Human T-cell Lymphotrophic Virus-1 Research Articles

Renal Prognosis of COVID-19 Associated Nephropathy

COVID-19 associated nephropathy (COVAN) is the most common cause of acute kidney injury (AKI) in Black patients with SARS-CoV-2 infection undergoing kidney biopsy. It presents with AKI and proteinuria, often nephrotic-range. The histopathology of COVAN is collapsing glomerulopathy (CG), the most severe form of focal segmental glomerulosclerosis.1 The pattern of injury of CG is also seen in the setting of other infections (including human immunodeficiency virus, human T-cell lymphotrophic virus-1, filariasis, leishmaniasis, parvovirus B19, cytomegalovirus, and loa loa), autoimmune disease, ischemia, anabolic steroids, and therapeutic or recreational drugs.

Structural insight into the Scribble PDZ domains interaction with the oncogenic Human T-cell lymphotrophic virus-1 (HTLV-1) Tax1 PBM.

Scribble (Scrib) is a highly conserved cell polarity regulator that harbours potent tumour suppressor activity and plays an important role in cell migration. Dysregulation of polarity is associated with poor prognosis during viral infections. Human T-cell lymphotrophic virus-1 (HTLV-1) encodes for the oncogenic Tax1 protein, a modulator of the transcription of viral and human proteins that can cause cell cycle dysregulation as well as a loss of genomic integrity. Previous studies established that Scribble interacts with Tax1 via its C-terminal PDZ-binding motif (PBM), leading to aggregation of polarity regulators and subsequent perturbation of host cell adhesion, proliferation, and signalling. Using isothermal titration calorimetry, we now show that all four PDZ domains of Scribble bind to Tax1 PBM. We then determined crystal structures of Scribble PDZ1, PDZ2 and PDZ3 domains bound to Tax1 PBM. Our findings establish a structural basis for Tax1-mediated subversion of Scribble-mediated cell polarity signalling and provide the platform for mechanistic studies to examine Tax1 induced mislocalization of Scribble and the associated changes in cellular architecture and subsequent tumorigenesis.

Danger of Biologic Therapy with Unrecognized Human T-cell Lymphotrophic Virus 1 (HTLV-1)

Danger of Biologic Therapy with Unrecognized Human T-cell Lymphotrophic Virus 1 (HTLV-1)

Development of an Anti-HTLV-1 Vaccine for the Treatment of Adult T-Cell Leukemia/Lymphoma

