Acute Adult T-cell Leukemia Research Articles

Serum Levels of Endogenous Thrombopoietin and Granulocyte-Colony Stimulating Factor in Patients with Acute or Lymphoma Type Adult T-cell Leukemia During Multicycle Chemotherapy

Recent multidrug chemotherapy for adult T-cell leukemia (ATL) showed improved findings, however, these protocols often induced persistent myelosuppression. Among 67 patients with acute and lymphoma type ATL treated between January 1996 and December 1998, 42 patients died during this period and showed chemotherapy-induced myelosuppression. To characterize the relation between the severity of myelosuppression and the endogenous thrombopoietin (TPO) or granulocyte-colony stimulating factor (G-CSF) levels in ATL patients, we measured these hematopoietic factors using ELISA method. Nineteen patients with acute or lymphoma type ATL and 16 healthy individuals were examined. During thrombocytopenia, the serum TPO levels were significantly higher than that of controls (P <0.0001) and were inversely correlated with the platelet counts (r = m 0.687 P <0.001). Later in the chemotherapy cycle, severe persistent thrombocytopenia occurred and TPO levels elevated and remained at a high level approximating the TPO levels of exogenous TPO administration (0.3 µ g/kg body weight). On the other hand, the serum G-CSF levels with absolute neutrophil counts (ANC) below 0.5 × 10 9 /L were significantly higher than controls (P =0.009) and inversely correlated with ANC (r = m 0.382 P =0.0034). However, G-CSF levels in six samples obtained after 6 h of G-CSF (100-150 µ g per body) administration was approximately 50-fold higher than that in the neutropenic states. These findings suggested that G-CSF can effectively reduce the severity and duration of intensified chemotherapy-induced neutropenia and higher dose exogenous TPO (higher than 0.6 µ g/kg per day) therapy may be required to enhance platelet recovery after intensive chemotherapy in ATL patients.

Matrix metalloproteinase-9 and vascular endothelial growth factor: a possible link in adult T-cell leukaemia cell invasion.

Plasma from a total of 57 patients with adult T-cell leukaemia (ATL) (acute ATL, 39 patients; lymphoma ATL, one patient; chronic ATL, 15 patients; smouldering ATL, two patients) and 20 healthy controls was analysed for the presence of type IV gelatinase activity with clinical features. A significant elevation of plasma matrix metalloproteinase-9 (MMP-9) was observed in some ATL patients, particularly in the patients with malignant cell infiltration. MMP-9 was found to be secreted into the conditioned medium from all ATL cell lines examined. Moreover, the corresponding mRNA was detectable both in all ATL cell lines examined and in the majority of primary acute ATL cells, indicating that ATL cells are capable of synthesizing and secreting MMP-9. We previously demonstrated that a high incidence of ATL cell infiltration was closely related to a high plasma level of vascular endothelial growth factor (VEGF) produced by ATL cells themselves. This present study showed that the presence of increased plasma MMP-9 was closely associated with elevated plasma VEGF in ATL patients. Furthermore, we showed that both increased plasma MMP-9 and VEGF were significantly related to high ATL cell infiltration. All these findings strongly suggest that MMP-9 and VEGF act co-operatively in the process of ATL cell invasion.

Arsenic Trioxide Induces Apoptosis in HTLV-I Infected T-cell Lines and Fresh Adult T-cell Leukemia Cells Through CD95 or Tumor Necrosis Factor α Receptor Independent Caspase Activation

Arsenic trioxide (As2O3) has been reported to induce apoptosis in human T-cell leukemia virus type-I (HTLV-I) infected T-cell lines and fresh adult T-cell leukemia (ATL) cells and to induce G1 phase accumulation in HTLV-I infected T-cell lines. The present study aimed to clarify the pathway of As2O3-induced apoptosis in HTLV-I infected T-cell lines, MT-1 and MT-2, and fresh ATL cells separated from peripheral blood of patients with acute or chronic type ATL. Cells were treated up to 72 h at clinically tolerable concentrations of As2O3 (1–2 µmol/l) shown to be safe in patients with acute promyelocytic leukemia (APL). Activation of caspases 3, 8, and 9, loss of mitochondrial transmembrane potential and cleavage of poly (adenosine diphosphate-ribose) polymerase (PARP) were observed during As2O3 treatment. Furthermore, prior exposure to a broad-spectrum caspase inhibitor blocked As2O3-induced apoptosis but not G1 phase accumulation. While pre-treatment with a CD95 receptor-blocking antibody (Ab) or a TNF-α neutralizing Ab did not show such inhibitions in these cells. In conclusion, As2O3 induces apoptosis in HTLV-I infected T-cell lines and fresh ATL cells through CD95 or TNF-α receptor independent caspase activation.

