNA positivity, morphology, and viability occurred post-infection with the virus. On day 2 only a few EBNA-positive lymphoblasts were observed. On days 11-21 positivity increased from 2 to 34% of cells. Simultaneously, the viable cell number declined to approximately 1/10th of original number. A significant proportion of the EBNA-positive cells corresponded to the original CLL cells. In 3 of 7 cases of CLL a Pan T-cell phenotype was demonstrated by Leu-1 monoclonal antibody testing. The infected cells did not react with two monoclonal antibodies, EBV-CS 1 and 4, which react with B-cell lymphoblastoid cell lines (B-LCL). Moreover, the B-LCL derived at 1-2 months post-infection of CLL cells did not express the Leu-1 antigen, but expressed EBV-CS 1 or 4 defined antigens. In the prolymphocytic leukemia, 64% of the cells showed EBNA positivity on day 7 and giant cells with huge round or multiple nuclei appeared which were EBNA-positive. CLL and prolymphocytic leukemia cells can be infected as demonstrated by EBNA-positivity. This infection does not lead to immediate transformation, but evokes lymphoblast and multinucleated giant cell production prior to the death of cells.     

Plasmacytoid blast crisis in B-cell chronic lymphocytic leukemia: effect of estradiol on growth and differentiation in vitro

Evolution of a case of chronic lymphocytic leukemia (CLL) into blast crisis was found to be characterized by three unusual features (1) the phenotype of the emerging blast cells was that of pre-plasmacytoid cells as shown by plasma cell morphology and an immunological phenotype corresponding partially with CLL- or intermediate B-cells, partially with plasma cells (terminal transferase-, common acute lymphocytic leukemia antigen-, Ia+, surface immunoglobulin heavy chains-, surface kappa light chains+, intracytoplasmic immunoglobulin A+ and G+, BA-1+, polyclonal gammaglobulin production); (2) cytogenetic analysis of spontaneous metaphases revealed that in addition to the typical CLL abnormality, trisomy 12, in all of the cells, an additional translocation between chromosomes 14 and 17 was present in 40% with a presumptive breakpoint on chromosome 14 (q12-3) never described before (commonly q32) and (3) the progression of the disease was associated with a striking increase in the expression by the transformed cells of specific binding sites for estradiol (E2) due to an actual increase in total cellular receptor proteins and not to a change in receptor affinity for E2. The functional status of the steroid receptors was confirmed by nuclear transfer of the cytoplasmic hormone-receptor complex upon temperature activation. Since the rise in E2-receptor display paralleled a large increase in the proliferative activity of the cells as well as a change in their maturation status the question was raised as to whether the E2-receptor should be considered as a physiological marker of growth rate or of cellular differentiation. Exposure of the patient's blast cells to E2 in vitro resulted in cessation of cell growth following at least one mitosis after addition of the inducer as seen from the replacement of the large blasts by small CLL-like cells without definite signs of alteration of the differentiation status. This suggests the association of E2-receptor expression with control of growth rather than cell maturation.     

Restrictions upon Epstein-Barr virus infection of the leukemic cell are demonstrated in patients with hairy cell leukemia

We tested the hypothesis that Epstein-Barr virus (EBV) might actually infect leukemic hairy cells in vivo by examining those cells for the EBV-receptor, EBV nuclear antigen (EBNA) and membrane antigen (MA), for spontaneous transformation and rescue of infectious virus and for presence of EBV genome. EBV-receptors were found on subpopulations of leukemic cells from each of 7 patients with hairy cell leukemia (HCL) tested. MA was present on low numbers (1-5 per cent) of fresh leukemic cells of 7 patients and in some instances occurred with a greater frequency after 3 to 5 days in culture, with or without 12-O-tetradecanoylphorbol-13-acetate. In 11 fresh leukemic cell preparations from 8 HCL patients, no EBNA was demonstrated. However, 2 samples after 4 days in culture expressed low frequencies of EBNA-positive cells. Spontaneous, EBV-positive cell lines were established with a high transformation efficiency from 3 HCL blood samples but not from 8 other specimens. Infectious EBV could be rescued from some hairy leukemic cell preparations by co-cultivation with cord blood lymphocytes. These results demonstrated that leukemic cell populations harbored infectious EBV, that the leukemic cells expressed virus receptors and suggested that a small subpopulation of leukemic cells might become infected in vivo at least transiently and possibly transformed in vitro by EBV. To test for the extent of occult in vivo infection of leukemic cells with EBV, Southern type hybridization studies were performed with a probe for EBV genome (Bam HI W). At a sensitivity level of 0.1 genome per cell, EBV genome was not detected in the leukemic cell populations of 7 patients. We conclude that host defence mechanisms protecting these individuals from EBV also prevent infections of the leukemic cell and/or most hairy leukemic cells are not suitable targets for both infection and transformation.     