Human T-cell Lymphotrophic Virus 1 (HTLV-1) is the etiologic agent of Adult-T cell Leukemia/Lymphoma (ATL). Therapeutic options for ATL patients are very limited and in aggressive forms of the disease survival rate is only 10% to 30% with conventional chemotherapies and bone marrow transplantation. Although some clinical trials gave encouraging results regarding the efficacy of new treatments, most of them are lifelong, aggressive and failed to achieve a significant impact on long-term survival. Consequently, new treatments for ATL patients are needed to limit relapses and side effects.Specific HTLV-1 cellular immune response is dramatically impaired in ATL patients, which could favor the initiation and the progression of the disease. Hence, stimulating immune responses against HTLV-1 can be an appropriate therapeutic option to treat ATL. THERAVECTYS has developed an anti-HTLV-1 vaccine, based on its lentiviral vector technology inducing a broad, intense and long-lasting cellular immune response after intra-muscular injection. THERAVECTYS was the first company to have launched a clinical trial based on lentiviral vectors technology with the THV01 vaccine for the treatment of HIV (NCT02054286). Results obtained demonstrated both safety and immunogenicity of THV01 in human, with polyfunctional and multi-specific CD4 and CD8 T-cells responses.The anti-HTLV-1 lentiviral vector, THV02 vaccine, encodes for a unique polypeptide derived from Tax, HBZ, p12I and p30II proteins, involved in HTLV-1 pathogenicity and known to be recognized by the immune system of HTLV-1 infected patients. Our preclinical results have demonstrated that THV02 can induce a cellular immune response in C57Bl/6j and BalbC mice and in Sprague Dawley rats, as demonstrated by IFN-γ Elispot. Safety of the THV02 vaccine has been demonstrated during carcinogenicity and regulatory GLP preclinical toxicity studies. Biodistribution and shedding studies demonstrated the very limited diffusion of THV02 after injection, its fast clearance and a non-dissemination in body fluids.As no relevant ATL immunocompetent animal model is available to assess the anti-tumor effect of THV02, THERAVECYTS is developing an ex-vivo efficacy model using blood samples of ATL patients. Briefly, monocyte-derived dendritic cells (MDDC) from blood of ATL patients are purified by isolation of CD14 positive cells from PBMC and differentiation in the presence of IL4 and GM-CSF. MDDC are then transduced with lentiviral vectors encoding for the anti-HTLV-1 antigen and maturation is induced upon TNFa and PGE2 exposure before the co-culture with autologous CD8+ T-cells for stimulation of the cellular immune response. Then, stimulated CD8+ are co-cultured with autologous CD4+ CD25+ ATL cells and the cytotoxic activity is monitored by flow cytometry. Preliminary results demonstrated that MDDC from a chronic ATL patient can be efficiently transduced and matured as attested by the CD40, CD86, HLA-DR, -A, -B and C markers on their surface. In addition, we have observed a specific stimulation of the CD8+, ie an increase of IFNg, TNFa, IL2 and perforin in the media of the co-culture of CD8+ with MDDC expressing anti-HTLV-1 antigen. These data are very encouraging and demonstrate for the first time the feasibility to develop an ex vivo model to assess vaccine efficacy using ATL blood sample. The development of this model is ongoing using several ATL donors representing the different subtypes of the disease and will be presented at the meeting.Regarding the indication and the safety profile of THV02, THERAVECTYS plans to begin a clinical trial in Q4 2015. This assay will be an open-label, dose escalation phase I/II study to assess the safety and the immunogenicity (cellular immune response) of the THV02 vaccination as a treatment of ATL patients. All ATL subtypes will be considered since THV02 vaccine can be combined with conventional ATL treatments. In addition, as the THV02 antigen contains peptides derived from Tax but also HBZ, p12I and p30II viral proteins, all ATL patients can be treated whatever the status of Tax expression. As secondary objectives, both humoral immune response and clinical effect will be assessed. HTLV-1 RNA expression and clonality of HTLV-1 infected cells will be studied as exploratory objectives. Finally, up to 16 patients will be enrolled in France, UK, French Guiana, Martinique and Guadeloupe before doing a phase of extension cohort in US. DisclosuresNo relevant conflicts of interest to declare.

Pilot study of Raltegravir, an Integrase inhibitor, in Human T-cell lymphotrophic virus-1 (HTLV-1) associated myelopathy, tropical spastic paraparesis (HAM/TSP). (P6.300)

Pilot study of Raltegravir, an Integrase inhibitor, in Human T-cell lymphotrophic virus-1 (HTLV-1) associated myelopathy, tropical spastic paraparesis (HAM/TSP). (P6.300)

HTLV 1 associated adult T cell lymphoma/leukemia a clinicopathologic, immunophenotypic tale of three cases from non-endemic region of south India

Adult T cell lymphoma/leukemia is a peripheral T-cell neoplasm caused by human T-cell lymphotrophic virus-1, affects mostly adults with systemic involvement and poor prognosis. Diagnosis of adult T-Cell leukemia/Lymphoma is challenging. The clinico-pathologic and immuno-phenotypic features of the three cases will be presented.