Possible involvement of interferon regulatory factor 4 (IRF4) in a clinical subtype of adult T-cell leukemia.

Interferon regulatory factor (IRF) 4 is the lymphoid‐specific transcription factor that is required for the proliferation of mitogen‐activated T cells. IRF4 has been suggested to be involved in tuniorigenesis because the overexpression of IRF4 caused the transformation of Rat‐1 fibroblasts in vitro. Here, we show that IRF4 is constitutively expressed in adult T‐cell leukemia (ATL)‐derived cell lines, which were infected with human T‐cell leukemia virus type‐I, but hardly expressed the trans‐activator protein, Tax. Similarly, constitutive expression of IRF4 was demonstrated in freshly isolated peripheral blood mononuclear cells (PBMC) from patients with either acute or chronic ATL. However, the high‐level expression of IRF4 was specifically associated with acute ATL. With mitogen‐activated PBMC from healthy donors, cell cycle analyses revealed that the induction of IRF4 occurred prior to cell cycle progression and the cells that had entered the cell cycle were predominantly IRF4‐positive cells. In addition, ectopic expression of IRF4 in Rat‐1 fibroblasts increased the S and G2/M phase population significantly. Taken together, our results indicate that IRF4 is involved in the pathogenesis of ATL through its positive effect on the cell cycle, and that IRF4 can be used as a molecular marker of clinical subtype in ATL.

Autocrine antiapoptotic stimulation of cultured adult T-cell leukemia cells by overexpression of the chemokine I-309

Adult T-cell leukemia (ATL) is an aggressive malignancy of CD4(+) T cells caused by the human T-cell leukemia virus type 1 (HTLV-1). The viral leukemogenesis is critically dependent on its oncoprotein Tax because the protein as well as the virus can immortalize primary human lymphocytes to permanent growth. As a transcriptional transactivator, Tax can stimulate the expression of distinct cellular genes. Alterations in the expression levels of unknown growth-relevant genes may contribute to the changed growth properties of Tax-immortalized and leukemic cells. To identify genes that are linked to Tax transformation and ATL leukemogenesis, this study systematically compared the gene expression of cultured cells from patients with acute ATL with that of stimulated peripheral blood T lymphocytes. Several overexpressed RNAs that encode signal transduction functions were identified. These include a dual-specific protein phosphatase (PAC1), an interferon-inducible factor (ISG15), a basic helix-loop-helix transcription factor (DEC-1), and the secreted antiapoptotic chemokine I-309. The ATL cell culture supernatants contained an antiapoptotic activity that could be specifically inhibited by antibodies directed against I-309. Inhibition of I-309 receptor (CCR8) signaling by pertussis toxin increased the apoptosis rate of ATL cell cultures in the presence and absence of external apoptotic stimuli. Both the I-309--specific antiapoptotic activity and the proapoptotic effect of inhibitors of I-309 signaling suggest the existence of an antiapoptotic autocrine loop in ATL cells. Thus, the overexpression of this chemokine may inhibit apoptosis in ATL cells and could substantially contribute to their growth. (Blood. 2001;98:1150-1159)

Expression of functional lung resistance–related protein predicts poor outcome in adult T-cell leukemia

