Benzamide riboside induced mitochondrial mediated apoptosis in human lung cancer H520 cells.
Benzamide riboside (BR) is a novel anticancer agent exhibiting pronounced activity against several human tumor cell lines via the inhibition of inosine 5'-monophosphate dehydrogenase (IMPDH), thereby restricting the biosynthesis of guanylates. Although it has been demonstrated that BR inhibits IMPDH and induces apoptosis, however, not much attention has been directed to the mechanism of apoptosis induction by this compound. The purpose of the present investigation was to investigate the mechanism of cytotoxicity induced by BR in human lung cancer cells. Non-small cell lung cancer [NSCLC] is the most prevalent type of lung cancer especially in India, and displays resistance to anticancer treatment. The results reveal that BR at a dose of 50 microM induces apoptosis in NSCLC H520 cells. This was ascertained by alteration in cellular morphology, TUNEL assay and flow cytometry. While Bax protein level was unaffected there was down regulation of anti-apoptotic Bcl-2 protein and up regulation of p53 as observed by Western blotting. Induction of apoptosis was accompanied by significant increase in caspase-3 activity. BR is a potent growth inhibitory pro-drug rationally synthesized to mimic NAD and inhibits PARP at high concentrations when assayed in permeabilized leukemic cells. Our observations showed that increased caspase-3 activity was accompanied by PARP cleavage. We also observed release of cytochrome c from mitochondria to the cytosol whereas no change was seen in the levels of apoptosis inducing factor (AIF). These findings indicate that BR induces apoptosis in H520 cells via the intrinsic mitochondrial pathway.
Khanna
Neeru
N
Department of Biochemistry, All India Institute of Medical Sciences, Room No 3027-Ansari Nagar, New Delhi-110029, India.
Jayaram
H N
HN
Singh
Neeta
N
D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002289:Carcinoma, Non-Small-Cell Lung / Q000378:metabolism*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors*; D020287:In Situ Nick-End Labeling; D008175:Lung Neoplasms / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*; D009705:Nucleosides / Q000378:metabolism / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D000970:Antineoplastic Agents; D009705:Nucleosides; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; C076145:3-(1-deoxyribofuranosyl)benzamide; D007168:IMP Dehydrogenase; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases