Adult T-cell leukemia cells expressing OKT 17 antigen in the activated state

The surface phenotype and the functional activities of leukemic cells from seven patients with adult T-cell leukemia (ATL) were studied using monoclonal antibodies OKT 3, 4 and 8, anti-Tac, and OKT 17. The latter defines the heterogeneity of the activated T4+ T cell subset. In all cases, ATL cells with the typical OKT3-T4+ T8- phenotype expressed OKT 17 antigen. In addition, in five out of the seven cases the fresh ATL cells possessed Tac antigen which is expressed on activated T cells in varying degree. After cultivation with PWM, most populations of ATL cells acquired Tac even in the cases expressing little antigen in uncultured preparations. However the PWM activated ATL cells did not lose OKT 17 antigen. Functional assays showed the suppressor activity of ATL cells on normal B cell differentiation in three out of six cases examined. These results suggest that ATL cells most probably arise from a particular subset characterized by OKT 17 antigen within the activated OKT4+ T cell subset.     

Clinical features of OKT4+/OKT8+ adult T-cell leukemia

Adult T-cell leukemia (ATL) has a range of clinical characteristics. Phenotypically the leukemic cells usually express the helper/inducer associated antigen OKT4 with lack of OKT8. We have observed three patients with acute ATL cytologically indistinguishable from OKT4+/OKT8- ATL but whose neoplastic cells had the unusual phenotype, OKT3+, OKT4+, OKT6-, OKT8+ OKT9+/-, OKT11+, Tac+/-, TdT-. All patients had abnormal karyotypes and antibodies against anti-ATL associated antigens as well as proviral DNA of human T-cell leukemia virus in the leukemic cells. The clinical course was complicated by skin eruptions, hypercalcemia, pulmonary infection and disseminated intravascular coagulopathy. All died of complications shortly after diagnosis. The clinical features of these patients were similar to those of OKT4+/OKT8- ATL. However, their acute course suggests that co-expression surface antigens OKT4 and OKT8 may be a sign of aggressive nature of the disease with poor prognosis.     

Regulatory T-cell function of adult T-cell leukemia/lymphoma cells

Adult T-cell leukemia/lymphoma (ATLL) patients are highly immunocompromised, but the underlying mechanism responsible for this state remains obscure. Recent studies demonstrated that FOXP3, which is a master control gene of naturally occurring regulatory T (Treg) cells, is expressed in the tumor cells from a subset of patients with ATLL. Since most ATLL cells express both CD4 and CD25, these tumors might originate from CD4(+)CD25(+)FOXP3(+) Treg cells, based on their phenotypic characteristics. However, whether ATLL cells actually function as Treg cells has not yet been clearly demonstrated. Here, we show that ATLL cells from a subset of patients are not only hypo-responsive to T-cell receptor-mediated activation, but also suppress the proliferation of autologous CD4(+) non-ATLL cells. Furthermore, ATLL cells from this subset of patients secrete only small amounts of IFN-gamma, and suppress IFN-gamma production by autologous CD4(+) non-ATLL cells. These are the first data showing that ATLL cells from a subset of patients function as Treg cells in an autologous setting. The present study provides novel insights into understanding the immunopathogenesis of ATLL, i.e., how HTLV-1-infected cells can survive in the face of host immune responses. It also adds to our understanding of ATLL patients' severely immunocompromised state.     

Poorly expressed CD2 antigen on the leukemic cells of adult T-cell leukemia and chronic lymphocytic leukemia of T-cell lineage

Adult T-cell leukemia (ATL) and T-cell chronic lymphocytic leukemia (T-CLL) are hematologic neoplasms in differentiated stages of peripheral mature T cells. This is suggested by the presence of CD2 (E rosette receptor) and mature and/or pan-T cell membrane surface antigens on their leukemic cells. We recently encountered one patient with ATL and another with T-CLL; their leukemic cells poorly expressed CD2 antigens, but clinical presentation, morphology of the leukemic cells and other marker studies were characteristic of either ATL or CLL. The clinical significance of the poor expression of CD2 remains to be further studied. The two patients reported here died of severe complications 4 and 9 weeks after diagnosis. The poor expression of CD2 in the peripheral T cells in these neoplasms is likely to indicate an aggressive nature of the disease.     

Aberrant expression of ganglioside and asialoglycosphingolipid antigens in adult T-cell leukemia cells

Gangliosides (GM3, GD3, GM2, gangliotetraose-series gangliosides) and their asialo derivatives of several adult T-cell leukemia (ATL) cell lines (ATL-1K, ATL-3I, ATL-5S, and MT-2 cells) and the lymphocytes from a patient with ATL were quantified by highly sensitive enzyme-immunostaining on silica gel thin layer chromatograms using specific antiglycolipid antibodies. GM2 and GD3 gangliosides and asialo GM1 (GA1) newly appeared in all cultured ATL cells and the lymphocytes from patients with ATL but not in normal human T-lymphocyte-rich fraction. Gangliotetraose-series gangliosides, GM1a, GD1a and GD1b, were also found in cultured ATL cells, but were not detected in normal human lymphocytes or the lymphocytes of a patient with ATL. Quantitative immunostaining analysis of GM2, GD3 gangliosides and GA1 in T-cell lines from non ATL leukemia (Molt-3, CEM and Jurkat) revealed GM2 gangliosides in all the T-cells from non ATL tested and GA1 in Jurkat cells, but no GD3 ganglioside was found in the non ATL leukemia cells tested. The above results indicate that ganglioside GD3 may be a T-cell glycosphingolipid antigen associated with ATL, and ganglioside GM2 and GA1 may be useful as surface markers related with ATL, as well as T-cell lymphoma. The contents of GA1, GM3, GD3, GM2 and gangliotetraose-series gangliosides in ATL cells were all different, even though all the cells used have a common antigen reactive with monoclonal OKT-4 antibody, indicating that there are several subsets of human inducer/helper T-cells, which possess different metabolism and expression of gangliosides.     

Immunosuppressive factors from adult T-cell leukemia cells

The mechanism of immunodeficiency in adult T-cell leukemia (ATL) patients was studied in vitro. Peripheral blood lymphocytes from ATL patients and ATL cell lines such as Hut 102, MT 1, and MT 2 were not activated to proliferate by the stimulation with concanavalin A and suppressed normal lymphocyte proliferative responses induced with concanavalin A when cultured together. The sera from ATL patients and the culture supernatants from ATL cells and ATL cell lines also suppressed normal lymphocyte proliferative responses induced with concanavalin A. By Sephacryl S-200 column chromatography, the suppressive factors were fractionated as a single peak with the molecular weights of 50,000 to 70,000. The suppressive factors were unstable to acid treatment but stable to the treatment with base, heat, freezing-thawing, and trypsin. The factors suppressed the production of interleukin 2 by T-cells and the responsiveness of T-cells to interleukin 2, but not the expression of interleukin 2 receptors on T-cells and the production of interleukin 1 by monocytes. These results suggest that the immunosuppressive factors produced by ATL cells have some roles in the induction of immunodeficient states in ATL patients.     

Capsaicin inhibits growth of adult T-cell leukemia cells

Human T-cell leukemia virus type 1 (HTLV-1)-associated adult T-cell leukemia (ATL) is resistant to conventional chemotherapy. We examined the in vitro effects of capsaicin, the principal ingredients of red pepper, on three ATL cell lines. Capsaicin treatment inhibited the growth of ATL cells both in dose- and time-dependent manner. The inhibitory effect was mainly due to the induction of cell cycle arrest and apoptosis. Capsaicin treatment also induced the degradation of Tax and up-regulation of I kappa-B alpha, resulting in the decrease of nuclear factor (NF)-kappa B/p65 DNA binding activity. In addition, the Bcl-2 level was found to be decreased. Based on these findings, capsaicin may be considered for chemoprevention of ATL.     