Chemotherapy of patients with adult T-cell leukemia (ATL) has been unsuccessful. The poor outcome is thought to be caused mainly by the drug resistance of ATL cells. Lung resistance-related protein (LRP) is a novel protein associated with drug resistance. The expression of LRP messenger RNA (mRNA) was evaluated by slot blot analysis in 55 patients with ATL. Of these patients, 36 had acute, 12 chronic, and 7 lymphoma-type ATL. The expression levels of LRP mRNA were significantly higher in chronic ATL than in lymphoma-type ATL (P =.007). The expression of LRP mRNA was higher in patients with white blood cell counts above 30,000/microL (P =.038) or with abnormal lymphocyte counts above 10,000/microL (P =.007) than in the remaining patients. The enhanced efflux of [(14)C]doxorubicin from nuclei isolated from ATL cells that expressed high levels of LRP was inhibited by a polyclonal antibody against LRP, and the accumulation of doxorubicin in the isolated nuclei was increased by the anti-LRP antibody. In acute and lymphoma-type ATL patients, high expression of LRP mRNA at diagnosis correlated with shorter survival, and a Cox proportional hazards model showed that LRP expression is an independent prognostic factor. These findings suggest that functionally active LRP is expressed in some ATL cells and that it is involved in drug resistance and poor prognosis in ATL. (Blood. 2001;98:1160-1165)

Existence of escape mutant in HTLV-I tax during the development of adult T-cell leukemia

AbstractAlthough Tax protein is the main target of cytotoxic T lymphocyte (CTL) on human T-cell lymphotropic virus type I (HTLV-I)–infected cells, and Tax peptide 11 through 19 binding to HLA-A*02 has been shown to elicit a strong CTL response, there are patients with adult T-cell leukemia (ATL) bearing HLA-A*02. To explore whether there is genetic variation in HTLV-I tax that can escape CTL recognition during the development of ATL, the HTLV-I tax gene was sequenced in 55 patients with ATL, 61 patients with HTLV-I–associated myelopathy/tropical spastic paraparesis (HAM/TSP), and 62 healthy carriers, and it was correlated with the presence of HLA-A*02. First, a premature stop codon in the 5′ half of the tax gene that looses transactivation activity on the viral enhancer was observed in 3 patients with acute and 1 patient with chronic ATL. This stop codon was revealed to emerge after the viral transmission to the patient from sequence analysis in family members with ATL. Second, amino acid change in Tax peptide 11-19 was observed in 3 patients with ATL. CTL assays demonstrated that this altered Tax 11-19 peptide, observed in ATL patients with HLA-A*02, was not recognized by Tax 11-19–specific CTL. Two patients with ATL had large deletions in tax by sequencing, and 5 patients with ATL had deletions in HTLV-I by Southern blotting. These findings suggest that at some stage of ATL development, HTLV-I–infected cells that can escape the host immune system are selected and have a chance to accumulate genetic alterations for further malignant transformation, leading to acute ATL.

HTLV-I Tax Related Dysfunction of Cell Cycle Regulators and Oncogenesis of Adult T Cell Leukemia

HTLV-I is causually related to the oncogenesis of adult T cell leukemia (ATL). However, the precise mechanism of HTLV-I oncogenesis is unclear. HTLV-I Tax protein functions as an activator of various cellular genes, including IL-2, IL-2 receptor-α, and c-fos through the activation of nuclear transfer factors such as NF-kB and SRF, and also potently activates trascription of viral genes through CREB/ATF sites in the viral LTR. However, Tax activation of HTLV-I infected T cells through the above pathways induces polyclonal proliferation of the cells in vitro; Tax however may function only transiently in the immediate post-infection period following infection in vivo. The long latent period of 60 years from infection to onset of disease suggests other mechanisms for ATL oncogenesis. Recent studies suggest that the malignant transformation of ATL is a multi-hit phenomena, suggesting that discrete genetic events are responsible for ATL oncogenesis. These genetic events could be responsible for the different stages of ATL: smoldering, chronic, lymphoma, and acute type. p16 and p53 genes are important negative regulators of the cell cycle and are often found to be mutated in neoplasms. Recent studies including ours demonstrated a high frequency of alteration of these two genes in primary ATL cells. Furthermore, alteration of the two genes is associated with acute but not chronic type ATL. In addition, p16 gene alteration is linked to the growth rate of ATL cells, suggesting that the alteration of these cell cycle regulatory genes may be related to progression from smoldering or chronic to acute or lymphoma type ATL. Tax may be involved in mutagenesis of these genes through suppression of DNA-β polymerase gene expression during the process from latent period to acute/lymphoma type. Once transformation occurs, activation of the pathway between Tax and the three nuclear transfer factors, NF-kB, SRF, and CREB/ATF, contributes to establish the aggressive manifestations of acute/lymphoma type ATL cells.