CP-690,550, a therapeutic agent, inhibits cytokine-mediated Jak3 activation and proliferation of T cells from patients with ATL and HAM/TSP

AbstractThe retrovirus, human T-cell–lymphotrophic virus-1 (HTLV-I) is the etiologic agent of adult T-cell leukemia (ATL) and the neurological disorder HTLV-I–associated myelopathy/tropical spastic paraparesis (HAM/TSP). The HTLV-I–encoded protein tax constitutively activates interleukin-2 (IL-2), IL-9, and IL-15 autocrine/paracrine systems that in turn activate the Jak3 (Janus kinase 3)/STAT5 (signal transducers and activators of transcription 5) pathway, suggesting a therapeutic strategy that involves targeting Jak3. We evaluated the action of the Jak3 inhibitor CP-690,550 on cytokine dependent ex vivo proliferation that is characteristic of peripheral blood mononuclear cells (PBMCs) from select patients with smoldering or chronic subtypes of ATL, or from those with HAM/TSP whose PBMCs are associated with autocrine/paracrine pathways that involve the production of IL-2, IL-9, IL-15, and their receptors. CP-690,550 at 50nM inhibited the 6-day ex vivo spontaneous proliferation of PBMCs from ATL and HAM/TSP patients by 67.1% and 86.4%, respectively. Furthermore, CP-690,550 inhibited STAT5 phosphorylation in isolated ATL T cells ex vivo. Finally, in an in vivo test of biological activity, CP-690,550 treatment of mice with a CD8 T-cell IL-15–transgenic leukemia that manifests an autocrine IL-15/IL-15Rα pathway prolonged the survival duration of these tumor-bearing mice. These studies support further evaluation of the Jak3 inhibitor CP-690,550 in the treatment of select patients with HTLV-I–associated ATL and HAM/TSP.

Abstract 2526: CP690550 inhibits interleukin-2 (IL-2), IL-9 and IL-15 mediated JAK3 activation and proliferation of T cells obtained from patients with HTLV-1-associated adult T cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP)

Abstract The retrovirus, human T cell lymphotrophic virus-1 (HTLV-1) is the etiologic agent of ATL and the neurological disorder, HAM/TSP. Infection with HTLV-1 results in viral tax protein mediated constitutive activation of the JAK3/STAT5 pathway in patients' T-lymphocytes that is believed to result in malignant transformation or autoimmunity. JAK3 plays a key role in cytokine receptor-mediated signal transduction including the receptors for IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21 that use the common γ- chain. This suggests that inhibitors of JAK3 may be of benefit in HTLV-1-associated diseases. We evaluated the effect of a novel small molecule JAK3 inhibitor, CP690550 on the cytokine-dependent cell line, NK92. CP690550 at 50 or 100 nM effectively inhibited the proliferation of IL-2-stimulated NK92 cells, but not NK92 that were stimulated with IL-12 or the combination of IL-6 and its receptor (IL-6R) that do not utilize the common γ-chain and JAK3 for signaling and proliferation. We also demonstrated the relative specificity of this inhibitor on another cytokine-dependent cell line, 32Dβ where at 50 or 100 nM CP690550 inhibited IL-2 and JAK3 mediated proliferation but did not affect murine IL-3-mediated cell proliferation. The activation mediated phosphorylation of STAT5 in NK92 cells in response to IL-2 stimulation was inhibited by CP690550. We evaluated the action of CP690550 on the ex vivo proliferation of peripheral blood mononuclear cells (PBMC) from selected patients with the smoldering or chronic subtypes of ATL, or HAM/TSP whose PBMC are associated with the production of cytokines, in particular, IL-2, IL-9 and IL-15 that use the common γ-chain and JAK3. CP690550 at 50 nM inhibited the ex vivo spontaneous proliferation of PBMC from 11 patients with ATL and 9 patients with HAM/TSP at 6-day by means of 66% and 86.4%, respectively. CP690550 also inhibited the proliferation of NK92 cells stimulated with the 6-day ex vivo PBMC culture supernatants from ATL and HAM/TSP patients by a mean of 83.9% and 82%, respectively. We also demonstrated that CP690550 effectively inhibited the activation of the JAK3/STAT5 pathway in isolated T-cells from ATL patients. On the basis of this study, the JAK3 inhibitor CP690550 deserve further study as a potential therapeutic agent for select patients with HTLV-1-associated ATL and HAM/TSP. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2526.

