Evaluation and investigation of the cardiotoxicity of the potential anti-cholangiocarcinoma drug lanatoside C

Objective Lanatoside C (Lan C), a cardiac glycoside, has demonstrated potent anti-cholangiocarcinoma activity. However, its potential cardiotoxicity at therapeutic doses remains poorly understood. This study aimed to systematically evaluate the cardiotoxicity of Lan C at cellular and organ levels and to elucidate its underlying molecular mechanisms. Methods Human AC16 cardiomyocytes (ATCC® CRL-3568™) were employed for in vitro studies, with untreated cells included as negative controls. Cytotoxic effects at anti-cholangiocarcinoma-relevant concentrations (50–200 nM) were evaluated using a combination of cell viability assays, transmission electron microscopy, flow cytometry for apoptosis and reactive oxygen species (ROS) detection, JC-1 staining to assess mitochondrial membrane potential, lactate dehydrogenase (LDH) release assays, and Western blot analysis of apoptotic proteins. Male BALB/c mice were administered either vehicle (PBS) or Lan C (80 mg/kg/day) by oral gavage for 28 consecutive days. Cardiac function was assessed by echocardiography, and myocardial architecture was examined through histopathological analysis. Results Lan C showed a dose-dependent inhibitory effect in vitro on AC16 cell viability and induced apoptosis. This process involved decreased mitochondrial membrane potential, increased reactive oxygen species (ROS), downregulation of the anti-apoptotic protein Bcl-2, and upregulation of cytochrome C, Cleaved Caspase-9, and Cleaved Caspase-3, indicating activation of the mitochondrial-dependent apoptotic pathway. In vivo , high-dose Lan C significantly impaired left ventricular systolic function, as indicated by reductions in LVEF and LVFS, and induced myocardial structural alterations. Conclusion Lan C exhibits measurable but relatively low cardiotoxicity at concentrations effective against cholangiocarcinoma, with cardiac effects may be manageable at therapeutic doses. These findings support Lan C as a promising anti-tumor candidate with controllable cardiotoxicity under optimized dosing and clinical monitoring.