Chromosome abnormalities of leukemia cells in adult patients with T-cell leukemia

Chromosomes of 30 patients with adult T-cell leukemia were analyzed. Chromosome abnormalities were found in all the patients examined. The modal chromosome number of abnormal cells was hypodiploid in 2 patients, diploid in 14, and hyperdiploid in 9. The remaining 5 patients had bimodal chromosome numbers (diploid and hyperdiploid modes). Although all the patients showed various numerical or structural chromosome abnormalities, they also had common chromosome abnormalities. Aberrations of chromosome 1 were noted in 20 of the 30 patients, aberrations of chromosome 3 were seen in 20, trisomy 6 or 6q- was found in 17, aberrations of chromosome 10 were noted in 16, aberrations of the long arm of chromosome 14 were seen in 9, and trisomy 18 was seen in 7. There was no particular relationship between the difference in clinical symptoms and disparity in chromosome abnormalities.     

Discordant gene and surface expression of the T-cell receptor/CD3 complex in adult T-cell leukemia cells.

Cell surface expression of the T-cell receptor (TCR)/CD3 complex on the cells from 11 acute type adult T-cell leukemia (ATL) and 4 lymphoma type ATL patients was examined by flow cytometry. Cells from 10 of 11 acute ATL patients were TCR alpha beta+ and CD3+, and their mean fluorescence intensities were low (TCR alpha beta, 25.3-84.6; CD3, 22.8-87.8). Cells from two of four lymphoma type ATL did not express this complex, and the other two were CD3+, TCR alpha beta-. In contrast, the mean fluorescence intensity of the TCR/CD3 complex in cells from a patient with T4 chronic lymphocytic leukemia was not low (TCR alpha beta, 129.9; CD3, 117.1). mRNA expressions of the TCR alpha, beta, and CD3 gamma, delta, epsilon, and zeta chains were examined by Northern blots. ATL cells from two acute and two lymphoma types expressed amounts of this complex equal to or greater than those expressed by T4 chronic lymphocytic leukemia. CD3 delta and TCR beta mRNA in ATL and T4 chronic lymphocytic leukemia cells were equally stable to actinomycin D treatment. The synthesis of CD3 zeta protein by ATL cells was detected by Western blotting assay. On the basis of these findings, we discuss the possible involvement of the TCR/CD3 complex in activation of ATL cells.     

Human T-cell leukemia virus type I and adult T-cell leukemia

Human T-cell leukemia virus type I (HTLV-I) causes adult T-cell leukemia (ATL) in about 5% of carriers after a long latent period. After its infection, HTLV-I promotes the clonal proliferation of HTLV-I infected cells in vivo by actions of encoded viral proteins, including Tax. However, leukemic cells frequently lack the expression of Tax by the genetic and epigenetic changes of HTLV-I provirus, suggesting that Tax is not always necessary after transformation. Alternatively, ATL cells without Tax protein could escape from the host immune system since Tax is the major target of cytotoxic lymphocytes. During the latent period, alterations of host genome accumulate, finally leading to onset of ATL.     

Familial adult T-cell leukemia

Two siblings who developed adult T-cell leukemia (ATL) are presented. The patient and 7 of 26 healthy family members examined had the serum antibodies against ATL-associated antigens (ATLA). This family study shows that two main routes of transmission of human T-cell leukemia virus (HTLV) may be involved: one is the route from parents to children and the other is horizontal transmission among spouses, especially from husband to wife; the anti-ATLA-positive family is considered to be a high-risk group for ATL.     

Multidrug resistance protein expression of adult T-cell leukemia/lymphoma

In adult T-cell leukemia/lymphoma (ATL), it is difficult to achieve remission and the reason for the resistance to chemotherapeutic agents may be linked to the presence of multidrug resistance (MDR) proteins. Lung resistance-related protein (LRP), multidrug resistance-associated protein and P-glycoprotein are three MDR proteins which we examined in ATL cells using multiparametric flow cytometry and real-time RT-PCR. LRP was highly expressed and suppressing LRP function increased doxorubicin accumulation in nuclei. This indicates LRP may be contributing to drug resistance in ATL patients, and the suppression of LRP function could be a new strategy for ATL treatment.     

