Effects of Doxorubicin (Adriamycin) and [(+)-1,2-bis(3,5-dioxopiperazinyl-1-yl)]propane (ICRF-187) on skeletal muscle protease activities.

Abstract

Adverse effects of doxorubicin (adriamycin) have been reported to be due to iron-catalyzed free radical formation, which can be prevented with the cytoprotective chelating agent [(+)-1,2-bis(3,5-dioxopiperazinyl-1-yl)]propane (dexrazoxane; ICRF-187). Affected tissues include the heart, gastrointestinal tract, and kidney. However, there is very little information on the effects of adriamycin on skeletal muscle, despite the fact that there is direct and indirect evidence to show that both adriamycin and ICRF-187 are myotoxic. To investigate the mechanisms of cytotoxicity of these agents in skeletal muscle, we have conducted a systematic investigation of the activities of the major lysosomal (dipeptidyl aminopeptidase I and II and cathepsins B, D, H, and L) and cytoplasmic (alanyl-, arginyl-, and leucyl aminopeptidase, dipeptidyl aminopeptidase IV, tripeptidyl aminopeptidase, and proline endopeptidase) muscle proteases. These enzymes play an important role in normal cellular function and represent potential targets for toxic and protective agents. Male Wistar rats (approx. 0.2 kg) were subjected to a pretreatment phase of 30 min followed by a treatment stage of either 2.5 or 24 h. The pretreatment involved injection of a single bolus of either saline (0.15 mol/l NaCl; 5 ml/kg ip) or ICRF-187 (100 mg/kg; 5 ml/kg ip). After 30 min, rats were injected again with a single bolus of either adriamycin (5 mg/kg; 10 ml/kg ip) or saline (0.15 mol/l NaCl; 10 ml/kg ip) in the treatment phase. At either 2.5 or 24 h after the last adriamycin or saline injection, rats were killed for subsequent dissection of the gastrocnemius muscle for analysis. In the 2.5-h study, there were significant reductions in cathepsin D activities of adriamycin-treated rats compared to saline injected control (p = 0.02). In both 2.5- and 24-h studies there were also significant differences (p = 0.05) in cathepsin H activities between rats treated with adriamycin and ICRF-187, although these differences were not significant when data were compared with corresponding saline-injected rats. There were no other overt effects for any of the other proteases at either 2.5 or 24 h. We conclude that both adriamycin and ICRF-187 have very little effect on the activities of muscle proteases and that altered proteolysis is not involved in the reported pathological reactions induced by these agents.