Relationship between Epstein-Barr virus (EBV) DNA and the EBV-determined nuclear antigen (EBNA) in Burkitt lymphoma biopsies and other lymphoproliferative malignancies

Twenty-six of 27 African Burkitt lymphomas with histologically confirmed diagnosis contained relatively large amounts of EBV DNA (10–101 viral genomes per cell), as determined by nucleic acid hybridization. Twenty-five of the 26 EBV DNA-positive lymphomas contained the EBV-determined nuclear antigen, EBNA, in the majority of the nuclei. Technical reasons may have accounted for the apparent EBNA-negativity of one EBV DNA-positive biopsy. Four African lymphoma biopsies, one with a definite diagnosis of Burkitt's lymphoma and three with a questionable diagnosis of the same disease, were all EBV DNA- and EBNA-negative. The same was true for a collection of Swedish cases of Hodgkin's disease, lymphocytic lymphoma, chronic lymphatic leukemia and some other lymphoproliferative malignancies. Thus, there is excellent agreement between the presence of EBV DNA and of EBNA in tumor biopsies. The EBNA antigen test therefore appears a relatively simple way of testing for the presence of the virus genome, provided it is carried out with appropriate controls. Several of the EBV-genome and EBNA-negative cases came from patients with high serum titers of EBV antibodies. It is concluded that the virus does not really travel along with malignant lymphomas as a passenger in the seropositive patients. In comparison with other lymphomas, African Burkitt's lymphoma of the high endemic areas is unique in that the tumors (with rare exceptions) represent the proliferation of an EBV-genome carrying clone. These findings stress the necessity to distinguish between EBV-seropositive status and evidence for EBV-genome-carrying neoplastic cells.     

Isoenzyme studies in human leukemia-lymphoma cell lines--1. Carboxylic esterase

The isoenzyme patterns of carboxylic esterase (E.C. 3.1.1.1) were studied in 74 proven human leukemia-lymphoma and 12 normal B-lymphoblastoid cell lines. These cell lines have been extensively phenotyped using poly- and monoclonal antibodies. Esterase isoenzymes were separated by isoelectric focusing and visualized by histo-cytochemical techniques. No leukemia-specific or (except for monocytes) blood cell type-specific isoenzyme or isoenzyme pattern could be detected. The monocytic element in some cell lines was characterized by a strong isoenzyme band which could be selectively and completely inhibited by sodium fluoride. The enzyme phenotypes were stably expressed in all subcultures of a given cell line and did not appear to have any cell cycle dependency. The leukemia-lymphoma cell lines have been subclassified into four major groups according to immunological parameters: T-cell, B-cell, myelomonocytic and non-T, non-B-cell. On the basis of immunological data the T-cell lines were assigned to five stages of differentiation. The number and staining intensity of the isoenzymes increased with differentiation of the T-cells paralleling the expression of immunological markers. The B-cell leukemia-lymphoma cell lines were divided into pre B-, B-, Burkitt lymphoma, multiple myeloma and hairy cell leukemia cell lines. Substantial variability among the isoenzyme patterns was detected ranging from immature profiles of pre B-cell lines to complete isoenzyme repertoires of multiple myeloma cell lines. No significant difference was seen between the isoenzymes of mature B-cell lines and normal B-lymphoblastoid cell lines. The most prominent feature seen in myelomonocytic cell lines was the monocytic band indicating a monocytic origin and separating the 'monocytoid' from the 'pure myeloid' cell lines. Considerable heterogeneity in the isoenzyme patterns was observed in the non-T, non-B cell groups which comprised erythroleukemia cell lines and cell lines arrested at a very early stage of lymphoid differentiation. These latter cell lines together with some T- and B-cell lines shared the common characteristics of positivity for cALLA, TdT and Ia antigens and an immature, incomplete isoenzyme profile. The results support the notions of maturation arrest and normal gene expression in leukemic cell populations. Furthermore, the importance of biochemical studies as part of the multiple marker analysis could be demonstrated.     

Cell surface glycoprotein changes in Epstein-Barr virus-positive and -negative human hematopoietic cell lines.