Adult T-cell leukemia cells over-express the multidrug-resistance-protein (MRP) and lung-resistance-protein (LRP) genes.

Adult T-cell leukemia (ATL) is a T-cell malignancy caused by human T-cell-leukemia-virus-I (HTLV-I) infection. ATL comprises 4 clinical forms: acute, chronic, smoldering and lymphoma types. ATL is usually resistant to conventional chemotherapy and has a relatively poor prognosis; however, the resistance mechanisms remain undetermined. To explore the multidrug-resistance (MDR) mechanisms of ATL, we examined the expression and functional activity of MDR-related genes in peripheral-blood mononuclear cells (PBMC) from ATL patients by semi-quantitative RT-PCR and FACScan with calcein-AM. PBMC from ATL patients expressed similar or higher levels of MRP, LRP and cMOAT mRNAs, as compared with normal PBMC. In normal controls and ATL patients, MDR1 mRNA expression was undetectable in this study. PBMC from acute and chronic ATL patients expressed significantly higher levels of MRP and LRP mRNA than did normal PBMC (p < 0.01 and p < 0.05 respectively). In chronic ATL, positive correlations were apparent between levels of MRP and LRP mRNA expression (r = 0.759, p = 0.018), and between each mRNA level and the absolute number of abnormal lymphocytes in peripheral blood. Probenecid, an inhibitor of the MRP pump, significantly increased the accumulation of calcein in PBMC from 3 chronic ATL patients. Our findings suggest that the MRP and LRP genes in ATL are often activated by HTLV-I infection and may confer MDR of ATL cells in vivo. Combined chemotherapy with inhibitors of these MDR genes may be promising in the treatment of ATL.

Alteration of p16 (CDKN2) gene is associated with interleukin-2–induced tumor cell growth in adult T-cell leukemia

Because tumorigenesis frequently involves the dysfunction of cell cycle-related proteins, we examined the effect of mutations in CDK inhibitor p16 and its linked genomic loci p15, cl.B, and 1063.7 on the growth of primary adult T-cell leukemia (ATL) cells. Southern blot analysis of primary ATL cells showed a significantly higher incidence of p16 gene alteration in acute ATL than in chronic ATL [67.7% (23/34) vs 26.1% (6/23), respectively; p < 0.003]. Similarly, polymerase chain reaction (PCR) analysis of p16 exon 2 revealed a higher incidence of alteration in acute ATL than in chronic ATL [52.9% (18/34) vs 26.1% (6/23), respectively; p < 0.05]. PCR-single strand conformation polymorphism analysis of exons 1 and 2 of p16 showed no mutations in the patients, with normal pattern by Southern blotting or PCR analysis. Notably five of six chronic ATL patients with abnormal p16 genes progressed to acute crisis within 4 months. PCR analysis of the p16 linked loci 1063.7, p15 exon 2, and cl.B found homozygous deletion in 55.9%, 20.6%, and 2.9% of acute ATL cells and 39.1%, 13.0%, and 0% of chronic ATL cells, respectively, showing no relationship of homozygous deletion in either loci with disease subtypes. In most cases, deletions were seen in multiple genes, including p16. Acute ATL cells had a higher frequency of multigene deletions than chronic ATL cells [44.1% vs 17.4%; p < 0.05]. When leukemic cells were analyzed for interleukin 2 (IL-2) responsive growth, only p16 gene alteration was directly associated with leukemic cell growth activity. Among leukemic cells showing high IL-2 responsiveness, 73.1% (19/26) had p16 gene alteration vs 27.8% (5/18) of leukemic cells that showed low IL-2 responsiveness ( p < 0.005). p16 gene alteration was found in 73.3% (14/19) of leukemic cells showing high autonomous growth rates but in only 40.0% (10/25) of those leukemic cells showing low autonomous growth ( p < 0.03). These results suggest the following: alteration of p16-related genomic regions in ATL is usually a wide rearrangement including the p16 gene; within this region, only p16 gene alteration is associated with disease aggressiveness; and p16 gene deletion may be a proximate event in leukemogenesis.

