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

Abstract Aim To 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 methods After the synthesis and IR, 1H, 13C NMR and mass spectral characterization of NPN, its antiproliferative efficacy was investigated against cells of breast cancer (MCF-7 and MDA-MB-231), lung cancer (HOP-62 and A-549), and of a non-cancerous epithelial line (VERO), using the 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 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 using staining the cells with DCFDA, Rhodamine 123, and acridine orange/ethidium bromide, respectively. Key findings NPN 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. NPN showed very little toxicity to the non-cancerous cell line, VERO. 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. Significance NPN can be explored further as a chemotherapeutic agent against TNBC.