Tubulin- and ROS-dependent antiproliferative mechanism of a potent analogue of noscapine, N-propargyl noscapine

Abstract

AimTo rationally-design, synthesize, characterize, biologically evaluate, and to elucidate the anticancer mechanism of action of a novel analogue of noscapine, N-propargyl noscapine (NPN), as a potential drug candidate against triple-negative breast cancer (TNBC). Materials and methodsAfter the synthesis and IR, 1H, 13C NMR and mass spectral characterization of NPN, its antiproliferative efficacy against different cancer cell lines was investigated using Sulforhodamine B assay. Cell cycle progression was analysed using flow cytometry. The drug-tubulin interactions were studied using tryptophan-quenching assay, ANS-binding assay, and colchicine-binding assay. Immunofluorescence imaging was used to examine the effect of NPN on cellular microtubules. Levels of reactive oxygen species (ROS), loss of mitochondrial membrane potential (MMP), and cell death were studied by staining the cells with DCFDA, Rhodamine 123, and acridine orange/ethidium bromide, respectively. Key findingsNPN strongly inhibited the viability (IC50, 1.35 ± 0.2 μM) and clonogenicity (IC50, 0.56 ± 0.06 μM) of the TNBC cell line, MDA-MB-231, with robust G2/M arrest. In vitro, the drug bound to tubulin and disrupted the latter's structural integrity and promoted colchicine binding to tubulin. NPN triggered an unusual form of microtubule disruption in cells, repressed recovery of cold-depolymerized cellular microtubules and suppressed their dynamicity. These effects on microtubules were facilitated by elevated levels of ROS and loss of MMP. SignificanceNPN can be explored further as a chemotherapeutic agent against TNBC.