Human T-cell Leukemia-lymphoma Virus Research Articles

Transformation of NIH 3T3 cells by a human c-sis cDNA clone.

The mechanism of leukaemogenic transformation by human T-cell leukaemia/lymphoma virus (HTLV), a retrovirus implicated in the aetiology of certain adult T-cell leukaemias and lymphomas, is unknown but is conceivably associated with the expression of the cellular analogues of retroviral oncogenes. The HUT-102 cell line, derived from a cutaneous T-cell lymphoma and infected with HTLV, expresses several cellular oncogenes. It is unusual among haemopoietic cell lines in that one of these is c-sis, the gene from which the oncogene v-sis of the simian sarcoma virus was derived, and perhaps the gene for platelet-derived growth factor (PDGF). To explore the possible role of c-sis expression in HTLV-induced disease, we have obtained cDNA clones of c-sis from HUT-102 cells. Here we describe two such clones and report that one of them transforms NIH-3T3 cells. This is the first example of transformation of NIH-3T3 cells by a human onc gene other than c-ras or Blym, as well as the first demonstration of transformation by a human cDNA clone.

IMMUNE T CELLS REACTIVE AGAINST HUMAN T-CELL LEUKAEMIA/LYMPHOMA VIRUS

Six patients with malignant disorders associated with human T-cell leukaemia/ lymphoma virus (HTLV) were studied to see whether long-term cultures of immune T cells reactive against HTLV-infected tumour cells could be achieved. Immune T-cell lines could not be developed from the cells of five patients who died or eventually had a relapse of disease, but in one patient who had an unusually long remission of his disease after therapy, immune T-cell lines were propagated that could produce their own T-cell growth factor and proliferate upon stimulation with autologous tumour cells and also specifically lyse HTLV-infected target cells. These immune T cells recognised the presence of circulating HTLV-bearing neoplastic cells in another patient with HTLV-associated T-cell leukaemia, who had been in clinical remission after chemotherapy when this study started, thereby providing early evidence of relapse.

Lymphokine production by cultured human T cells transformed by human T-cell leukemia-lymphoma virus-I.

Cell-free conditioned media from human T cells transformed by human T-cell leukemia-lymphoma virus (HTLV-I) were tested for the production of soluble biologically active factors, including several known lymphokines. The cell lines used were established from patients with T-cell leukemia-lymphoma and from human umbilical cord blood and bone marrow leukocytes transformed by HTLV-I in vitro. All of the cell lines liberated constitutively one or more of the 12 biological activities assayed. These included macrophage migration inhibitory factor (MIF), leukocyte migration inhibitory factor (LIF), leukocyte migration enhancing factor (MEF), macrophage activating factor (MAF), differentiation inducing factor (DIF), colony stimulating factor (CSF), eosinophil growth and maturation activity (eos. GMA), fibroblast activating factor (FAF), gamma-interferon and, in rare instances, T-cell growth factor (TCGF). Some cell lines produced interleukin 3 (IL-3), platelet-derived growth factor (PDGF), or B-cell growth factors (BCGF). Such cells should prove useful for the production of lymphokines and as sources of specific messenger RNA's for their genetic cloning.

Functional Properties of Neoplastic T Cells in Adult T Cell Lymphoma/Leukemia Patients from the Caribbean

The neoplastic T cells from five patients with adult T cell lymphoma/leukemia (ATLL), born in the Caribbean, were studied with respect to immunoregulatory activity on pokeweed mitogen (PWM) driven immunoglobulin (Ig) synthesis as well as surface-marker phenotypes with monoclonal antibodies. The neoplastic T cells in all patients had an OKT1+4+8-11+M1-I1-3A1- phenotype, but differed in the reactivity with OKT3. None of the patients' cells exerted helper activity on PWM-induced Ig synthesis. The neoplastic cells of three patients had suppressor activity on PWM-induced Ig synthesis. All patients were positive for human T cell leukemia/lymphoma virus (HTLV) or had antibodies against HTLV antigens. It has previously been shown that the neoplastic cells in Japanese ATLL patients and in patients from the Caribbean are indistinguishable by morphology and marker phenotype. We now show them to be also similar with respect to their functional properties.