It has previously been shown that differential fucose labelling of many normal and homologous tumor cells, followed by proteolytic release and degradation, yields glycopeptides which upon gel filtration shown an increase in fast-eluting glycopeptides for the tumor cells. This technique has now been applied to cell-surface glycoproteins of different human hematopoietic cell lines. These lines included Epstein-Barr virus (EBV)-carrying lymphoblastoid cell lines of presumed non-neoplastic origin, and malignant EBV-genome-positive Burkitt lymphoma and EBV-negative non-Burkitt lymphoma, leukemia and myeloma lines. As compared with normal peripheral lymphocytes, both the lymphoblastoid type of cell lines and the different types of lines of proven malignant ancestry contained the fast-eluting glycopeptides on their cell surface with very few exceptions. It is therefore concluded that (I) malignant conversion of human lymphoid cell in vivo is commonly, but not obligatorily, associated with a specific change in the composition of the fucosyl glycopeptides, and (2) EBV infection of B lymphocytes does not lead only to the well-documented immortalization in vitro but also, as a rule, to the same type of alteration in fucosyl glycopeptides as was demonstrated for the neoplastic cell lines. It proved possible to distinguish several categories of hematopoietic cell lines due to the effect that pretreatment of the glycopeptides with neuraminidase or mild acid exerted on their subsequent chromatographic behavior.     

Surface glycoprotein patterns of normal and malignant human lymphoid cells. II. B cells, B blasts and Epstein-Barr virus (EBV)-positive and -negative B lymphoid cell lines.

Surface glycoproteins of normal human B lymphocytes, B blasts and various types of lymphoid cell lines were labelled by the galactose oxidase-tritiated sodium borohydride method. The labelled glycoproteins were separated by polyacryUmide slab gel electrophoresis in the presence of sodium dodecy! sulfate and visualized by modified autoradiography (fluorography). The battery of examined hematopoietic cell lines included Epstein-Barr virus (EBV) carrying lines of proven malignant origin (Burkitt's lymphoma) and presumed non-neoplastic origin (lymphoblastoid cell lines), and EBV-genome-negative lymphocytic lymphoma, histiocytic lymphoma, myeloma and myeloid leukemia lines. The presence of possible EBV-associated surface glycoproteins, detectable by the labelling method, was studied by use of two EBV-negative cell lines (BJAB and Ramos) and their EBV-converted sublines. The six Burkitt lymphoma and three lymphocytic lymphoma lines had all the basic surface glycoprotein pattern of resting B cells and, in addition to individually distinct bands, two characteristic pairs of glycoproteins sf apparent molecular weights of 87,000/85,000 and 71,000/69,OOO. These glycoproteins were not detected on the normal B celts, B blasts or non-neoplastic lymphoblastoid lines. Neither were they found on the other types of neoptastic line. No consistent difference in the surface glycoprotein patterns was detected between the EBV-genome-negative and EBV-con-verted BJAB and Ramos sublines. The glycoprotein pattern of the six lymphoblastoid lines resembled that of the B blasts. The histiocytic lymphoma, myeloma and leukemia lines all had distinct patterns. These results confirm that the Burkitt's lymphoma and the lymphoblastoid cell lines represent two biologically distinct EBV-carrying B lymphoid celts and that the galactose oxidase NaB[³H]₄ surface labelling technique can be used as a reliable molecular mapping method to distinguish between these two and other types of lymphoid cell lines.     

Surface glycoproteins of human non-T, non-B acute lymphocytic leukemia cell lines

The surface glycoproteins of 4 human acute lymphocytic leukemia (ALL) cell lines with immunologic surface marker profiles suggestive of a non-T, non-B lymphocyte derivation (absence of surface immunoglobulin, complement and Fc receptors presence of non-T, non-B common ALL (cALL) and Ia-like antigen) have been analyzed by the galactose oxidase tritiated sodium borohydride labelling technique. The surface glycoprotein patterns have been compared with those of normal B-, T-, O-blood lymphocytes and with fresh ALL and chronic lymphocytic leukemia cells (CLL). All cALL lines express GP 42, GP 31 and GP 24, which have been identified as HLA, and Ia-like antigens, respectively. With one exception (the KM3 line) the cALL lines have GP 120 and GP 130 as major surface GPs a feature shared with fresh ALL and acute myelocytic leukemia cells. Two of the lines (NALL-1 and Nalm-1) also express a GP 210 K found as a major band on O- and B-lymphocytes and on fresh ALL cells. The dominating surface GP on KM 3 has an apparent mol. wt. of 1000,000 Daltons which is not clearly seen on other cells examined. The analyses suggest a common surface glycoprotein pattern of cALL (GP 210, GP 130, GP 120, GP 42, GP 31 and GP 24) but also re-emphasizes that cALL, as defined by immunologic surface marker analysis, is heterogenous neoplasia as the major surface GPs in KM 3 and Nalm-16 differ from those of the other cALL lines and the fresh cALL cells.     

