Arsenic trioxide (ATO) is an inorganic arsenic derivative that is effective in patients with relapsed acute promyelocytic leukemia (APL) and is being evaluated as therapy for other leukemias and solid tumors. However, its use is limited by toxicity. In contrast, organic arsenic derivatives (OAD) are much less toxic ; hepatic methylation of inorganic arsenic compounds (like ATO) to yield OAD is a mechanism of detoxification. We hypothesized that, compared to ATO, selected OAD have similar clinical benefit with significantly lower toxicity. We have synthesized over 100 OADs and found some to have potent anti-cancer activity in the NIH 60 cell line screen. Based on its activity, we selected S-dimethylarsino-glutathione (SGLU1) for further study in our laboratory and have confirmed its potent activity against human leukemia, lymphoma, and multiple myeloma cell lines. In vivo anti-cancer activity of SGLU1 was evaluated by NIH in the P388 mouse lymphocytic leukemia model. SGLU1 administered IP at 100 mg/kg (highest dose tested) for 9 days resulted in a 64% increase in survival ; in contrast, others have shown in a murine model of APL, the disease most sensitive to ATO, that IP administration of ATO for 28 days at the maximum tolerated dose of 5 mg/kg was required for a 100% increase in survival. SGLU1 also exerts cytotoxic effects on leukemic cells from patients with leukemia. In vitro anti-leukemic activity of SGLU1 is comparable to that of ATO, but its toxicity against healthy donor cells is significantly less than that of ATO. Studies to determine the toxicity of a single dose injection of SGLU1 showed the concentration of SGLU1 that kills 50% of mice (LD50) to be 350 mg/kg, and for ATO to be 10 mg/kg. When administered IV daily for 5 days, LD50 for SGLU1 is 250 mg/kg/d (1250 mg/kg total dose), and 5 mg/kg/d for ATO (25 mg/kg total dose). For toxicology evaluation, 5 mice per sex received 5 daily IV injections ; doses escalated from a starting dose of 50 mg/kg/day. All surviving mice were sacrificed after 28 days. Microscopic observations included drug-related lesions in lung, liver, thymus and testes, but only at very high doses. The no-observed effect level (NOEL) for female mice was 150 mg/kg/day based on a single observation of centrilobular hypertrophy of hepatocytes in 1 of 5 female mice treated at a dose of 200 mg/kg/day. This event was not unexpected since SGLU1 is likely metabolized in the liver. The NOEL for male mice was 100 mg/kg/day based on testicular seminiferous tubular degeneration observed in 1 of 5 male mice treated at 150 mg/kg/day. Pharmaceutical evaluation of SGLU1 revealed its synthesis to be straightforward, yielding highly pure material, and sufficient solubility and stability for administration in a clinical setting. Ex vivo cardiotoxicity testing to determine the potential effects of escalating doses of SGLU1 on cardiovascular function in guinea pig hearts (the Langendorff preparation test) yielded no adverse effects in clinically relevant doses (up to 100 microM). A 28-day toxicity study with pharmacokinetics in Beagle dogs is underway. Studies designed to understand the mechanism of action of SGLU1 show that it has different mode of action than ATO (reported separately ASH 2003 abstract). Results of our work reflect a high potential for identification of a new therapeutic approach for patients with leukemia.

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The significance of terminal deoxynucleotidyl transferase (TdT) expression in acute myelogenous leukemia (AML) remains controversial. Therefore, we studied TdT expression by flow cytometry in 120 previously untreated patients with AML or myelodysplastic syndrome (MDS) to determine the distribution of TdT-positive blasts and the intensity of TdT expression and to seek clinically significant associations. TdT expression measured by flow cytometry (flow TdT%) was heterogeneous, ranging from 0.1% to 87% (median, 8.5%), and 74 patients (62%) had at least 5% TdT-positive blasts. TdT positivity was associated with the M0 or M1 subtype and with expression of CD34 and CD7. No significant correlation was found between TdT expression and type of cytogenetic abnormality or rearrangement of immunoglobulin or T-cell receptor genes. Remission lasted longer in patients with a flow TdT% < 5 than in patients with a flow TdT% > 5 (median, 95 weeks vs 55 weeks, p = 0.02); however, complete remission rates did not differ when patients were classified by initial flow TdT%. Survival was slightly better for patients with flow TdT% less than 5%. Among patients with a flow TdT% > 5%, those with a higher TdT intensity survived longer than those with a lower intensity. These data suggest that quantitative TdT measurement may contribute to prognostic estimate in AML patients.

A prognostic factor analysis for survival and response was conducted on 374 previously treated patients who were treated with one of four fludarabine regimens. As several prognostic factors were associated with response and survival, multivariate analyses were conducted to identify the combination of factors which best describe the prognosis as regards response to fludarabine and survival. Statistical models to identify the hazard for survival and probability of response were developed. The characteristics which were included in the model predicting for survival were sex, age, number of prior treatments, performance status, hemoglobin level, serum albumin, and alkaline phosphatase levels. The characteristics most strongly associated with response were the hemoglobin levels, Serum albumin, and the number of prior treatments. These models can be used to identify risk groups to guide physicians in decision-making and to assist in the design and analysis of clinical trials.