Functional properties of neoplastic T cells in adult T cell lymphoma/leukemia patients from the Caribbean

The neoplastic T cells from five patients with adult T cell lymphoma/leukemia (ATLL), born in the Caribbean, were studied with respect to immunoregulatory activity on pokeweed mitogen (PWM) driven immunoglobulin (Ig) synthesis as well as surface-marker phenotypes with monoclonal antibodies. The neoplastic T cells in all patients had an OKT1+4+8–11+M1-I1–3A1- phenotype, but differed in the reactivity with OKT3. None of the patients' cells exerted helper activity on PWM- induced Ig synthesis. The neoplastic cells of three patients had suppressor activity on PWM-induced Ig synthesis. All patients were positive for human T cell leukemia/lymphoma virus (HTLV) or had antibodies against HTLV antigens. It has previously been shown that the neoplastic cells in Japanese ATLL patients and in patients from the Caribbean are indistinguishable by morphology and marker phenotype. We now show them to be also similar with respect to their functional properties.

DEMONSTRATION OF VIRAL ANTIGEN p24 IN CIRCULATING IMMUNE COMPLEXES OF TWO PATIENTS WITH HUMAN T-CELL LEUKAEMIA/LYMPHOMA VIRUS (HTLV) POSITIVE LYMPHOMA

Circulating immune complexes from two patients with human T-cell leukaemia/ lymphoma virus (HTLV)-related lymphoma were shown to contain the major internal antigen of the virus, p24. The amount of complex-bound p24 in sequential serum samples correlated roughly with tumour cell mass. Small amounts of complex-bound p24 were detected in samples before a relapse became clinically manifest. Measurement of complex-bound p24 in patients with HTLV-associated lymphomas and leukaemias might thus be helpful in management of malignancies and offer the possibility of detecting imminent relapse and preventing it by intensification of treatment.

Transformation of different phenotypic types of human bone marrow T-lymphocytes by HTLV-1.

Fresh bone marrow mononuclear leukocytes were used as a target for infection by human T-cell leukemia-lymphoma virus subgroup 1 (HTLV-1) by co-cultivation with virus-positive cell lines. The lines were established from T-cell leukemia-lymphoma patients or HTLV-1-transformed human umbilical cord-blood T cells. Clumps of transformed cells became visible by 2-3 weeks after infection and developed at a high frequency (greater than 90%) in the bone marrow samples used. Stimulation of target cells with lymphocyte mitogens facilitated this process but was not absolutely required. Unlike fresh or cultured cells from HTLV-1-positive adult leukemia-lymphoma patients and HTLV-1-transformed cord-blood T cells, which usually have an OKT 4/leu 3a surface phenotype, the transformed bone marrow cells frequently fell into one of three categories based on their reactivity with cell-specific monoclonal antibodies. These included populations of cells that were predominantly; (1) OKT 4/leu 3a-positive, (2) OKT 8/leu 2a-positive, and (3) cells expressing neither phenotype. Fresh bone marrow provided a rapid and efficient system for the assessment of HTLV-1 infection. The type of bone marrow cells transformed in vitro suggests that HTLV-1 can infect several subsets of T lymphocytes, possibly including immature cells or that these cells can undergo phenotypic modulation.

Immunological characterization of the low molecular weight gag gene proteins p19 and p15 of human T-cell leukemia-lymphoma virus (HTLV) and demonstration of human natural antibodies to them

Small molecular weight gag gene proteins p19 and p15 of human T-cell leukemia virus (HTLV) were purified to homogeneity from density banded virus, using differential affinities to phosphocellulose at pH 7.9 and 6.5. Using specific radioimmunoassays, p19 and p15 were shown to band with HTLV on sucrose gradients. These proteins were present only in HTLV-I and in cell lines expressing HTLV-I. They were not expressed in a B-cell line from patient CR, although his T cells were HTLV producers. No significant immunological cross-reactivity was detected between these proteins and HTLV-II MO P19 and p15 did not share any common antigenic determinants with each other or with HTLV p24 in competition radioimmunoassays. Sera of HTLV-positive leukemia-lymphoma patients and some of their relatives also had natural antibodies to p19 and p15.