Alemtuzumab (Campath 1-H) in Patients with HTLV-1 Associated Adult T-Cell Leukemia/Lymphoma.

This study investigated the antitumor activity of alemtuzumab (Campath 1-H) with regard to response rate, time to progression, overall survival and adverse events in patients with human T-cell lymphotrophic virus-1 (HTLV-1)-associated adult T-cell leukemia/ lymphoma (ATL). In a single institution non-randomized open-label phase II trial, 24 patients with chronic (n=2), acute (n=13) and lymphomatous (n=9) ATL were treated with intravenous alemtuzumab 30 mg three times weekly for a maximum of 12 weeks. Patients received antimicrobial prophylaxis from the initiation of treatment continuing for a minimum of 2 months after treatment, or until the peripheral blood CD4+ T-cells were ≥ 200 cells/μL. Tumor responses were assessed at the end of 4, 8, and 12 weeks. All patients were evaluable for toxicity and one patient was not evaluable for response due to early withdrawal. The overall objective response rate was 10 out of 23 patients (95% CI: 23 to 66%). Based on the 10 patients who responded, median time to response was 1.1 months (range, 1.0 to 3.0 months). The median response duration was 3.9 months. Median progression free survival of patients with chronic subtype was 2.0 months, 1.0 month for lymphomatous subtype and 2.4 months for acute subtype. Median time to treatment failure was 1.7 months. At data cut-off, (April 24, 2008), 6 (25%) patients were alive. Overall median survival was 5.9 months. Median overall survival by subtypes was 3.9 months for chronic, 4.8 months for lymphomatous, and 7.6 months for acute. Median survival for all responders from on study date was 8.4 months. Clinical benefit was observed both in responders and in patients with stable disease. The most common adverse events were related to infusion and were generally grade 1 or 2, occurring mainly in the first week of alemtuzumab administration. All patients developed CMV antigenemia; however, only 3 (12.5%) were symptomatic. Grade 3 or 4 infections were reported in 6 (25%) patients. Median time to recovery of ALC to >200 cells/μL was 1.9 months.

Viral myelitis: An update

Viral infections of the central nervous system are uncommon but are important in the differential diagnosis of acute myelopathy. Acute viral myelitis can present as acute flaccid paralysis (poliomyelitis) or neurologic dysfunction due to involvement of the white matter. The latter usually affects only part of the transverse expanse of the spinal cord and manifests as asymmetric motor and sensory symptoms. When both halves of the spinal cord are affected, the entity is referred to as acute transverse myelitis and patients exhibit uniformly symmetric weakness, sensory loss, and urinary bladder involvement. Acute flaccid paralysis is due to cytolytic infection of anterior horn cells. When the involvement is mainly white matter, virus-specific and autoimmune host cellular immune responses are believed to contribute to spinal cord damage. Acute flaccid paralysis is caused by polioviruses-1, -2, and -3; coxsackieviruses A and B; enterovirus-71; and flaviviruses, including West Nile virus. Involvement of spinal cord white matter may be associated with infection by many different viruses; however, in most cases a specific viral cause is never determined. Chronic myelitis may be due to either direct infection of the spinal cord by human T-cell lymphotrophic virus-1 (HTLV-1), or a metabolic disturbance due to HIV-1 infection in AIDS patients; no other human virus is known to chronically infect the spinal cord without involvement of the brain. The principal treatment is antiviral drugs immediately upon virus isolation or the identification of a viral sequence by PCR and, when indicated, high doses of methylprednisolone.

