A microtubule inhibitor, ABT-751, induces autophagy and delays apoptosis in Huh-7 cells

Journal Article · Tue Nov 15 04:00:00 EST 2016 · Toxicology and Applied Pharmacology

DOI:https://doi.org/10.1016/J.TAAP.2016.09.021· OSTI ID:22690852

Wei, Ren-Jie ^[1]; Lin, Su-Shuan ^[2]; Wu, Wen-Ren ^[1]; Chen, Lih-Ren ^[3]; Li, Chien-Feng ^[3]; Chen, Han-De; Chou, Chien-Ting; Chen, Ya-Chun ^[1]; Liang, Shih-Shin ^[1]; Chien, Shang-Tao ^[2]; Shiue, Yow-Ling ^[1]

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1. Institute of Biomedical Sciences, National Sun Yat-sen University, Kaohsiung, Taiwan (China)
2. Department of Pathology, Kaohsiung Armed Forces General Hospital, Kaohsiung, Taiwan (China)
3. Department of Biotechnology, Southern Taiwan University of Science and Technology, Tainan, Taiwan (China)

The objective was to investigate the upstream mechanisms of apoptosis which were triggered by a novel anti-microtubule drug, ABT-751, in hepatocellular carcinoma-derived Huh-7 cells. Effects of ABT-751 were evaluated by immunocytochemistry, flow cytometric, alkaline comet, soft agar, immunoblotting, CytoID, green fluorescent protein-microtubule associated protein 1 light chain 3 beta detection, plasmid transfection, nuclear/cytosol fractionation, coimmunoprecipitation, quantitative reverse transcription-polymerase chain reaction, small-hairpin RNA interference and mitochondria/cytosol fractionation assays. Results showed that ABT-751 caused dysregulation of microtubule, collapse of mitochondrial membrane potential, generation of reactive oxygen species (ROS), DNA damage, G{sub 2}/M cell cycle arrest, inhibition of anchorage-independent cell growth and apoptosis in Huh-7 cells. ABT-751 also induced early autophagy via upregulation of nuclear TP53 and downregulation of the AKT serine/threonine kinase (AKT)/mechanistic target of rapamycin (MTOR) pathway. Through modulation of the expression levels of DNA damage checkpoint proteins and G{sub 2}/M cell cycle regulators, ABT-751 induced G{sub 2}/M cell cycle arrest. Subsequently, ABT-751 triggered apoptosis with marked downregulation of B-cell CLL/lymphoma 2, upregulation of mitochondrial BCL2 antagonist/killer 1 and BCL2 like 11 protein levels, and cleavages of caspase 8 (CASP8), CASP9, CASP3 and DNA fragmentation factor subunit alpha proteins. Suppression of ROS significantly decreased ABT-751-induced autophagic and apoptotic cells. Pharmacological inhibition of autophagy significantly increased the percentages of ABT-751-induced apoptotic cells. The autophagy induced by ABT-751 plays a protective role to postpone apoptosis by exerting adaptive responses following microtubule damage, ROS and/or impaired mitochondria. - Highlights: • An anti-microtubule agent, ABT-751, induces autophagy and apoptosis in Huh-7 cells. • ABT-751 induces early autophagy and delays apoptosis. • ABT-751 induces autophagy via modulations of nuclear TP53 and AKT/MTOR pathways. • ABT-751-induced apoptosis is ROS-, mitochondria- and caspase-dependent. • Inhibition of autophagy enhances ABT-751-induced apoptosis.

• OSTI ID: 22690852
• Journal Information: Toxicology and Applied Pharmacology, Journal Name: Toxicology and Applied Pharmacology Vol. 311; ISSN TXAPA9; ISSN 0041-008X
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
AGAR
APOPTOSIS
CELL CYCLE
DNA
DNA DAMAGES
DRUGS
FLUORESCENCE
HEPATOMAS
INHIBITION
LYMPHOMAS
MICROTUBULES
MITOCHONDRIA
MODULATION
PHOSPHOTRANSFERASES
POLYMERASE CHAIN REACTION
POLYMERASES
RNA
SERINE
THREONINE