Doxurubicin (Adriamycin, Dox) is an anthracycline antineoplastic agent that is commonly used in the treatment of neoplasms, including breast cancer, leukemias and lymphomas. Its clinical use is limited due to the development of a cumulative dose-dependent irreversible heart muscle disease similar to congestive cardiomyopathy of a variety of etiologies. The histopathological findings of Dox-treated myocytes generally include a characteristic picture of vacuolar degeneration of the sarcoplasmic reticulum, swelling of cardiac mitochondria with disorganization of the cristae, interstitial edema, and disruption and loss of myofibrils. Alteration of the Z-band structure and disarray of the thin filaments are observed with Dox treatment in vivo and in vitro. Although the precise subcellular mechanism of Dox-induced cardiotoxicity is not defined, some of the pathogenic mechanisms include: DNA binding leading to impaired RNA and protein synthesis, free radical formation resulting in lipid peroxidation and membrane damage, and mitochondrial impairment. Previously, we showed that Dox selectively inhibits muscle-specific gene expression in the myocardium both in vivo and in vitro. One of the interesting aspects of those observations was that Dox induces the expression of the Id (inhibitor of differentiation) gene, a negative regulator of myogenesis, whereas overexpression of E2-5, a natural molecular antagonist of E2-5, can prevent the inhibition of the expression of muscle-specific genes by Dox. Id is overexpressed in myocardiocytes and myogenic cells. The mechanisms involved appear to be by activation of a kinase pathway. The experiments proposed are designed to test the hypothesis that doxorubicin acts, in part, by specific effects on the transcriptional regulatory machinery of tissue-specific genes. The studies proposed involve molecular genetic analysis of events related to Dox effects in cells in culture, in heart muscle in vivo and in transgenic animals. Some of these experiments are of relatively high risk but have the possibility of providing major understanding of critical mechanisms involved in Dox induced cardiomyopathy and leading to methods that could ameliorate or prevent it. Furthermore, the apparent ability of this anthracycline drug to selectively inhibit molecular genetic events related to tissue specificity also raises the general possibility that pharmacological agents could be used to regulate and control gene transcription in highly selective ways. A long term objective of this proposal is to develop rational therapeutic approaches to this cardiomyopathy, by taking advantage of the knowledge gained from the study of cultured heart cells exposed to Dox. These studies may lead to prevention or reduction of this serious side effect of Dox. To this end, it is essential to understand the precise molecular mechanisms by which Dox causes cardiomyopathy.