A phase II study of the efficacy and toxicity of alemtuzumab for the therapy of human T cell lymphotrophic virus-1 (HTLV-1)-associated adult T-cell leukemia/lymphoma (ATL)

2535 Background: HTLV-1-associated ATL is an aggressive lymphoproliferative disorder with limited effective therapy. Alemtuzumab is a humanized monoclonal antibody directed against CD52. High levels of CD52 are expressed on the surface of ATL cells. Preclinical studies in the MET-1 mouse model of ATL showed alemtuzumab to be highly active. Methods: A single institution open-label phase II study examining the response and toxicity of alemtuzumab in patients with ATL. Eligibility criteria: HTLV-1-associated ATL (chronic, acute and lymphoma type), ≥10% CD52+ malignant cells, measurable disease, age ≥18 years, AGC ≥1000/μL, platelets ≥50,000/μL, adequate physiologic status and informed consent. Results: Ten (10) patients, 6 with leukemia, 4 with lymphoma, 8 Afro-Caribbean, 1 African-American and 1 Japanese, 7 women, 3 men, median age 49 years (range, 36–62), all having received previous treatments including CHOP (7), other monoclonal antibodies (5), radioimmunotherapy (2), and radiation (1), median WBC 48,500/μL (leukemia Pts.) received alemtuzumab standard induction followed by 30 mg 3 times weekly for a minimum of 4 weeks and maximum of 12 weeks. All patients experienced infusion reactions (grade, 1–3) including hypotension, fever, rigors, chills, pruritus and urticaria; however, no patient required discontinuation of treatment. All patients developed grade 4 lymphopenia, and transient cytomegalovirus (CMV) antigenemia. Four (4) patients with leukemia responded (1 CR, 3 PR). No patient with lymphoma responded. Conclusions: Alemtuzumab has antitumor activity in HTLV-1-associated adult T cell leukemia and is well tolerated. Lymphomatous ATL appears unresponsive to alemtuzumab. The reason for this is unknown; however, antibody levels achieved in lymph nodes may be suboptimal. Accrual continues. No significant financial relationships to disclose.

Development of a quantitative cell-based ELISA, for a humanized anti-IL-2/IL-15 receptor β antibody (HuMikβ 1), and correlation with functional activity using an antigen-transfected murine cell line

The HuMikβ 1, a humanized IgG1 monoclonal antibody directed toward the IL-2/IL-15 receptor β-chain (CD122), inhibits the actions of the inflammatory cytokine IL-15, and may be useful for immunotherapy of an array of autoimmune disorders as well as diseases associated with the retrovirus human T-cell lymphotrophic virus 1 (HTLV-1). In order to facilitate the production of material for clinical investigation, we developed a cell-based ELISA (CbELISA) for measuring the binding activity, as a potential biological activity marker, of the HuMikβ 1 monoclonal antibody to a transfected 32D mouse cell line (32Dβ) expressing IL-2Rβ antigen on the cell surface. There is specific binding of HuMikβ 1 to the transfected cell line, titrating out in the concentration range of 1–1000 ng/ml. Under identical conditions, there was no binding of HuMikβ 1 to the parent cell line 32D. Satisfactory binding curves with HuMikβ 1 were obtained with 32Dβ cells grown between 3 and 19 passages in culture and at seed densities of 2 × 10 5–4 × 10 6 cells/well. The binding was specific for Mikβ antibodies recognizing the IL-2/IL-15 receptor β subunit as demonstrated by binding of HuMikβ 1, Mikβ 2 and Mikβ 3 antibodies, and lack of binding of irrelevant humanized and chimeric antibodies and isotype-matched human IgG1 to the 32Dβ cell. Also, the human IgG1 and irrelevant humanized and chimeric antibodies did not interfere with the HuMikβ 1 binding. The assay could detect changes in binding activity of HuMikβ 1 antibody under stressful conditions (heat and low pH) and the results paralleled the effect of stress on the physicochemical characteristics. More importantly, the binding activity shows an apparent correlation to inhibition of IL-15-induced proliferation of 32Dβ cells with HuMikβ 1. In conclusion, the cell-based ELISA method represents a simple, reproducible accurate quantitative assay for monitoring HuMikβ 1 activity and could be used as a potency marker assay for monitoring the lot–lot consistency and functional stability of HuMikβ 1 product.

Complete Cytogenetic Response in a Case of T-PLL with a Complex Karyotype after Treatment with Alemtuzumab.