Centrosome amplification in adult T-cell leukemia and human T-cell leukemia virus type 1 Tax-induced human T cells

Centrosomes play pivotal roles in cell polarity, regulation of the cell cycle and chromosomal segregation. Centrosome amplification was recently described as a possible cause of aneuploidy in certain solid tumors and leukemias. ATL is a T-cell malignancy caused by HTLV-1. Although the precise mechanism of cell transformation is unclear, the HTLV-1-encoded protein, Tax, is thought to play a crucial role in leukemogenesis. Here we demonstrate that lymphocytes isolated from patients with ATL show centrosome amplification and that a human T cell line shows centrosome amplification after induction of Tax, which was suppressed by CDK inhibitors. Micronuclei formation was also observed after centrosome amplification in Tax-induced human T cells. These findings suggest that Tax deregulates CDK activity and induces centrosome amplification, which might be associated with cellular transformation by HTLV-1 and chromosomal instability in HTLV-1-infected human T cells.     

Integration of human T-cell leukemia virus type 1 in genes of leukemia cells of patients with adult T-cell leukemia

Adult T-cell leukemia (ATL) occurs after a long latent period of persistent infection by human T-cell leukemia virus type 1 (HTLV-1). However, the mechanism of oncogenesis by HTLV-1 remains to be clarified. It was reported that the incidence curve of ATL versus age was consistent with a multistage carcinogenesis model. Although HTLV-1 is an oncogenic retrovirus, a mechanism of carcinogenesis in ATL by insertional mutagenesis as one step during multistage carcinogenesis has not been considered thus far, because the exact integration sites on the chromosome have not been analyzed. Here we determined the precise HTLV-1 integration sites on the human chromosome, by taking advantage of the recently available human genome database. We isolated 25 integration sites of HTLV-1 from 23 cases of ATL. Interestingly, 13 (52%) of the integration sites were within genes, a rate significantly higher than that expected in the case of random integration (P = 0.043, chi(2) test). These results suggest that preferential integration into genes at the first infection is a characteristic of HTLV-1. However considering that some of the genes are related to the regulation of cell growth, the integration of HTLV-1 into or near growth-related genes might contribute to the clonal selection of HTLV-1-infected cells during multistage carcinogenesis of ATL.     

Correlation of aberrant proliferation with T-cell growth factor in adult T-cell leukemia cells

Peripheral blood lymphocytes from seven patients with adult T-cell leukemia (ATL) were found to lack PHA-responsiveness. However, in most of the cases, minute but distinct proliferation could be induced and maintained by human spleen cell conditioned medium containing PHA or by a combination of PHA and conditioned medium of gibbon cell line, MLA-144 (MLA-144 CM). These results indicate that the lack of response to mitogens of ATL cells might be attributed not only to the failure of these cells to produce T-cell growth factor (TCGF) upon activation, but also to their poor responsiveness to TCGF. Furthermore, a direct proliferative response to mitogen-free MLA-144 CM was shown in two out of seven patients; these two patients experienced rapidly progressive clinical courses. This observation raises the possibility that TCGF promotes the growth of ATL cells in vivo, and is related to the clinical course of the disease.     

Binding of adult T-cell leukemia virus to various hematopoietic cells

A newly found human retrovirus, adult T-cell leukemia virus (ATLV) was shown by means of membrane immunofluorescence to bind to various hematopoietic cells including T-, B- and non-T, non-B-cell lines. Partially purified viral gp46 from culture fluids of ATL virus producer lines also bound efficiently to an ATLV-negative T-cell line, CCRF-CEM cells. When the viruses were pre-incubated with anti-ATLV-positive human sera, ATLV binding to the cells was clearly inhibited but not by pre-incubation with anti-ATLV-negative sera. These data suggest that: (1) ATLV binds not only to T-cells but also to multiple types of cells of hematopoietic origin; (2) anti-ATLV antibody-positive human sera have the blocking antibody for the binding of ATLV to lymphoid cells.     

Production of parathyroid hormone-related protein in adult T-cell leukemia cells

Human parathyroid hormone-related protein (PTHrP) mRNA was detected in peripheral leukemic cells obtained from adult T-cell leukemia (ATL) patients as well as in cultured human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines. In contrast, PTHrP mRNA was not detected in other types of leukemic cells. Using radioimmunoassay, immunoreactive PTHrP was also detected in the spent media of HTLV-I-infected T-cell lines. These results suggest that PTHrP plays an important role in developing the hypercalcemia frequently observed in ATL patients.