Epstein-Barr virus DNA load in chronic lymphocytic leukemia is an independent predictor of clinical course and survival

The relation between Epstein-Barr virus (EBV) DNA load and clinical course of patients with chronic lymphocytic leukemia (CLL) is unknown. We assessed EBV DNA load by quantitative PCR at CLL presentation in mononuclear cells (MNC) of 220 prospective patients that were enrolled and followed-up in two major Institutions. In 20 patients EBV DNA load was also assessed on plasma samples. Forty-one age-matched healthy subjects were tested for EBV DNA load on MNC. Findings were validated in an independent retrospective cohort of 112 patients with CLL. EBV DNA load was detectable in 59%, and high (≥2000 copies/µg DNA) in 19% of patients, but it was negative in plasma samples. EBV DNA load was significantly higher in CLL patients than in healthy subjects (P < .0001). No relation was found between high EBV load and clinical stage or biological variables, except for 11q deletion (P = .004), CD38 expression (P = .003), and NOTCH1 mutations (P = .05). High EBV load led to a 3.14-fold increase in the hazard ratio of death and to a shorter overall survival (OS; P = .001). Poor OS was attributable, at least in part, to shorter time-to-first-treatment (P = .0008), with no higher risk of Richter''s transformation or second cancer. Multivariate analysis selected high levels of EBV load as independent predictor of OS after controlling for confounding clinical and biological variables. EBV DNA load at presentation is an independent predictor of OS in patients with CLL.     

Classification and biological nature of established human hematopoietic cell lines.

Over 200 established human hematopoietic cell lines of normal and malignant origin have been investigated by morphological and functional parameters. Employing morphology as the overriding parameter four types of lines were identified. (1) Lymphoblastoid cell lines, derived from normal and neoplastic hematopoietic tissue, were characterized by the wide morphologic flexibility of individual lymphoblastoid cells, constant association with Epstein-Barr virus (EBV), polyclonal derivation, differentiation for immunoglobulin production (secretion) and their diploids. (2) Lymphoma cell lines. This type of line was established at a high frequency from Burkitt's lymphoma and rarely from other types of lymphoma, but never from patients without malignancy or with non-lymphoma malignancies. Important characteristics were morphologic stereotypia within each line, monoclonal derivation, common but not obligatory association with EBV, variability in the expression of Ig synthesis (no production, or membrane bound Ig, or secretion) and aneuploidy. (3) Myeloma cell lines could only rarely be obtained from patients with myeloma. The basis for classification of these lines is their production of Ig identical to the myeloma protein in vitro. Other important distinguishing features were: plasma cell morphology, absence of EBV and aneuploidy. (4) The leukemia cell line (MOLT 4) was the only line with T-cell characteristics and was easily distinguished from the other types. Important characteristics were a typical surface ultrastructure, absence of EBV and absence of immunoglobulin production, Individual lymphoblastoid lines were in principle identical whereas each line of the other three types had its own characteristic profile. The phenotypic characteristics of the lymphoblastoid lines were very stable during prolonged serial cultivation. Only in a few cases were secondary chromosomal, functional or morphologic alterations noted. We conclude that EBV-carrying lymphoblastoid lines can be obtained from non-neoplastic precursor cells from healthy as well as from diseased individuals. Lymphoma, myeloma and leukemia lines are only obtained from the respective neoplastic tissue but generally at a low frequency. With the exception of Burkitt's lymphoma, malignant hematopoietic tissue and leukemia frequently give rise to established cell lines in vitro of the lymphoblastoid type rather than lines derived from the neoplastic cells;     

EBV-transformed lymphoblastoid cell lines down-regulate ebna in parallel with secretory differentiation

Four monoclonal and one polyclonal lymphoblastoid cell line (LCL) were studied with regard to cytoplasmic immunoglobulin (clg) expression, presence of the Epstein-Barr virus (EBV)-determined nuclear antigen (EBNA) and DNA synthesis. Each line was found to consist of two subpopulations, with only minimal overlap. Proliferating, EBNA-positive, clg-negative cells formed the majority. The minority were EBNA-negative, contained abundant clg and were largely non-proliferating. This suggests the continuous occurrence of a maturation process within each LCL. The concomitant downregulation of EBNA raises the interesting question whether continued synthesis of the nuclear antigen is incompatible with differentiation for epigenetic reasons, or, alternatively, whether differentiation takes place when the viral genomes are suppressed or lost.     

