Mechanistic insight into SAHA-induced autophagic cell death in breast cancer cells

Abstract

In recent years histone deacetylase inhibitors (HDACis) a novel class of agents that targets mechanistic abnormalities in cancers have shown promising anti-cancer activity in both haematological and solid cancers Among them SAHA (Vorinostat a pan-HDACi) is approved by FDA to treat cutaneous T-cell lymphoma and is being evaluated in other cancer types including breast cancer Although several studies have shown that SAHA exhibits potent anti-proliferative effects and induces autophagic cell death the exact mechanisms by which SAHA induces autophagic cell death have not been fully understood Therefore the aim of this study is to investigate the underlying mechanisms of SAHA-induced autophagic cell death in cancer cells In our study we observed that SAHA a pan-HDAC inhibitor downregulated the expression of survivin and XIAP in both concentration- and time-dependent manner Through Western blotting lysotracker and MDC staining indunction of LC-3B conversion and autophagosome/autolysosome formations were observed in the cells treated with SAHA MTT cell viability assay Western blotting showed that the addition of YM155 and Embelin (survivin and XIAP inhibitor respectively) further depleted survivin and XIAP levels respectively thereby enhancing the autophagic and cytotoxic effects of SAHA in the treated cells Downregulation of survivin by siRNA also increased the sensitivity of MCF-7 cells to SAHA At the molecular level RT-PCR analysis revealed that at high concentration (2xIC50) SAHA inhibited survivin gene transcription Subsequent protein stability assay and Western blotting showed that SAHA reduced the protein stability of survivin and XIAP possibly through modulating 26S proteasome and heat-shock protein 90 We further performed Western blotting with the addition of various selective HDAC inhibitors (BML281 MGCD0103 TSA and MC1568) where the results showed that HDAC 1 2 3 and 6 may be the contributors in SAHA-mediated survivin and XIAP depletion in MCF-7 cells By specifically silencing HDAC1-6 isoforms with siRNA we demonstrated that HDAC 3 and HDAC 6 isoforms are the major contributors in suppressing survivin and XIAP in both cancer cells Taken together proteasomal degradation of survivin and XIAP promoted by HDAC 3 and 6 isoforms inhibition might play important roles in the SAHA-induced autophagy and autophagic cell death in breast cancer models Importantly identification of how each HDAC isoforms regulate members of IAP family can potentially be applied to predict novel HDACI in targeted or combinational therapies in the future

Date of Award	2014 Jul 28
Original language	English
Supervisor	Chun Hei Antonio Cheung (Supervisor)