In Vivo Anti-Leukemia Activity Of Novel C-Ring Modified Prodigiosenes In A Zebrafish Xenograft Model

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Acute myeloid leukemia (AML) remains fatal for 40% of patients, both due to refractory disease and toxicity from traditional therapeutic agents. Zebrafish provide an efficient and robust leukemia model with the capacity for embryonic screens to test new drugs in vivo. Naturally-occurring Prodigiosin has demonstrated potential as an anti-cancer agent, but toxicity prevents its use as a chemotherapeutic. Here, we present four novel derivatives, prodigiosenes designated DSB-8, -31, -39, -50, synthesized using a multi-step sequence beginning with simple pyrroles. These synthetic compounds demonstrated anti-leukemia activity in vitro against cells in the NIH/NCI Developmental Therapeutics Program, including K562, CCRF-CEM, MOLT-4, RPMI-8226 and SR cell lines. We have previously xenografted human leukemia cells into 48 hour zebrafish embryos and developed a cell quantification assay to evaluate drug responses (Corkery et al BJH, 2011). To assess the anti-leukemia activity and toxicity of our novel prodigiosenes in vivo, we first conducted toxicity curves using 48-hour zebrafish embryos treated for 72 hours to determine an optimal dose (50% of maximum tolerated dose [MTD]). Prodigiosenes DSB-39 (1.0 uM) and DSB-50 (1.5 uM) were tolerated better than DSB-8 (0.2 uM) and DSB-31 (0.2 uM). Subsequently, 40-50 CM-DiI-labeled K562 chronic myelogenous leukemia (CML) cells were injected into the yolk sac of 48-hour zebrafish embryos, which were treated with prodigiosenes at 50% MTD. Engrafted embryos were followed using live cell microscopy. Embryos treated with 0.03% DMSO served as negative control, showing abundant K562 cell proliferation and entry of cells into circulation. Embryos treated with 20 µM imatinib mesylate (IM), a targeted inhibitor of the BCR-ABL1 oncoprotein in K562 cells, served as a positive control, demonstrating no cell proliferation or migration. By comparison, prodigiosenes inhibited K562 cell activity to a similar or greater degree than IM. Qualitatively, the effects of prodigiosenes were graduated: DSB-8 (n=6/7) was cytostatic, DSB-31 (n=5/8) and DSB-50 (n=6/9) were moderately cytotoxic, and DSB-39 (n=7/7) was strongly cytotoxic. To quantify this effect, we are applying a modified ex vivo proliferation assay. Embryos are dissociated to a single cell suspension at 24 and 72 hours post-injection followed by immunohistochemistry directed to human CD34. Our work extends the use of zebrafish xenografts to determine the in vivo sensitivity of human cancer cells to novel drugs and suggests that tailored prodigiosenes may represent novel therapeutic agents for leukemia with improved anti-cancer potency and reduced toxicity. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4258. doi:1538-7445.AM2012-4258