Synergistic anticancer effects of doxorubicin in combination with tilorone in breast cancer

Abstract Background Tilorone is an orally active antiviral interferon inducer with anticancer effects. Doxorubicin has been previously reported to exhibit synergism with type 1 interferons. The present study investigated the anticancer effects of doxorubicin in combination with tilorone in breast cancer. Methods Effects of different concentrations of tilorone (5μM–80μM) and doxorubicin (0.03μM–32μM) on the viability of breast cancer cells (MDA-MB-231 and MDA-MB-468) were measured by MTT assay. For combination studies, drugs were combined in equipotent ratios. CompuSyn analysis was performed to determine the synergistic combinations (CI < 1). Colony formation and Western blot assays were performed to measure proliferation and apoptosis. In vivo efficacy of the combination was tested in the DMBA-induced breast cancer model. After 3 weeks of treatment, tumors were excised for tumor-specific and immunohistochemical analysis. CompuSyn analysis was applied to assess in vivo drug interactions. Results Combination of equipotent doses of tilorone and doxorubicin synergistically (CI < 1) caused significant reduction in cell viability with an increase in the pro-apoptotic proteins at the combination of IC50/2, IC50, and, 2 × IC50 dose combinations of doxorubicin and tilorone in comparison to single drug treatments. The combination of doxorubicin and tilorone also exhibited synergism in DMBA-induced breast cancer at all tested dose combinations. Tumor tissue showed increased expression of IFN-β, caspase-3, P53, and E-cadherin with decreased N-cadherin expression in treatment groups, with more prominent changes observed in combination groups. Medium dose combination of tilorone and doxorubicin (T2D2) most effectively inhibited tumor progression with reduced hepatic and renal toxicities. Conclusion Doxorubicin in combination with tilorone exerts synergistic effects against breast cancer with reduced side effects. The combination can be further explored in clinical settings for effective management of breast cancer with reduced side effects caused by doxorubicin chemotherapy. © 2025 Elsevier B.V.