Discovery of cytotoxic and pro-apoptotic compounds against leukemia cells: Tert-butyl-4-[(3-nitrophenoxy) methyl]-2,2-dimethyloxazolidine-3-carboxylate

Aims: We evaluated biological activity in leukemia cells lines of R and S enantiomers of tert-butyl 4-[(3-nitrophenoxy)-methyl]-2,2-dimethyloxazolidine-3-carboxylate (BNDC).

Main methods: Cytotoxic activity was assessed by MTT assay. Flow cytometry assays were used to determined DNA fragmentation (Propidium Iodide-PI staining) and phosphatidylserine exposure (Annexin-V and PI staining). DNA condensation was evaluated by fluorescence microscopy using double-staining in leukemia cells (Hoechst and PI). Caspase activities were measured using Z-VAD-FMK, a non-selective Caspase Inhibitor, by flow cytometry and Z-DEVD-AMC, a selective Caspase-3 substrate, by fluorescence spectrometry.

Key findings: Both enantiomers displayed cytotoxic activity against leukemia cell lines (HL60, HL60.Bcl-2, HL60.Bcl-xL and Jurkat) with low toxicity against human peripheral blood mononuclear cell--PBMC based on IC50 values. In HL60 cell lines, compounds induce exposure of phosphatidylserine and DNA fragmentation, which could be blocked by pretreatment of cells with Z-VAD-FMK. Confirming this observation, both enantiomers induced Caspase-3 activation. Additional analysis revealed an increased percentage of apoptotic cells (defined as those with fragmented nuclei and condensed chromatin) after treatment with compounds.

Significance: Taken together, the results indicate that BNDC compounds exhibited cytotoxic and pro-apoptotic activities and have a potential for developing a new class of Anticancer drugs.