A Family of the Super High Frequency of ATL

P140 We report a very rare case of a family with acute adult T-cell leukemia (ATL) in all six siblings. Sibling I; 77 yr. female. She developed lymphadenopathy and skin lesion in November, 1989. Anti-HTLV-I (+). Her white blood cell (WBC) count was 100,000/μl, and 95% of them were ATL cells (CD3+4+8−) with convoluted nuclei. Pathology of the lymph node (LN) revealed non-Hodgkin's lymphoma (NHL), diffuse, pleomorphic type. Sibling II; 72 yr. female. She noticed lymphadenopathy in July, 1988. Anti-HTLV-I (+). Her WBC count was 3,200/μl and 10% were ATL cells with indented nuclei. Pathology of the LN showed NHL, diffuse, pleomorphic type (T cell). Sibling III; 60 yr. female. She had lymphadenopathy in 1982. Test for anti-HTLV-I antibodies was not done. A large number of abnormal lymphocytes with convoluted nuclei appeared in the blood. Pathology of the LN revealed NHL, diffuse, medium type. Sibling IV; 64 yr. female. She had lymphadenopathy in Feburary, 1988, and multiple skin tumor occurred in her clinical course. Anti-HTLV-I (+). Her WBC count was 7,700/μl, and 33% were ATL cells with convoluted nuclei. Pathology of the skin tumor revealed NHL, diffuse, pleomorphic type (CD2+3−4+25+30+). Sibling V; 62 yr. male. He had lymphadenopathy in the abdominal cavity and ascites. Test for anti-HTLV-I antibodies was not done. His WBC count was 68,700/μl, and 84% were abnormal lymphocytes. Pathology of the LN was NHL, diffuse, large cell type (T cell). Sibling VI; 48 yr. male. He was admitted to hospital because of cough and dyspnea in 1978. He had lymphadenopathy. His WBC count was 49,600/μl and 90% were abnormal lymphocytes with convoluted nuclei. Other examinations about ATL were not tested. Although examinations about HTLV-I in III, V and VI were not tested, these three cases were highly suspected of acute ATL due to their clinical manifestations and blood pictures. The super high frequency of ATL in the siblings may show this family members have disposition of ATL.

Successful Treatment of HTLV-1-associated Acute Adult T-Cell Leukemia by Allogeneic Bone Marrow Transplantation

P121 A 42 year-old male with HTLV-1-associated acute adult T-Cell leukemia (ATL), treated with carcinostatic agents, followed by successful allogeneic bone marrow transplantation(BMT), and remaining in complete remission 5 months after BMT is described. He was admitted in November, 1997 by bilateral cervical lymphadenopathy but free from both hepatosplenomegaly and skin lesion. His white blood cell count was 16.5× 109/l with 7% of abnormal lymphocytes. Pathology of a lymph node revealed malignant lymphoma (diffuse, pleomorphic type with T-cell phenotype). Serum anti-HTLV-1 antibody was positive. These findings lead him to the diagnosis of acute type of ATL. He was treated with cyclophosphamide, adriamycin, vincristine, and prednisolone followed by BMT from an HLA identical sibling on June 10, 1998 resulting in a complete remission. The conditioning regimen consisted of L-PAM and total body irradiation. GVHD prophylaxis was performed with cyclosporine A and short term methotrexate administration. His WBC count increased more than 1.0×109/l 14 day after BMT and platelet count elevated to 50.0×109/l on the 27th day. Complications of acute GVHD (grade 2) appeared with watery diarrhea and cytomegalovirus antigenemia, which were resolved with predonisolone and gancyclovir respectively. He still remains in complete remission 5 months after BMT. Although the efficacy of allogeneic BMT (allo-BMT) for ATL was still uncertain, he was treated successfully by allo-BMT without any life threatening complications. Conclusion: allo-BMT can be one of the best choice for the treatment of ATL.

