LAUSR

Impaired mitochondrial function has been implicated as a mechanism of doxorubicin-induced cardiotoxicity, however the precise genes that regulate this process in human cardiomyocytes remain to be elucidated. We hypothesized that doxorubicin significantly alters expression of genes involved in mitochondrial function in human cardiomyocytes, which in turn impairs mitochondrial respiration. Towards this, we treated human inducible pluripotent stem cell (iPSC)-cardiomyocytes with doxorubicin at 15 different time and dose conditions. Gene expression was assessed by RNAseq for each condition and 169 genes involved in mitochondrial function were analyzed for differential expression between control and treated conditions. Mitochondrial respiration (basal respiration, ATP production, maximal respiration, and spare respiratory capacity) was measured using the Seahorse Bioscience XFe96 Cell Mito Stress Test kit and correlated to gene expression levels. Of the 169 genes analyzed, 25 were significantly differentially expressed (P < 0.05) including GPX4 (P = 5.70 x 10 -3 ). Expression of GPX4 remained significant in pairwise comparisons by dose for day 2 and qRT-PCR validation confirmed a dose-dependent decrease in GPX4 expression. Maximal respiration (r = -0.62; P = 0.031) and spare respiratory capacity (r = -0.67; P = 0.017) correlated with GPX4 expression in doxorubicin-treated iPSC-cardiomyocytes. GPX4 encodes a glutathione peroxidase that is responsible for protecting the cell against oxidative damage. Damage due to reactive oxygen species (ROS) is one of the established mechanisms of anthracycline-induced cardiac damage. Our findings underscore a role for mitochondrial function and ROS in the development of doxorubicin-induced cardiotoxicity and implicates GPX4 in this process. The assessment of doxorubicin-altered gene expression in iPSC-cardiomyocytes may provide insight into how impaired mitochondrial respiration leads to cardiotoxicity and heart failure in cancer survivors treated with doxorubicin and other anthracyclines.