bafilomycin-a1 and Necrosis

A novel series of methylene-substituted DIMs (C-DIMs), namely 1,1-bis(3'-indolyl)-1-(p-substituted phenyl)methanes containing t-butyl (DIM-C-pPhtBu) and phenyl (DIM-C-pPhC6H5) groups inhibit proliferation of invasive estrogen receptor-negative MDA-MB-231 and MDA-MB-453 human breast cancer cell lines with IC50 values between 1-5 uM. The main purpose of this study was to investigate the pathways of C-DIM-induced cell death.. The effects of the C-DIMs on apoptotic, necrotic and autophagic cell death were determined using caspase inhibitors, measurement of lactate dehydrogenase release, and several markers of autophagy including Beclin and light chain associated protein 3 expression (LC3).. The C-DIM compounds did not induce apoptosis and only DIM-C-pPhCF3 exhibited necrotic effects. However, treatment of MDA-MB-231 and MDA-MB-453 cells with C-DIMs resulted in accumulation of LC3-II compared to LC3-I protein, a characteristic marker of autophagy, and transient transfection of green fluorescent protein-LC3 also revealed that treatment with C-DIMs induced a redistribution of LC3 to autophagosomes after C-DIM treatment. In addition, the autofluorescent drug monodansylcadaverine (MDC), a specific autophagolysosome marker, accumulated in vacuoles after C-DIM treatment, and western blot analysis of lysates from cells treated with C-DIMs showed that the Beclin 1/Bcl-2 protein ratio increased.. The results suggest that C-DIM compounds may represent a new mechanism-based agent for treating drug-resistant ER-negative breast tumors through induction of autophagy.

Topics: Amino Acid Chloromethyl Ketones; Animals; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Autophagy; Beclin-1; Biomarkers, Tumor; Blotting, Western; Breast Neoplasms; Caspase Inhibitors; Caspases; Cell Line, Tumor; Cell Proliferation; Cysteine Proteinase Inhibitors; Female; Humans; Immunohistochemistry; Indoles; L-Lactate Dehydrogenase; Macrolides; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Nude; Microtubule-Associated Proteins; Necrosis; Proto-Oncogene Proteins c-bcl-2; Receptors, Estrogen; Recombinant Fusion Proteins; Time Factors; Transfection; Xenograft Model Antitumor Assays