Abstract 1673: Adiponectin induces autophagic cell death in breast cancer cells through SIRT1 mediated deacetylation of LKB1 leading to ATG1 activation.

Abstract

Abstract Introduction: Obesity is an independent risk factor for the development of breast cancer. Molecular effects of obesity are mediated via perturbations in the adipocytokine profile. Adiponectin is widely considered as an adipocytokine with therapeutic potential as it inhibits growth of breast cancer cells but the mechanisms underlying growth inhibitory effects of adiponectin are still elusive. The present study was designed to systematically elucidate the underlying mechanisms by which adiponectin inhibits growth of breast cancer cells. Methods: Characteristic of autophagy were evaluated using transmission electron microscopy, acridine orange staining and western blot analysis of key effector molecules of autophagy. RT-PCR, western blot and immunofluorescence analysis were used to examine SIRT1 (Sirtuin 1), liver kinase B1 (LKB1), and AMP-activated protein kinase (AMPK) axes. Functional importance of AMPK activation and LKB1 overexpression in the biologic effects of adiponectin was examined by using AMPK-null and AMPK-wild type (WT) immortalized mouse embryonic fibroblasts (MEFs) and isogenic LKB1-knockdown cell line pairs. LKB1 immunoprecipitates were used to examine LKB1 deacetylation in response to adiponectin. Results: We provide strong evidence that adiponectin causes autophagy in breast cancer cells. Adiponectin treated breast cancer cells exhibit autophagosomes and major autophagosomal protein level changes including ATG1 (autophagy related 1) activation, cleavage of LC3 and suppression of p62 (p62/SQSTM1) expression. Adiponectin-induced autophagy leads to breast cancer cell death as evident by increased levels of cleaved-caspase 9, cleaved-PARP, and Tunel positivity. Adiponectin-mediated autophagy-induction as well as autophagic cell death is attenuated in the presence of autophagy inhibitors. Analysis of the underlying molecular mechanisms reveals that adiponectin treatment increases AMPK activation that is required for adiponectin-mediated ATG1 overexpression and phosphorylation. Intriguingly, we discover that adiponectin increases SIRT1 expression which deacetylates LKB1 leading to its activation. LKB1 knock-down inhibits adiponectin mediated autophagy induction as evidenced by modulations in major autophagosomal protein levels including ATG1. Analysis of breast tumors treated with adiponectin reveals significant increase in the levels of SIRT1, LKB1, phospho AMPK, phospho ATG1 as well as decreased p62. Conclusions: These data uncover a novel role of adiponectin as an inducer of autophagic cell death and provide the first in vitro and in vivo evidence of the integral role of the SIRT1-LKB1-ATG1 axis in adiponectin mediated autophagic cell death of breast cancer cells. Citation Format: Seung J. Chung, Neeraj Saxena, Dipali Sharma. Adiponectin induces autophagic cell death in breast cancer cells through SIRT1 mediated deacetylation of LKB1 leading to ATG1 activation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1673. doi:10.1158/1538-7445.AM2013-1673