Abstract 195: Epigenetic Mechanisms Underlying Anthracycline-Induced Cardiotoxicity

Abstract

Anthracycline-induced cardiotoxicity (AC) is a major cause of morbidity in childhood cancer survivors. Doxorubicin cardiotoxicity however, is consistently observed to develop chronically over time. We sought to develop a mouse model of pediatric AC to test our hypothesis for an epigenetic mechanism, which may be operating to explain delayed-onset AC. Three week-old male and female C57Bl/6J mice were administered 5 weekly injections with doxorubicin (1mg/kg) (n=24) or saline (n=8). The dosage used was intended to closely mimic the clinical scenario. Half of the doxorubicin-treated (DOX) mice (n=12) and all controls (n=8) were sacrificed 1 week after the last injection; the rest of DOX mice (n=12) were given 4 weeks of drug-free holiday before sacrifice. Echocardiographic measurements were obtained before the first injection, 1 week after the final injection, and before sacrifice. Gene expression profiles were assessed by RNA-sequencing, reduced representation bisulphite sequencing (RRBS), and by ATAC-seq. These analyses were performed on purified pools of isolated cardiac myocytes, procured using a published Langendorff-free method. There was no death in the cohort. LV chamber dimension and wall thickness as well as fractional shortening did not differ between DOX and controls, implying the lack of obvious functional insufficiency from this dose. Similarly, DOX mice did not differ in body weight, heart weights (HW), and HW-to-tibia length ratios compared with controls. Cardiomyocyte dimensions and myocardial interstitial fibrosis were the same between groups. Despite the lack of echo and histological changes in DOX mice, RNA-seq analysis revealed profound and significant differentially expressed genes between the DOX mice and controls. Importantly, a significant proportion of downregulated and upregulated genes persisted, despite the period of drug-free holiday. Moreover, despite being taken off DOX, cardiomyocytes appeared to display new gene expression changes taking place even 4 weeks after stopping DOX. Detailed results of RNA-sequencing, RRBS and ATAC-Seq will be presented during the meeting. These results suggest that cardiomyocytes display DOX-related epigenetic alterations, which may implicate the basis for delayed-onset AC.