LEUKEMOGENESIS AND PROGRESSION OF ADULT T-CELL LEUKEMIA

O-15 Following infection with human T-cell leukemia virus type I (HTLV-I), a long latent period is required for the development of frank adult T-cell leukemia (ATL), suggesting that accumulated genetic or epigenetic changes of the host genes are involved in leukemogenesis. In HTLV-I+ carriers, CD4+ memory T-cells increase in accordance with HTLV-I provirus load. In contrast, both CD4+ and CD8+ naive T-cells decrease, which might account for the immunodeficiency associated with HTLV-I infection. Phenotypes of increased CD4 memory T-cells are similar to those of ATL cells, suggesting that HTLV-I viral protein(s), especially Tax, promotes the proliferation of infected cells, leading to the onset of ATL. Cyclin-dependent kinase inhibitor, p16 gene, is one of tumor suppressor genes, whose deletions have been shown to be associated with high-grade malignant ATL. We examined the methylation profile of p16 in various HTLV-I-infected cell populations using methylation-specific PCR (MSPCR) following sodium bisulfite treatment. A high frequency of methylation of p16 gene was identified in ATL samples and HTLV-I-transformed cell lines. Sequencing of p16 revealed that the CpG sites of p16 were more frequently methylated in fresh leukemic cells than detected with MSPCR. Fresh leukemic cells isolated from acute and lymphoma-type ATL cases had more methylated sites than those from chronic or smoldering ATL cases. These data suggest that the CpG sites of p16 are methylated at greater levels as the disease progresses, resulting in the suppression of p16 expression. Although Tax has been shown to functionally inactivate p16, the methylation of p16 also appears to play a critical role in regulating p16 in HTLV-I-associated leukemogenesis.

Treatment of HTLV-1 associated adult T-cell leukemia lymphoma with a combination of Zidovudine and alpha interferon. Report of the French experience

O-05 The combination of Zidovudine (AZT) and alpha interferon (INF) has been shown to induce a good response rate in small group of patient with adult T-cell leukemia (ATL) either as a first line or in relapse after chemotherapy (Gill et al, Hermine et al, NEJM, 1995, 332 ). In the present study, 21 patients with ATL (Acute 16, Lymphoma 3, Chronic 1, Smoldering 1) received the combination of AZT (1 gr/day) and INF (9MUI/d) for two months except two patients who received lower doses. All are seropositive for HTLV-1 with a clonal integration of the provirus in the tumoral cells. Median age was 35 years (18-75). Most of patients have bad prognosis factors ECOG>2 (14), LDH>1N (20), Hypercalcemia (8), Age >40 (13). Sixteen patients were previously untreated, and 4 pts received AZT+INF after standard chemotherapy, and one after autologous bone marrow transplantation. For the 20 pts with an evaluable disease, 16 responses were observed (9 CR, 7 PR). The highest response rate was observed among the 13 previously untreated acute ATL pts (5 CR, 5 PR). However, despite this very high response rate, relapses occured with an evaluated EFS of 8 months, and an overall survival of 14 months. For previously untreated acute ATL patients, the median overall survival was of 18 months. Four patients are still alive 5 years after diagnosis. A high frequency of neurological relapses (4), and initial hypercalcemia (4) were observed despite a good response for other criteria. Toxicity was mild in all pts and was mainly hematological. In conclusion, the combination of AZT+INF exhibits a high response rate in ATL pts, especially in the previously untreated acute ATL with a significant prolongation of survival in this subgroup. In order to avoid relapses, prophylaxis intrathecal chemotherapy as well as post remission treatment are warranted. However, the probability of cure of ATL remains low. Thus new drugs are needed. Based on in vitro studies (Bazarbachi et al, Blood, 1999, Jan 1 ) we are now investigating in vivo the effects of the combination of arsenic and INF as a second line therapy after AZT and INF in ATL patients. Preliminary results will be discussed.