Phenotypic and functional characterization of HTLV positive neoplastic T cells cultured with interleukin-2 — I. Retention of morphology, phenotype and selective cytotoxic properties in long term culture

T cells from the peripheral blood of a T-cell chronic lymphocytic leukemia (T-CLL) patient, cultured in the presence of interleukin-2 (IL-2), were found to express the p19 structural core protein of the human T-cell leukemia/lymphoma virus (HTLV) and to release type C virus particles. Comparison of the T-CLL cell line with the original leukemic T cells revealed that both the fresh and the proliferating T-CLL cells were pleomorphic cells that showed a convoluted nucleus and formed rosettes with sheep erythrocytes (E-rosettes). They were reactive with the monoclonal antibodies OKT1, OKT4 and OKT11, but not with OKT3, OKT6 or OKT8, indicating that they were mature T cells but that they differed from normal T cells in their lack of reactivity with OKT3. In addition they did not bind peanut agglutinin or OKM-1, and were negative for Epstein-Barr nuclear antigen, surface immunoglobulin, non-specific esterase activity or Fc- or complement receptors. Part of the fresh T-CLL cells reacted with a monoclonal antibody recognizing HLA-DR antigens (p29, 34) (36%) and with anti-Tac (62%), a monoclonal antibody directed at the IL-2 receptor, indicating that the T-CLL cells were partially activated already in vivo. After culture in vitro all proliferating T-CLL cells expressed HLA-DR and Tac antigens. The fresh T-CLL cells were found to be defective in cell-mediated lympholysis (CML) generated in mixed lymphocyte culture (MLC), antibody-dependent cellular cytotoxicity (ADCC) and lectin-dependent cellular cytotoxicity (LDCC). In addition they failed to exhibit natural killer (NK) cell activity against targets that are usually very susceptible to lysis, such as K562, but were able to kill two tumor-derived cell lines, the melanoma NKI-4 and the neuroblastoma CHP-100. The same pattern of selective killing was observed using the proliferating T-CLL cells as effectors, or cloned T-CLL cultures obtained from them by limiting dilution procedures. Therefore, it was concluded that the T-CLL cells represented a clonal expansion of neoplastic T cells that retained their phenotype and cytotoxic properties after culture in vitro.

Phenotypic and functional characterization of HTLV positive neoplastic T cells cultured with interleukin-2 — II. Inhibition of lymphoproliferative responses

T cells from the peripheral blood of a T-cell chronic lymphocytic leukemia (T-CLL) patient, were expanded by culture in the presence of interleukin-2 (IL-2). In contrast to the original leukemic T cells that showed a complete lack of suppressor activity, the proliferating T-CLL cells were extremely potent inhibitors of mitogen, antigen and alloantigen induced lymphoproliferative responses. Compared to the fresh T-CLL cells, the proliferating T-CLL cells showed an enhanced expression of the human T-cell leukemia/lymphoma virus (HTLV) p19 antigen and released type C virus particles into their culture supernatant. A direct involvement of the virus particles in the suppression was however unlikely, since the inhibition was found to be mediated by a soluble inhibitor in the culture supernatant of proliferating T-CLL cells. This inhibitor was very hydrophobic and was inactivated by treatment with trypsin, by heating to 56°C for 2 h and by storage at −70°C for more than 14 weeks. It could be excluded that the suppression of lymphoproliferation was due to competition for IL-2, to toxic effects, to nutrient depletion or to a shift in the kinetics of the target cell responses. Furthermore, it could not be attributed to suppressor inducer activity of the OKT4+ proliferating T-CLL cells, since normal T cells enriched for OKT4+ T cells and depleted for OKT8+ T cells were also inhibited in their proliferation. Since other hemopoietic cell lines, not of OKT4+ T lymphocyte origin, also were suppressed to proliferate, it is concluded that the proliferating T-CLL cells represent a neoplastic T-cell clone that had differentiated into suppressor effector T cells after prolonged culture in the presence of IL-2.

Antibodies against three purified structural proteins of the human type‐C retrovirus, HTLV, in Japanese adult T‐cell leukemia patients, healthy family members, and unrelated normals

The immunological relationship of human T-cell leukemia/lymphoma virus (HTLV) and the virus found in Japanese adult T-cell leukemia/lymphoma (ATL) was investigated in detail by testing the specific binding of serum antibodies from Japanese ATL patients and normal Japanese donors to the purified HTLV proteins p24, p19, and p15. Sera were prescreened for antibodies to p24. Of those positive, 67% of the ATL sera and 78% of the normal sera were further shown to have antibodies against p19. In both groups 17% had antibodies to p15. Generally, the average antibody titers were twice as high in ATL as in normal sera. Competition radioimmunoprecipitation assays done with various sera and involving HTLV-producing cells, virus-positive cells from a Japanese ATL patient, and virus-positive cultured T cells of one of his healthy family members as competing materials demonstrated no differences between the p24, p19, and p15 found in these cells. These results provide strong and detailed immunological evidence that the human retrovirus isolates first made from US patients with cutaneous T-cell malignancies, and those made later in Japanese ATL are either identical or very closely related strains of the same virus, HTLV, a finding verified in other detailed analyses of the HTLV genomes of the respective isolates. To date only HTLV-II, isolated from a US case of hairy-cell leukemia of a T-cell type, is a distinct additional human retrovirus class.