Antagonistic effects of c-myc and Epstein-Barr virus latent genes on the phenotype of human B cells

Epstein-Barr virus (EBV) immortalized cells and Burkitt lymphoma cells have a completely different growth pattern and phenotype. EBV immortalized cells express a set of 11 viral genes to accommodate B cell activation and proliferation, whereas EBV-positive Burkitt lymphoma cells highly express the c-myc oncogene that is activated through translocation into 1 of the immunoglobulin loci and EBNA1 as the only viral protein. We have developed a primary human B cell line conditionally immortalized by Epstein-Barr virus in which the viral gene program responsible for the induction of proliferation can be switched on and off by the addition or withdrawal of estrogen (cell line EREB2-5). Starting from this cell line we have generated 2 daughter cell lines that proliferate in a c-myc dependent fashion, 1 with a highly active exogenous c-myc gene (cell line A1) and 1 with a regulatable c-myc gene that can be switched on by withdrawal and switched off by addition of tetracycline (cell line P493-6). The comparison of the 3 cell lines has allowed us to dissect the contribution of c-myc and EBV genes to the regulation of the growth pattern and expression of cell surface molecules. We show that MYC and EBNA2 (and their respective target genes) have opposing effects on the expression of several surface markers involved in B cell activation. We show that MYC contributes to the phenotype of Burkitt lymphoma cells by upregulating CD10 and CD38 and downregulating activation markers. The phenotype of the cells is determined on one hand by the absence of the viral gene products EBNA2 and LMP1 that mediate the phenotype of activated lymphoblasts and to a lesser extent by an active contribution of the c-myc gene.     

Chromosome 14 translocation in african and north american burkitt's lymphoma

Two established North American Burkitt lymphoma cell lines were studied by chromosomal banding techniques. The SU-AmB-1 line previously shown to be negative for the Epstein-Barr virus (EBV) was found to have, among other changes, a translocation from the long arm (q) of chromosome 8 onto 14q. The SU-AmB-2 line, which contains the EBV genome, also displayed the same 8/14 translocation. These results were compared with data from three EBV-positive tumor cell lines derived from patients with African Burkitt's lymphoma. Our findings indicate that a translocation from 8q onto 14q occurs in both African and North American Burkitt lymphomas, and that this abnormality apparently is not related directly to EBV. This chromosome translocation therefore may be an important event in the development of human lymphocytic malignancy, analogous to the occurrence of the Philadelphia chromosome rearrangement in chronic myelogenous leukemia.     

The establishment of Epstein-Barr virus nuclear antigen-positive (SP-50B) and Epstein-Barr virus nuclear antigen-negative (SP-53) cell lines with t(11;14)(q13;q32) chromosome abnormality from an intermediate lymphocytic lymphoma

Two lymphoma cell lines, SP-50B and SP-53, were established from peripheral blood of a 58-year-old woman with leukemic conversion of intermediate lymphocytic lymphoma. These cell lines grew in suspension with or without forming clumps of cells. SP-50B was morphologically similar to the common Epstein-Barr (EB) virus-transformed lymphoblastoid cell lines and was positive for EB virus nuclear antigen (EBNA), whereas SP-53 closely resembled the patient's lymphoma cells and was negative for EBNA. Both cell lines expressed the same phenotypic markers as original lymphoma cells (CpIg+, SmIg+, OKIa1+, Leu12+) and possessed t(11;14)(q13;q32) chromosome translocation. These results indicate that although morphologically different, SP-50B and SP-53 were both derived from patient's lymphoma cells. The long-term cultivation of EBNA-positive and EBNA-negative B-cell lymphoma lines from a single donor has not been previously reported. These cell lines would provide useful tools for studying the oncogenic role of EB virus and bcl-1 oncogene that is located on chromosome 11q13.     

Ultrastructural expression of virus-like particles in human leukaemic cells growing in vitro

The ultrastructure of newly discovered virus-like particles in human leukaemic myeloblasts and lymphoblasts growing in vitro has been studied. Three EBNA-negative cultures contained cytoplasmic inclusion bodies which consist probably of viroplasms. After addition of 5-bromo-2-deoxyuridine (BUDR) or dimethyl sulphoxide (Me₂SO) to the cultured cells there was an approximately 50-fold increase in the number of inclusion bodies, supporting the notion that these bodies are the site of synthesis of viral structural proteins. A suggested sequence of development of inclusion bodies into virus-like particles is described and illustrated. Similar inclusions have been reported to date only in fresh human leukaemic cells but not in cell cultures. The availability of these particles for further studies will help to determine whether they play a role in the aetiology of human leukaemia. EBNA-positive human haemic cell lines studied did not contain similar intracytoplasmic inclusion bodies, and treatment of two of them with BUDR did not produce inclusion bodies.