PURPOSE:To evaluate the efficacy and toxicity of Hyper-CVAD (fractionated cyclophosphamide, vincristine, doxorubicin, and dexamethasone), a dose-intensive regimen, in adult acute lymphocytic leukemia (ALL).PATIENTS AND METHODS:Adults with newly diagnosed ALL referred since 1992 were entered onto the study; treatment was initiated in 204 patients between 1992 and January 1998. No exclusions were made because of older age, poor performance status, organ dysfunction, or active infection. Median age was 39.5 years; 37% were at least 50 years old. Mature B-cell disease (Burkitt type) was present in 9%, T-cell disease in 17%. Leukocytosis of more than 30 x 10(9)/L was found in 26%, Philadelphia chromosome-positive disease in 16% (20% of patients with assessable metaphases), CNS leukemia at the time of diagnosis in 7%, and a mediastinal mass in 7%. Treatment consisted of four cycles of Hyper-CVAD alternating with four cycles of high-dose methotrexate (MTX) and cytarabine therapy, together with intrathecal CNS prophylaxis and supportive care with antibiotic prophylaxis and granulocyte colony-stimulating factor therapy. Maintenance in patients with nonmature B-cell ALL included 2 years of treatment with mercaptopurine, MTX, vincristine, and prednisone (POMP).RESULTS:Overall, 185 patients (91%) achieved complete remission (CR) and 12 (6%) died during induction therapy. Estimated 5-year survival and 5-year CR rates were 39% and 38%, respectively. The incidence of CNS relapse was low (4%). Compared with 222 patients treated with vincristine, doxorubicin, and dexamethasone (VAD) regimens, our patients had a better CR rate (91% v 75%, P <.01) and CR rate after one course (74% v 55%, P <.01) and better survival (P <.01), and a smaller percentage had more than 5% day 14 blasts (34% v 48%, P =.01). Previous prognostic models remained predictive for outcome with Hyper-CVAD therapy.CONCLUSION:Hyper-CVAD therapy is superior to our previous regimens and should be compared with established regimens in adult ALL.

Preclinical data have shown that all-trans retinoic acid (ATRA) with interferon-alpha (IFN-alpha) can exert significant suppressive effects on Philadelphia-chromosome (Ph)-positive cells. The aim of this study combining IFN-alpha, low-dose cytosine arabinoside (ara-C) and ATRA was to increase the proportion of patients achieving a major cytogenetic response, in comparison with a group of 140 patients previously treated with IFN-alpha plus low-dose ara-C. Forty three patients with Ph-positive CML in early chronic phase were treated with IFN-alpha 5 MU/m2 s.c. daily, low-dose ara-C 10 mg s.c. daily and ATRA 45 mg/m2 orally daily, for 7 consecutive days every other week. Overall, 76% of patients achieved a complete hematologic response (CHR). A cytogenetic response was in observed 59% (major in 38% and complete in 17%). Compared with patients treated with IFN-alpha and low-dose ara-C, those receiving additional ATRA had a lower CHR rate (p. 014), but other response rates were similar. Severe toxicities were common with the triple regimen (64%), mostly related to ATRA therapy. Two patients experienced pseudotumor cerebri; two patients had leukocytosis during the week on ATRA treatment, decreasing during the week off (one suffered a severe asthma-like reaction followed by pulmonary edema, resembling ATRA syndrome). Six patients had other unusual side-effects: aseptic necrosis of the hip (1 patient), ataxic syndrome (1 patient), paranoid syndrome (2 patients), syncopal episodes (1 patient), pure red cell aplasia (1 patient). In conclusion the results of IFN-alpha and low-dose ara-C combined with ATRA in patients with early CML-chronic phase were disappointing, due to excessive toxicity. Whether different ATRA dose schedules may result in fewer side-effects and improve hematologic and cytogenetic response remains to be determined.

The recently characterized immunotoxin HuM195-gelonin consists of a humanized anti-CD33 monoclonal antibody conjugated to the single-chain plant toxin gelonin. Binding of the immunotoxin to hematopoietic cells that express the CD33 differentiation antigen has been demonstrated and results in cytotoxicity due to ribosomal inactivation by gelonin. Blast cells from most patients with acute myelogenous leukemia express CD33, whereas normal stem cells necessary for maintenance of hematopoiesis do not. We asked whether an immunoconjugate using recombinant gelonin rather than plant gelonin is toxic to acute myelogenous leukemia (AML) cell lines and primary AML blasts obtained from patients and exposed to the immunotoxin in vitro. The CD33pos cell lines HL60, OCI/AML2, and OCI/ AML5 showed decreased proliferation when exposed to immunotoxin for 24-72 h. The CD33neg cell line OCI/AML3 was relatively resistant to HuM195, and all cell lines were resistant to equimolar concentrations of unconjugated antibody and gelonin. Primary blast cultures from seven patients with AML had CD33 detectable on 75.7-99.8% of cells by flow cytometry, and all showed dose-dependent decreases in clonogenic cell survival during 24-h incubation with the immunotoxin. Cells selected for low CD33 expression by cell sorting or by prolonged incubation with immunotoxin could reexpress CD33 at baseline levels and remained sensitive to immunotoxin. We conclude that humanized M195 conjugated to recombinant gelonin has antileukemic activity and should be considered for clinical testing in Phase I trials.