FDC Like Cell Line HK With IL-2, IL-4 and OX40 Ligand Supports The Growth Of ATLL Cells

Adult T-cell leukemia/lymphoma (ATLL) is an aggressive neoplasm derived from CD4+ T-cells with HTLV-I infection, and its mechanisms of tumorigenesis still remain to be elucidated. The fact that tumor cells rarely proliferate in vitro is one of the most important problems to be solved. The establishment of cell line from ATLL patient samples has been difficult even in the presence of interleukins. Previously we established one cell line (HU-ATTAK) from acute or lymphoma types of 10 ATLL cases which did not proliferate in the presence of IL-2 and/or IL-4. HU-ATTAK is critically dependent on IL-2 and human umbilical cord vein endothelial cells (HUVEC) as feeder cells. In HU-ATTAK, adding anti-OX40 ligand antibody into the culture system completely inhibited its proliferation. So, OX40 ligand as well as L-2 and/or IL-4 is suggested be necessary for the proliferation of ATLL cells, and feeder cells may also confer the favorable environment. As the substitute of HUVEC, follicular dendritic cell like cell line HK which expresses OX40 ligand was used by introducing human OX40 ligand cDNA (HK-OX40L). When 9 ATLL patient samples were co-cultured with HK-OX40L in the presence of L-2 and/or IL-4, two ATLL cells proliferate vigorously only in the presence of both IL-2 and IL-4 simultaneously. These cell lines were confirmed to be derived from original tumor cells by array CGH analysis, and continued the growth for more than a year. Depletion of IL-2 and IL-4 made these cell lines stop growing within 6 days even on HK-OX40L. In the presence of IL-2 and IL-4, the conditions such as HK alone without OX40 ligand or OX40 ligand alone without HK made the cell lines growing for three months at most. In the presence of IL-2 and IL-4 without HK-OX40L, these cell lines vigorously proliferated for more than three months but finally stopped growing. These data suggested that for the growth of these two cell lines, the cell division is dependent on IL-2 and IL-4, and the maintenance of immortalization is dependent on OX40 ligand and HK cells. This culture analysis would provide important factors for cell growth of ATLL which will explore new targets for ATLL treatment. *HK cells are kindly provided by Dr. Young S Choi at Ochsner Cancer Center, New Orleans. Disclosures: No relevant conflicts of interest to declare.

Phase I Study of KW-0761, a Humanized Anti-CCR4 Antibody, in Patients (Pts) with Relapsed or Refractory Adult T-Cell Leukemia-Lymphoma (ATLL) and Peripheral T-Cell Lymphoma (PTCL): Preliminary Results.

Introduction: KW-0761 is a defucosylated humanized IgG1 monoclonal antibody against CC chemokine receptor 4 (CCR4). Previous studies revealed that CCR4 was over-expressed on tumor cells from 88% of pts with ATLL and 38% of pts with PTCL. CCR4 expression was associated with unfavorable prognosis in both diseases. These evidences suggest that CCR4 could be a reasonable molecular target for treatment of CCR4-positive ATLL and PTCL. KW-0761 exerted very potent ADCC but not CDC against cultured ATLL cell lines using normal effecter cells. KW-0761 also exhibited potent ADCC against freshly isolated ATLL cells using the auto-effecter cells. Methods: KW-0761 is being investigated in a Phase I, single-agent, dose-escalation, multicenter study for relapsed or refractory pts with ATLL or PTCL after undergoing the initial chemotherapy. Eligible pts were required to express CCR4 on their tumor cells by FCM and/or IHC. This phase I trial was designed to evaluate toxicity, pharmacokinetics (PK), immunogenicity and anti-tumor activity. Toxicity was evaluated by NCI CTCAE (v3.0). Pts were planned to receive four weekly intravenous injections of KW-0761 at the doses of 0.01, 0.1, 0.5 and 1.0 mg/kg. Results: As of August 10, 2007, 6 pts (1 PTCL and 5 ATLL) have been treated with KW-0761 at the dose of 0.01 (N=3) and 0.1 mg/kg (N=3). Three pts were evaluable for toxicity, PK, immunogenicity and 4 pts were evaluable for anti-tumor activity. KW-0761 was well tolerated with no dose-limiting toxicities. Reversible grade (G) 3 toxicities were lymphocytopenia and herpes zoster. G1-2 adverse events were; rash, constipation, nausea, EF decrease, neutropenia, thrombocytopenia, eosinophilia, lymphocytopenia, ALP increase and QT prolongation. In one leukemic ATLL pt with swollen lymph nodes (LNs) treated at 0.01 mg/kg, peripheral ATLL cells disappeared and the LNs were decreased in size, attaining PR. In another leukemic ATLL pt with skin involvement treated at 0.1 mg/kg, peripheral ATLL cells disappeared and the skin achieved PR by Physicians Global Assessment of Clinical Conditions (PGA). One ATLL pt with skin and bone involvement at 0.01 mg/kg showed SD. Preliminary PK analysis at 0.01 mg/kg showed that Cmax and T1/2 after the 4th dosing was 323.7 56.7 ng/ml and 244 117 hrs, respectively. KW-0761 did not exhibit immunogenicity at 0.01 mg/kg. Conclusions: Preliminary phase I data warrant further investigations of KW-0761, a first-in-class humanized anti-CCR4 antibody, against CCR4-positive peripheral T-cell malignancies including ATLL. Patient accrual is ongoing and the updated results will be presented at the meeting.

Immunophenotypic diversity and prognosis of adult T-cell leukemia/lymphoma

17565 Background: Adult T-cell leukaemia/lymphoma (ATLL) is an aggressive disease associated with human T-cell lymphotropic virus type-I (HTLV-I) with heterogeneous clinical presentation and outcomes, described in Southern Japan, Europe, Caribbean and previously on Pacific coast of South America including Peru (EHA 2001, abst. 304). Shimoyama’s ATLL classification (BHJ 1991:79), includes four types: acute, lymphomatous, chronic and smouldering; recently a new clinical cutaneous type had been described (BJD 2005:152). Methods: We described our experienced previously (ASH 2005, abst. 4797). Herein we analyzed immunophenotypically our ATLL patient population by flow cytometry looking for aberrant phenotypes and their correlation with overall survival. The statistical method was descriptive and survival was calculated using the Kaplan-Meier method. Results: From July 1997 to March 2005 our registry had 69 cases, 37 had flow pre-treatment: acute type (n = 31) and lymphomatous (n = 6). Over our 37 flow done we were able to analysis only 33. The incidence of the typical (CD4+/CD8-) phenotype, the double-negative (CD4-/CD8-), the double-positive (CD4+/CD8+), and the CD8 positive (CD4-/CD8+) phenotypes was 58%, 12%, 15%, and 12% respectively. The median overall survival (OS) for the 33 patients was 4.1 months (range: 0.5–46.1). The patients with typical phenotypes had a median OS of 4.1 months better than the patients with the double-negative phenotype whom median OS was 2.0 (range: 1.5–9.7) but not significant. Median OS in the double positive and the CD4-/CD8+ phenotype population were 15.2 and 4.8 months respectively with no significance Conclusions: ATLL has a diversity of immunophenotype, our data suggest that there is not any correlation with survival, major accrual will gave us a final conclusion. No significant financial relationships to disclose.

A Human T-Cell Lymphotropic Virus Type 1 Enhancer of Myc Transforming Potential Stabilizes Myc-TIP60 Transcriptional Interactions

ABSTRACT The human T-cell lymphotropic virus type 1 (HTLV-1) infects and transforms CD4+ lymphocytes and causes adult T-cell leukemia/lymphoma (ATLL), an aggressive lymphoproliferative disease that is often fatal. Here, we demonstrate that the HTLV-1 pX splice-variant p30II markedly enhances the transforming potential of Myc and transcriptionally activates the human cyclin D2 promoter, dependent upon its conserved Myc-responsive E-box enhancer elements, which are associated with increased S-phase entry and multinucleation. Enhancement of c-Myc transforming activity by HTLV-1 p30II is dependent upon the transcriptional coactivators, transforming transcriptional activator protein/p434 and TIP60, and it requires TIP60 histone acetyltransferase (HAT) activity and correlates with the stabilization of HTLV-1 p30II/Myc-TIP60 chromatin-remodeling complexes. The p30II oncoprotein colocalizes and coimmunoprecipitates with Myc-TIP60 complexes in cultured HTLV-1-infected ATLL patient lymphocytes. Amino acid residues 99 to 154 within HTLV-1 p30II interact with the TIP60 HAT, and p30II transcriptionally activates numerous cellular genes in a TIP60-dependent or TIP60-independent manner, as determined by microarray gene expression analyses. Importantly, these results suggest that p30II functions as a novel retroviral modulator of Myc-TIP60-transforming interactions that may contribute to adult T-cell leukemogenesis.