The anthracycline doxorubicin (Dox) is an effective antitumor agent. However, its use is limited because of its toxicity in the heart. N-Benzyladriamycin-14-valerate (AD 198) is a modified anthracycline with antitumor efficacy similar to that of Dox, but with significantly less cardiotoxicity and potentially cardioprotective elements. In the present study, we investigated the possibility of in vivo protective effects of low-dose AD 198 against Dox-induced cardiomyopathy. To do this, rats were divided into four groups: vehicle, Dox (20 mg/kg; single injection day 1), AD 198 (0.3 mg/kg per injection; injections on days 1, 2, and 3), or a combination treatment of Dox + AD 198. Seventy-two hours after beginning treatment, hearts from the Dox group had decreased phosphorylation of AMP kinase and troponin I and reduced poly(ADP-ribose) polymerase, β-tubulin, and serum albumin expression. Dox also increased the phosphorylation of phospholamban and expression of inducible nitric-oxide synthase in hearts. Each of these Dox-induced molecular changes was attenuated in the Dox + AD 198 group. In addition, excised hearts from rats treated with Dox had a 25% decrease in left ventricular developed pressure (LVDP) and a higher than normal increase in LVDP when perfused with a high extracellular Ca(2+) solution. The Dox-induced decrease in baseline LVDP and hyper-responsiveness to [Ca(2+)] was not observed in hearts from the Dox + AD 198 group. Thus Dox, with well established and efficient antitumor protocols, in combination with low levels of AD 198, to counter anthracycline cardiotoxicity, may be a promising next step in chemotherapy.