The investigation of ceranib-2 on apoptosis and drug interaction with carboplatin in human non small cell lung cancer cells in vitro
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Ceramide is found to be involved in inhibition of cell division and induction of apoptosis in certain tumour cells. Ceranib-2 is an agent that increases ceramide levels by inhibiting ceramidase in cancer cells. Therefore, we aimed to investigate the effects of ceranib-2 on cell survival, apoptosis and interaction with carboplatin in human non-small cell lung cancer cells. The cytotoxic effect of ceranib-2 (1–100 µM) was determined by MTT assay in human lung adenocarcinoma (A549) and large cell lung carcinoma (H460) cells. Carboplatin (1–100 µM) and lung bronchial epithelial cells (BEAS-2B) were used as positive controls. Morphological and ultrastructural changes were analysed by light microscope and TEM. Apoptotic/necrotic cell death and acid ceramidase activity were analysed by ELISA. Combination effects of ceranib-2 and carboplatin were investigated by MTT. The expression levels of CASP3, CASP9, BAX and BCL-2 were examined by qRT-PCR. The IC50 of ceranib-2 was determined as 22 μM in A549 cells and 8 μM in H460 cells for 24 h. Morphological changes and induction of DNA fragmentation have revealed apoptotic effects of ceranib-2 in both cell lines. Ceranib-2 and carboplatin has shown synergism in combined treatment at 10 and 25 μM doses in H460 cells for 24 h. Ceranib-2 inhibited acid ceramidase activity by 44% at 25 µM in H460 cells. Finally, CASP3, CASP9 and BAX expressions were increased while BCL-2 expression was reduced in both cells. Our results obtained some preliminary results about the cytotoxic and apoptotic effects of ceranib-2 for the first time in NSCLC cell lines.
KeywordsCeranib-2 Ceramidase inhibitor Cytotoxicity Apoptosis Combined therapy
This publications was based completely on M.Sc. thesis of the first author, Merve YILDIZ.
This study was supported by TUBITAK (The Scientific and Technical Research Council of Turkey), (Project Number: 214Z159).
- Baran Y, Salas A, Senkal CE, Gunduz U, Bielawski J, Obeid LM, Ogretmen B (2007) Alterations of ceramide/sphingosine 1-phosphate rheostat ınvolved in the regulation of resistance to ımatinib-induced apoptosis in K562 human chronic myeloid leukemia cells. J Biol Chem 282:10922–10934. doi: 10.1074/jbc.M610157200 CrossRefGoogle Scholar
- Cheng JC, Bai A, Beckham TH, Marrison ST, Yount CL, Young K, Lu P, Bartlett AM, Wu BX, Keane BJ, Armeson KE, Marshall DT, Keane TE, Smith MT, Jones EE, Drake RR, Bielawska A, Norris JS, Liu X (2013) Radiation-induced acid ceramidase confers prostate cancer resistance and tumor relapse. J Clin Invest 123:4344–4358. doi: 10.1172/JCI64791 CrossRefGoogle Scholar
- Obeid LM, Linardic CM, Karolak LA, Hannun YA (1993) Programmed cell death induced by ceramide. Science 259:1769–1771. http://www.ncbi.nlm.nih.gov/pubmed/8456305
- Pettus BJ, Chalfant CE, Hannun YA (2002) Ceramide in apoptosis: an overview and current perspectives. Biochim Biophys Acta 1585:114–125. http://www.ncbi.nlm.nih.gov/pubmed/12531544
- Sawada M, Nakashima S, Banno Y, Yamakawa H, Hayashi K, Takenaka K, Nishimura Y, Sakai N, Nozawa Y (2000) Ordering of ceramide formation, caspase activation, and Bax/Bcl-2 expression during etoposide-induced apoptosis in C6 glioma cells. Cell Death Differ 7:761–772. doi: 10.1038/sj.cdd.4400711 CrossRefGoogle Scholar
- Soans E, Evans SC, Cipolla C, Fernandes E (2014) Characterizing the sphingomyelinase pathway triggered by PRIMA-1 derivatives in lung cancer cells with differing p53 status. Anticancer Res 34:3271–3283Google Scholar
- Taha TA, Hannun YA, Obeid LM (2006a) Sphingosine kinase: biochemical and cellular regulation and role in disease. J Biochem Mol Biol 39:113–131. http://www.ncbi.nlm.nih.gov/pubmed/16584625