PURPOSE: Doxorubicin (Dox) is clinically limited due to its dose-dependent cardiotoxicity. Andrographolide (Andro) has been confirmed to exert cardiovascular protective activities. This study aimed to investigate protective effects of Andro in Dox-induced cardiotoxicity (DIC). METHODS: The cardiotoxicity models were induced by Dox in vitro and in vivo. The viability and apoptosis of H9c2 cells and the myocardial function of c57BL/6 mice were accessed with and without Andro pretreatment. Network pharmacology and RNA-seq were employed to explore the mechanism of Andro in DIC. The protein levels of Bax, Bcl2, NLRP3, Caspase-1 p20, and IL-1β were qualified as well. RESULTS: In vitro, Dox facilitated the downregulation of cell viability and upregulation of cell apoptosis, after Andro pretreatment, the above symptoms were remarkably reversed. In vivo, Andro could alleviate Dox-induced cardiac dysfunction and apoptosis, manifesting elevation of LVPWs, LVPWd, EF% and FS%, suppression of CK, CK-MB, c-Tnl and LDH, and inhibition of TUNEL-positive cells. Using network pharmacology, we collected and visualized 108 co-targets of Andro and DIC, which were associated with apoptosis, PI3K-AKT signaling pathway, and others. RNA-seq identified 276 differentially expressed genes, which were enriched in response to oxidative stress, protein phosphorylation, and others. Both network pharmacology and RNA-seq analysis identified Tap1 and Timp1 as key targets of Andro in DIC. RT-QPCR validation confirmed that the mRNA levels of Tap1 and Timp1 were consistent with the sequenced results. Moreover, the high expression of NLRP3, Caspase-1 p20, and IL-1β in the Dox group was reduced by Andro. CONCLUSIONS: Andro could attenuate DIC through suppression of Tap1 and Timp1 and inhibition of NLRP3 inflammasome activation, serving as a promising cardioprotective drug.