Assessment of markers of cardiac toxicity following combined treatment of cardiomyocytes with epirubicin and trastuzumab

Background Advances in cancer therapy have significantly improved long term survival rates of those with breast cancer. Concurrently, chemotherapy-induced cardiac dysfunction is receiving increased attention in breast cancer survivors. Reduced left ventricular systolic function related to cancer therapies has recently been designated cancer therapeutics-related cardiac dysfunction (CTRCD). The addition of Trastuzumab to Epirubicin treatment has been shown to slow progression of disease, reduce mortality and extend duration of survival in patients with HER2+ Breast Cancer. However, both drugs have off target cardiotoxic effects. Currently echocardiography is the standard diagnostic test for detecting left ventricular systolic dysfunction in patients receiving these agents, but often becomes abnormal only when significant irreversible cardiac damage has occurred. There is an emerging need for blood-based biomarkers to aid in diagnosing subclinical cardiac dysfunction and to stratify those at risk prior to therapy. Methods AC16 cardiomyocytes were treated with Epirubicin (26ug/ml) and Trastuzumab (150ug/ml), both together and in monotherapy, over 10hr and 26hr studies. Cell viability was assessed via MTT cell viability assay. Protein and gene expression of Troponin I and BNP were assessed via western blot analysis and RT-PCR. Western blot analysis and fluorescent microscopy staining of oxidative stress markers was also carried out to investigate potential mechanisms of cardiac damage. Results Morphological changes occurred in all cells treated over 26hrs, particularly with combined treatment. Cell viability decreased significantly compared to control in the combined treatment group over 26hrs (p 0.0126*). Troponin I expression also increased significantly in the combined treatment group compared to monotreatment with Epirubicin for the same timepoint (p 0.015). BNP levels significantly increased in cells treated with a combination of Epirubicin and Trastuzumab compared to Trastuzumab alone (p=0.023). Conclusion Combined treatment with Epirubicin and Trastuzumab produces significantly more cell damage than monotreatment with either therapy. Troponin I and BNP are biomarkers that should be explored as a diagnostic tool for prediction of subclinical CTRCD.