T-prolymphocytic leukemia (T-PLL) is an aggressive T-cell leukemia characterized by the proliferation of prolymphocytes with a mature postthymic T-cell phenotype and commonly involves the blood, bone marrow, lymph nodes, liver, spleen, and skin. Leukemic cells appear as small to medium-sized prolymphocytes or small variant cells, occasionally with cerebriform nuclei. T prolymphocytes are CD2+, CD3+, and CD7+ and may express CD4+CD8−, CD4+CD8+, or CD4−CD8+ markers. The most frequent chromosomal abnormalities are t(14;14)(q11;q32), inv(14)(q11q32), t(X;14)(q28;q11), i(8)(q10), and t(8;8)(p12;q11). Although patients treated with conventional chemotherapy generally have a poor prognosis, alemtuzumab (Campath®) has been associated with good clinical responses in patients with T-PLL. Routine cytogenetic analysis to determine the response to alemtuzumab has not been performed in patients with T-PLL. This study reports the complete cytogenetic remission achieved following alemtuzumab treatment in a single patient with T-PLL. The 41-year old male patient presented with weight loss, night sweats, diffuse lymphadenopathy and hepatosplenomegaly, in the absence of any skin rash. Peripheral blood counts were: platelets 128 x 109/L and leukocytes 93.6 x 109/L, and hemoglobin values were 12.5 g/dL. The lymphocytes were mostly medium-sized with round to irregular nuclei; some cerebriform cells were also observed, with intermediate chromatin, single evident nucleoli, and a basophilic cytoplasm. Human T-cell lymphotrophic virus-1 serology was negative. Immunophenotypic studies showed CD2+, CD3+, CD5+, CD7+, CD4−, CD8+, CD1−, terminal deoxynucleotidyl transferase-negative (TdT-), and T-cell receptor a/b+. Classic cytogenetics and spectral karyotyping (SKY) were performed and an abnormal karyotype was observed in 18 metaphases analyzed: 46, t(X;14)(q28;q11), t(Y;14)(q12;q11), r[i(8)(q10)]. These abnormalities have not been previously reported in cases of T-PLL. The patient then underwent chemotherapy with CHOP, and subsequent therapy with fludarabine. The patient was then treated with alemtuzumab, 30 mg twice a week, for a total of 7 doses. The patient achieved complete clinical, hematological, and cytogenetic remission, characterized by lack of symptoms, reduction of lymph nodes, liver, and spleen size to normal volumes, and the disappearance of prolymphocytes from peripheral blood. Cytogenetic studies performed 7 months after treatment by classic cytogenetics and SKY revealed a karyotype of 46,XY[20]. This study is the first to demonstrate a complete cytogenetic remission following treatment with alemtuzumab in a patient with T-PLL that was refractory to standard chemotherapy. This is especially impressive in a patient with a complex karyotype and with different cytogenetic alterations not previously described.

Combination therapy for adult T-cell leukemia–xenografted mice: flavopiridol and anti-CD25 monoclonal antibody

AbstractAdult T-cell leukemia (ATL) develops in a small proportion of individuals infected with human T-cell lymphotrophic virus-1. The leukemia consists of an overabundance of activated T cells, which express CD25 on their cell surfaces. Presently, there is no accepted curative therapy for ATL. Flavopiridol, an inhibitor of cyclin-dependent kinases, has potent antiproliferative effects and antitumor activity. We investigated the therapeutic efficacy of flavopiridol alone and in combination with humanized anti-Tac antibody (HAT), which recognizes CD25, in a murine model of human ATL. The ATL model was established by intraperitoneal injection of MET-1 leukemic cells into nonobese diabetic/severe combined immunodeficient mice. Either flavopiridol, given 2.5 mg/kg body weight daily for 5 days, or HAT, given 100 μg weekly for 4 weeks, inhibited tumor growth as monitored by serum levels of human β-2-microglobulin (β2μ; P < .01), and prolonged survival of the leukemia-bearing mice (P < .05) as compared with the control group. Combination of the 2 agents dramatically enhanced the antitumor effect, as shown by both β2μ levels and survival of the mice, when compared with those in the flavopiridol or HAT alone group (P < .01). The significantly improved therapeutic efficacy by combining flavopiridol with HAT provides support for a clinical trial in the treatment of ATL.

