Abstract 19146: Disruption of TRAF2-TAK1-IKKB Signaling Triggers Mitochondrial Perturbations and Necrotic Cell Death in Doxorubicin Cardiotoxicity

Abstract

The anthracycline doxorubicin (dox) is a highly effective anti-tumour agent, however, its use is limited by its severe cardiotoxic effects that manifests as heart failure. The decline in cardiac performance induced by doxorubicin remains poorly defined. A critical survival role for the canonical IKKβ -NF-κB signaling pathway has been demonstrated in ventricular myocytes. In this report, we demonstrate that, dox impairs IKKβ- NF-κB signaling in ventricular myocytes accompanied by mitochondrial perturbations including mPTP, loss of membrane potential and ROS production. IKKβ- NF-κB signaling involves TRAF 2 mediated ligation of K63 ubiquitin chains to RIP1 (Receptor Interacting Protein 1) which serves as scaffold for recruitment of ubiquitylated Tak1 complex and phosphorylation-dependent activation of IKKβ -NF-kB signaling. Interestingly, ventricular myocytes treated with dox demonstrated reduction in expression levels of TRAF2 and TAK1. This was accompanied by a decline in K63- ubiquitylation chains and concomitant increase in K-48 polyubiquitination of RIP1, impaired NF-kB activation and necrotic cell death. Interestingly, inhibiting the kinase activity of RIP1 with Necrostatin-1, (Nec1) suppressed necrotic cell injury induced by dox but not NF-kB activation. Concordant with these findings was a marked increase in necrotic cell death in cardiac myocytes defective for IKKB signaling or MEF cells deficient for p65 treated with dox. Notably, mitochondrial perturbations, including PT-pore opening , ROS production, calcium uptake, LDH, Tn(T) and HMGB-1 release and necrotic cell injury induced by dox were completely abrogated by restoring IKKB-NF-kB signaling in cardiac myocytes or Nec-1. Herein, we provide novel evidence that K-48 ubiquitylation of RIP1 by doxorubicin provides a functional switch that signals necrotic cell death in cells defective for IKKβ-NF-kB signaling. Hence, interventions that modulate IKKB- NF-kB activation may prove beneficial in mitigating the cardiotoxic effects of dox.