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Cassia fistula has a wide array of biologically active and therapeutically important class of compounds. Leishmania donovani important drug targets, sterol 24-c methyltransferase ( Ld SMT), trypanothione reductase ( Ld TR), pteridine reductase ( Ld PTR1), and nucleoside hydrolase ( Ld NH), were modelled, and molecular docking was performed against the abundant phytochemicals of its leaf extract. Molecular docking results provided the significant prima facie evidence of the leaf extract to have antileishmanial potential. To confirm this, we performed in vitro antileishmanial and cytotoxicity assays. Methanolic extract of C. fistula leaves showed growth inhibition and proliferation of L. donovani promastigote with an IC 50 value of 43.31 ± 4.202 μg/mL. It also inhibited the growth of intra-macrophagic amastigotes with an IC 50 value of 80.76 ± 3.626 μg/mL. C. fistula extract was found cytotoxic at a very high concentration on human macrophages (CC 50 = 626 ± 39 μg/mL). Annexin V/propidium iodide (PI) staining assay suggested partial apoptosis induction in parasites by C. fistula to exert its antileishmanial activity. Here, for the first time, we have shown the antileishmanial potential of C. fistula leaves. Overall, our results could open new insight for an affordable and natural antileishmanial with high efficacy and less toxicity., Competing Interests: The authors declare no competing financial interest., (© 2021 The Authors. Published by American Chemical Society.)