Correlation of telomerase activity with development and progression of adult T-cell leukemia

Telomerase is an enzyme that adds hexameric TTAGGG nucleotide repeats to the ends of vertebrate chromosomal DNAs (i.e. telomeres) to compensate for losses that occur with each round of DNA replication. Telomerase activity, demonstrable in most human tumors, enables them to maintain telomere stability. Peripheral blood mononuclear cells were sampled from 57 patients seropositive for human T-lymphotropic virus type I (HTLV-I), including 24 asymptomatic viral carriers, ten smoldering type, five chronic type, and 18 acute type adult T-cell leukemia (ATL). Telomerase activity was determined in samples using a modified telomeric repeat amplification protocol. We semiquantitatively determined telomerase activity by serial dilution of each sample. All of 23 samples from acute and chronic type ATL patients were positive, seven of ten (70%) smoldering type patients and seven of 24 (29.2%) asymptomatic viral carriers were positive. Disease progression from asymptomatic viral carrier to acute type correlated with telomerase activity. Two samples from chronic type ATL patients with relatively high telomerase activity progressed to the acute type within 1 month. Serum lactate dehydrogenase level also correlated with telomerase activity. These results indicate that reactivation of telomerase activity is a key event in development and progression of ATL, and telomerase could be a useful marker for predicting the course of disease. Accordingly, ATL could be a good candidate disease for trials of telomerase inhibitors, as novel anticancer drugs.

The methylthioadenosine phosphorylase gene is frequently co-deleted with the p16INK4a gene in acute type adult T-cell leukemia.

Adult T-cell leukemia (ATL) is a retrovirus-associated leukemia with poor prognosis and often has deletions of the p16INK4a and p15INK4b genes on chromosome 9p21. The gene for methylthioadenosine phosphorylase (MTAP), a purine and methionine metabolic enzyme, resides approximately 100 Kb telomeric to the p16INK4a gene and is frequently co-deleted with the tumor suppressor gene in a variety of cancers. This enzyme deficiency can be exploited for selective chemotherapy with de novo purine synthesis inhibitors and/or methionine depletion. To determine whether ATL can be a candidate for selective chemotherapy based on genetic alterations on chromosome 9p21, we analyzed the MTAP gene in 41 samples from ATL patients (27 acute type and 14 chronic type ATL) and 3 cell lines established from ATL patients. Five samples from the acute type had deletions of the MTAP gene (4 total deletions and 1 partial deletion of exons 6-8). The MTAP gene was always co-deleted with p16INK4a. No deletion of the MTAP gene was detected in samples from the chronic type. Of 3 cell lines, 2 showed partial deletions of exons 5-8 of the MTAP gene, and 1 lost all exons. The p16INK4a gene was deleted in all cell lines. In conclusion, deletions of the MTAP gene were found in 5 of 27 acute type ATL samples. Acute type ATL with MTAP deficiency can be a good candidate for selective chemotherapy by depleting purines and/or methionine.

Adult T Cell Leukaemia

Adult T cell leukaemia (ATL) is an aggressive clonal malignancy of mature activated T cells, caused by the human T cell lymphotropic virus type I (HTLV-I) infection. ATL continues to have a very bad prognosis because of the intrinsic resistance of leukaemic cells to conventional or even high-dose chemotherapy and because of the associated severe immunosuppression. Even though conventional chemotherapy remains the standard treatment in acute and lymphomatous types of the disease, the benefit of this intensive approach is not well established in chronic and smouldering ATL. Combination chemotherapy regimens, in particular those designed for the treatment of aggressive non-Hodgkin's lymphomas or acute lymphoblastic leukaemia, have little effect in the treatment of ATL. Different combination regimens of conventional chemotherapy have been investigated in Japan, and recent results of intensive induction therapy showed a complete remission (CR) rate of about 40% in the aggressive forms of the disease. However, most of the patients relapsed and eventually died. Allogeneic bone marrow transplantation has been used in the treatment of a very small number of patients with ATL. High toxicity and transplant-related mortality were observed in these immunocompromised patients. New cytotoxic agents have also been used in pilot phase II studies in refractory or relapsed ATL patients, but significant results have only been observed with deoxycoformycin. Recently, promising results have been obtained with anti-Tac monoclonal antibodies directed against the alpha-chain of the interleukin-2 receptor (CD25), which is highly expressed on ATL cells but not on normal resting lymphocytes. Promising results were also reported in 2 phase II studies with combination therapy comprising the antiretroviral agent zidovudine and interferon-alpha (IFNalpha). In previously untreated patients with acute ATL, high and rapid (usually within 2 weeks) response rates were reported. In contrast, in lymphomatous ATL, both studies suggested a milder and slower effect of zidovudine and IFNalpha. In this case, this combination may be used as maintenance therapy after a CR or good partial response induced by polychemotherapy. Since cross resistance between chemotherapy and zidovudine and IFNalpha does not seem to occur, a combination with best induction polychemotherapy is warranted.