Antibodies against human T-cell leukemia/lymphoma virus (HTLV) and expression of HTLV p19 antigen in relatives of a T-cell leukemia patient originating from Surinam.

Serum and peripheral blood cell samples from eleven relatives of a T-cell leukemia patient with human T-cell leukemia/lymphoma virus (HTLV)-associated disease, were investigated for the presence of HTLV antibody and antigen expression. In addition to the patient, three family members were seropositive and a wide range in HTLV antibody titer was observed. The father of the patient showed the highest titer (173,000) and carried HTLV p19+ cells in his peripheral blood. Upon induction of proliferation of these cells by short-term culture in the presence of phytohemagglutinin (PHA) and 12-O-tetradecanoyl phorbol-13-acetate (TPA), an increase from 1% to 28% HTLV p19+ cells was observed confirming previous findings that HTLV p19 expression was correlated with proliferative activity of the host cells. In none of the other seronegative or seropositive relatives of the patient HTLV p19 expression was revealed when tested in freshly isolated mononuclear cells, upon short-term incubation in PHA + TPA or after prolonged culture for 2 or 3 passages in medium containing T-cell growth factor. The results from HLA typing studies did not provide any evidence for HTLV related abnormalities in haplotype expression. Our data confirmed the earlier notion of a prevalence of HTLV infection within families of patients with HTLV-associated disease. It is furthermore obvious from our results that a normal individual may possess high titer HTLV antibody and circulating HTLV p19+ cells without showing signs of disease.

Transformation of human umbilical cord blood T cells by human T-cell leukemia/lymphoma virus.

Several isolates of human T-cell leukemia/lymphoma virus (HTLV) were transmitted to normal human T cells obtained from the umbilical cord blood of newborns. T cells from seven specimens were immortalized by infection with different HTLV isolates and their properties were compared with those of activated uninfected normal T cells grown in the presence of T-cell growth factor (TCGF) and with those of HTLV-positive neoplastic T-cell lines derived from patients with T-cell malignancies. The HTLV-infected cells generally belonged to a class of mature T cells (OKT4+ and Leu 3A+) and differed from the normal uninfected cells in that they could be propagated in culture indefinitely; possessed altered morphology, including convoluted nuclei and some bi- and multinucleated giant cells; formed large clumps in culture; demonstrated a diminished requirement for TCGF; had an increased density of TCGF receptors; often became completely independent of exogenous TCGF; and expressed HLA-DR determinants. These properties of the HTLV-infected cord blood T cells contrasted to those of uncultured cord blood T cells and of cord blood cells stimulated with mitogen and grown with TCGF but resembled the characteristics of T-cell lines established previously from patients with HTLV-associated T-cell malignancies. This in vitro system offers a unique opportunity to study the basic mechanism involved in abnormal growth and neoplastic transformation of a specific class of human T cells.

Cell lines producing human T-cell lymphoma virus show altered HLA expression

Human T-cell leukaemia/lymphoma virus (HTLV) can be identified in fresh and cultured T-lymphocytes from patients with adult T-cell malignancies. HLA typing of the peripheral blood lymphocytes and cultured cell lines from the patient from which the virus was originally isolated suggested the expression of additional HLA-A and -B locus antigens on the HTLV positive cultured T-cells that were not present on the EBV transformed B-cell line or on the peripheral blood lymphocytes. Peripheral blood lymphocytes (PBLs) and T-cell lines established from patients and cord blood lymphocytes, infected with virus by co-culture with T-cell lines, were typed for HLA antigens with alloantisera and in addition tested for reactivity with a monoclonal antibody (4D12) which recognizes a polymorphic HLA class-I antigen. In all HTLV positive cells, with demonstrable provirus replication, altered HLA alloantigen expression was observed. This may be explained by the observations reported in the accompanying paper which shows homology between the envelope gene region of HTLV and the region of an HLA-B locus gene which codes for the extracellular portion of a class I histocompatibility antigen.

