Abstract 3547: In vitro and ex vivo anti-leukemia activity of AR-67, a novel lipophilic camptothecin

Abstract

Abstract BACKGROUND: The overall survival and disease-free rates for Acute Myelogenous Leukemia (AML) patients treated with the gold-standard therapy comprised of Cytarabine (Ara-C) and Daunorubicin (DNR) remain significantly low. Camptothecins are potent anticancer drugs that confer their cytotoxic effect by interfering with Topoisomerase-I (Top1). Semi-synthetic camptothecin analogues have been used in the past to treat AML patients with relatively low response rates. Here, we examined the anti-leukemia activity of AR-67, a novel camptothecin analogue, alone or in combination with standard agents for the treatment of AML. METHODS: The anti-leukemia activity of AR-67, TPT, Ara-C and DNR was studied in HL60, KG1 and K562 cells and in primary leukemia cell cultures after exposure to increasing drug concentrations for 48 hours. Cell proliferation was determined using a colorimetric assay (MTT) assay and IC50s were obtained using nonlinear-regression analysis. Drug combination treatments were assessed for synergy. The expression of Top1, phosphohistone (γ-H2AX), Bcl-2 and c-PARP were evaluated using biochemical assays. RESULTS: The potency of AR-67 was in the nM range in the leukemic cell lines. Consistent with the lower proliferation rates of primary cells grown in culture, their IC50 values were variable and in the µM range. IC50s estimated for TPT were higher than the respective ones for AR-67. The IC50 values for Ara-C and DNR were in the μM range for the HL60, K562 and primary cells, and higher than the ones for AR-67. However, the KG-1 cell line appeared to be more sensitive to the AML gold standard therapy, but not as sensitive as to AR-67. AR-67 treatment decreased Top1 expression, increased γ-H2AX expression in a concentration-dependent manner suggesting removal of DNA-Top1 complexes and formation of double strand DNA breaks. Induction of apoptosis was manifested by decreased Bcl-2 and increased c-PARP expression suggesting a caspace-3 mediated effect. AR-67 combination treatments that included DNR and Ara-C showed synergy in HL60 cells. Additionally, pretreatment with Ara-C revealed synergy between AR-67 and DNR in both the KG1 and K562 cells, accompanied by increased γ-H2AX and c-PARP expression. CONCLUSIONS: Our results indicate AR-67 as a potent anticancer agent that induces DNA damage and apoptosis. Using in vitro leukemia models we have shown that AR-67 is a potentially efficacious agent when combined with standard agents for the treatment of AML and may benefit AML patients resistant to Ara-C. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3547. doi:10.1158/1538-7445.AM2011-3547