Quantitative characterization and potential function of membrane Fas/APO-1 (CD95) receptors on leukaemic cells from chronic B and T lymphoid leukaemias.

The expression and function of the Fas-receptor (Fas-R) were examined in chronic lymphocytic leukaemia (CLL), hairy cell leukaemia-variant (HCL-v) and adult T-cell leukaemia (ATL). The expression of Fas-R in freshly isolated leukaemic cells was qualitatively and quantitatively different between each disease; faint in B-CLL, moderate in HCL-v and strong in ATL. Both full-length and alternatively spliced truncated forms of Fas mRNA were detected even in CLL B cells with faint to negative Fas-R, and Fas mRNA was also shown to be capable of increasing in vitro expression, i.e. the message was functional. In contrast, Fas-R expression on ATL cells was heterogenous and usually intense with a mean density approximately 3-fold higher than that of normal T cells. Fas-R was confirmed to have the potential function for anti-Fas monoclonal antibody-mediated cell death in vitro in Fas-R+ ATL cells. The expression level of Fas-R on the cells was higher in chronic than acute ATL (10,360 v 6260 antibody-binding capacity per cell, mFasABC; P<0.05) and was inversely correlated with serum LDH activity, suggesting that the strong Fas-R accounts for the slow progression of chronic ATL and the negative Fas-R protects from Fas-mediated cell death. These results show that Fas-R expression on leukaemic cells is valuable in their characterization and perhaps their function, and may contribute to the progression and immune evasion of malignant clones.

Frequent expression of interleukin-9 mRNA and infrequent involvement of interleukin-9 in proliferation of primary adult T-cell leukemia cells and HTLV-I infected T-cell lines

To examine the possibility that interleukin-9 (IL-9) may be involved in oncogenesis and the proliferation of adult T-cell leukemia (ATL) cells, we examined the expression of IL-9 mRNA and growth response to IL-9 in five human T-lymphotropic virus type-I (HTLV-I) infected T-cell lines and in primary leukemia cells in peripheral blood from eight patients with ATL (four acute ATL and four chronic). Four out of five cell lines expressed IL-9 mRNA not correlated with Tax expression. Primary ATL cells from all patients also expressed IL-9 mRNA not correlated with the clinical forms. Recombinant IL-9 showed growth enhancing activity in only one out of five cell lines and one out of eight patients' primary leukemic cells. These results suggest the infrequent involvement of IL-9 in the proliferation of ATL cells, both primary tumor cells and HTLV-I infected T-cell lines.

Serum dehydroepiandrosterone and DHEA-sulfate in patients with adult T-cell leukemia and human T-lymphotropic virus type I carriers

The serum levels of dehydroepiandrosterone (DHEA) and DHEA-sulfate (DHEA-S) were determined by radioimmunoassay in 38 patients with adult T-cell leukemia (ATL). Levels of serum DHEA and DHEA-S were also measured in 60 human T-lymphotropic virus type I (HTLV-I) carriers, and did not differ from those in 60 healthy control subjects. Serum levels in patients with ATL were lower than those in the age- and sex-matched healthy controls and in HTLV-I carriers with statistical significance. Serum DHEA and DHEA-S in male patients with acute and lymphoma-type ATL were 1.06 +/- 0.77 ng/ml and 245.8 +/- 192.9 ng/ml, respectively. Levels in male patients with chronic and smoldering-type ATL were 1.69 +/- 0.68 ng/ml and 477.6 +/- 251.5 ng/ml, respectively. Serum levels of DHEA and DHEA-S in patients with acute and lymphoma-type ATL were significantly lower than those in patients with chronic and smoldering-type ATL (P < 0.05). These data suggest that a decrease in serum levels of DHEA and DHEA-S may be associated with patients who have some clinical subtypes of ATL. Moreover, androgens may have a therapeutic role in patients with ATL, as administered in patients with hairy-cell leukemia. Because there is at present no curative chemotherapy for ATL, a trial combination of androgens and standard chemotherapy may be a reasonable therapeutic option in such patients.