The Human T-Cell Leukemia-Lymphoma Virus in the Southeastern United States

The human T-cell leukemia-lymphoma virus (HTLV) is a recently described RNA tumor virus associated with human T-cell malignant neoplasms. In two geographic areas, Japan and the Caribbean basin, clusters of adult T-cell leukemia-lymphoma are "sentinel diseases" and have suggested an underlying prevalence of HTLV infection in both family members of the index cases and in the population. Four cases of lymphoma from the United States are described as illustrative of the sentinel disease. Serological studies of families and of a small population sample suggest that HTLV infection is endemic in certain parts of the southeastern United States at rates similar to those seen in Caribbean blacks but at a lower rate than that observed in southwestern Japan.

Human T-Cell Leukemia-Lymphoma Virus and Adult T-Cell Leukemia

THE ISOLATION and characterization of the human T-cell leukemia-lymphoma virus (HTLV)^1-5and its relationship to adult T-cell leukemia (ATL)^6-8represent the remarkable convergence of scientific discoveries made half a world apart. The discovery of HTLV represents the culmination of a search for a true human retrovirus (type C virus) that dates back more than 50 years. Almost simultaneous to this breakthrough was the remarkable recognition of a distinctive clinicopathologic entity, ATL.^9-11The molecular and clinical studies summarized in this review represent a scientific tale of two cities, Bethesda, Md, and Kyoto, Japan. <h3>HTLV: A Prototype Human Retrovirus</h3> In numerous animal species, naturally occurring leukemias or lymphomas are caused by retroviruses (also known as type C viruses, RNA tumor viruses, leukemia viruses).^12-14This class of virus is characterized by a replication cycle requiring integration of the viral genetic material in the form of a provirus into host

Human T-cell leukemia-lymphoma virus and adult T-cell leukemia

Human T-cell leukemia-lymphoma virus and adult T-cell leukemia

The human T-cell leukemia-lymphoma virus in the southeastern United States

The human T-cell leukemia-lymphoma virus in the southeastern United States

The human T-cell leukemia/lymphoma virus associated with American adult T-cell leukemia/lymphoma

The human T-cell leukemia/lymphoma virus (HTLV) is a novel type-C retrovirus isolated from patients with T-lymphoproliferative malignancies. Thirteen cases of HTLV-associated malignancy from US centers were studied in detail. Ten of these cases share common clinical features and define a typical virus-associated adult T-cell leukemia/lymphoma (ATL). All ten patients presented with Ann Arbor stage IV lymphoma because of skin involvement, bone marrow involvement, or lymphomatous leptomeningitis. Lymphadenopathy occurred in 7 of 10 patients at presentation, and the malignant cells were cytologically pleomorphic. Leukemia occurred in 60% of the patients at presentation. Hypercalcemia was found initially in two-thirds of the patients, with lytic bone lesions or positive bone scans in 7 of 10. Complete remission occurred in 40%, but all have relapsed. These cases closely resemble those virus-positive cases of adult T-cell leukemia/lymphoma (ATL) reported from Japan and the Caribbean. Three additional virus- positive patients had atypical presentations and diagnoses (acute lymphocytic leukemia, Sezary's syndrome, leukemic reticuloendotheliosis), usually with less aggressive clinical courses and atypical demographic and laboratory features. Presence of HTLV serum antibodies in cases of ATL (with hypercalcemia and circulating malignant cells) appears to define a distinct clinicopathologic entity that may occur in geographic clusters.

Restricted expression of human T-Cell leukemia-lymphoma virus (HTLV) in Transformed human umbilical cord blood lymphocytes

The productive infection and transformation of fresh human lymphocytes by several HTLV isolates have recently been reported. We extend these observations here with the description of multiple immortalized, non-producer, human umbilical cord blood lymphocyte cultures developed by cocultivation or fusion of fresh cells with T cells cultured from leukemia-lymphoma patients. These transformed neonatal leukocytes exhibit morphological, cytochemical, and other phenotypic characteristics similar to those of other HTLV-infected cells but, in contrast to the usual productive infection seen, these cells contain only low amounts of viral proteins and do not release virus particles. These cells contain at least one copy of HTLV proviral DNA/cell and transcribe viral RNA similar in size to virus-producing cells. Virus expression in these cultures was not enhanced by IUdR treatment. These cell cultures should be useful in studies of the regulation of viral expression in human cells and of the viral proteins and nucleic acids involved in T-cell immortalization and growth.