From Microarray to Bedside: Targeting NF-κB for Therapy of Lymphomas

From Microarray to Bedside: Targeting NF-κB for Therapy of Lymphomas

Borrelia valaisianain Cerebrospinal Fluid

<i>Borrelia valaisiana</i>in Cerebrospinal Fluid

Cytotoxic T-lymphocyte antigen-4 gene polymorphisms and human T-cell lymphotrophic virus-1 infection: their associations with Hashimoto's thyroiditis in Japanese patients.

Cytotoxic T-lymphocyte antigen-4 (CTLA-4) decreases the immune response of T cells by inactivating the signal that occurs with interaction between CD28 on T cells and B7 on antigen-presenting cells. Gene polymorphisms involving CTLA-4 promoter (-318 C/T), exon 1 (49 A/G), and exon 4 (microsatellite (AT)n) have been linked to Hashimoto's thyroiditis (HT) and other autoimmune diseases. HT also has a reported association with human T-cell lymphotrophic virus-1 (HTLV-1) infection. We investigated the occurrence of CTLA-4 polymorphisms in Japanese patients with HT with and without anti-HTLV-1 antibodies (HTLV-1 Ab). DNA samples from 143 patients with HT and 199 controls were subjected to polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis using the restriction enzymes, Bbv 1, Tse 1, and Mse 1. In the HTLV-1 Ab-positive group the exon 1 G allele was more frequent in patients with HT than in controls (67% vs. 53%, p = 0.0377), and in HTLV-1 Ab-negative group it was also frequent in patients with HT than in controls (68% vs. 53%, p = 0.0041). Frequency of the G allele in HT with HTLV-1 Ab was comparable to those without HTLV-1 Ab. Frequency of polymorphism in the promoter did not differ between patients with HT and controls, nor between controls with and without HTLV-1 Ab. HTLV-1 infection is not associated with CTLA-4 polymorphisms in either HT or controls. HTLV-1 infection is not regulated by genetic factor such as CTLA-4, and may affect occurrence of HT as an independent purely environmental factor.

DNA Triplex Formation Selectively Inhibits Granulocyte-Macrophage Colony-stimulating Factor Gene Expression in Human T Cells

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hemopoietic growth factor that is expressed in activated T cells, fibroblasts, macrophages, and endothelial cells. Although GM-CSF does not appear to be essential for normal hemopoiesis, overexpression of GM-CSF has been implicated in the pathogenesis of some diseases such as myeloid leukemia and chronic inflammation. An NF-kappaB/Rel binding site within the GM-CSF promoter, termed the kappaB element appears to be important for controlling expression in reporter gene assays in response to a number of stimuli in T cells. We investigated oligonucleotide-directed triple helix formation across this regulatory sequence as a potential tool to inhibit GM-CSF gene transcription. A 15-base oligonucleotide, GM3, was targeted to a purine-rich region in the GM-CSF proximal promoter, which overlaps the kappaB element. Gel mobility shift assays and DNase I footprinting demonstrated that GM3 formed a sequence-specific collinear triplex with its double-stranded DNA target. Triplex formation by GM3 blocked recombinant and nuclear NF-kappaB proteins binding to the GM-CSF element. GM3 also caused selective inhibition of the human T-cell lymphotrophic virus-1 Tax transactivator-induced luciferase activity from a reporter construct driven by the GM-CSF promoter in Jurkat T cells. Finally, GM3 greatly reduced the concentration of endogenous GM-CSF mRNA induced by different stimuli in Jurkat T cells but did not affect interleukin 3 mRNA levels in the same cells. We conclude that the kappaB element in the GM-CSF promoter plays a central role in the transcriptional activation of the endogenous GM-CSF gene. Colinear triplex formation acts as a selective transcriptional repressor of the GM-CSF gene and may have potential therapeutic application in cases of undesirable overexpression of this protein.