Assessment of the molecular mechanisms of drug-induced hidden cardiotoxicity by a multi-omics approach: The example of rofecoxib

Abstract

Background and Purpose: Hidden cardiotoxicity is defined as drug-induced cardiotoxicity that becomes obvious only in the presence of comorbidities. However, the molecular mechanisms of hidden cardiotoxicity are not always known. Therefore, unbiased multi-omics approaches could assist in revealing regulatory pathways. The most notable representative of hidden cardiotoxic drugs is the cyclooxygenase-2-inhibitor, rofecoxib. We previously reported increased mortality in rats because of proarrhythmic effects of rofecoxib in ischaemic hearts. Here, we aimed to identify molecular mechanisms of hidden cardiotoxicity exemplified by rofecoxib that present prior to comorbidities. Experimental Approach: Rats were treated with rofecoxib or its vehicle for 4 weeks. RNA sequencing and proteomic datasets of heart samples were used for differential expression and pathway reconstruction analyses. Key Results: In this model, mechanisms of hidden cardiotoxicity could not be revealed by transcriptomic analyses. However, mass-spectrometry-based proteomics showed conspicuous changes, revealing 132 proteins that were dysregulated in expression or on phosphorylation sites. Importantly, the phospho-proteomics allowed us to identify two kinases that may mediate cardiotoxicity. Finally, pathway reconstruction maps a complex molecular machinery whose clustered proteins regulate processes involving cytoskeleton binding, mRNA processing, proteolysis, translation, citrate acid cycle and calcium ion signalling. Conclusion and Implications: This is the first demonstration that multi-omics characterisation can reveal underlying regulatory pathways of hidden cardiotoxicity. Importantly, our study shows that transcriptomics gives limited information on the hidden cardiotoxic effects of rofecoxib, which are mainly mediated by changes in posttranslational modifications and protein expression. These changes, among other mechanisms, may disturb the cardiac calcium handling, which could explain the fatal arrhythmias following ischaemia/reperfusion observed with rofecoxib.