Astragalus membranaceus (AM) herb is a component of traditional Chinese medicine used to treat various cancers. Herein, we demonstrate a strong anti-leukemic effect of AM injected (Ai) into the mouse model of erythroleukemia induced by Friend virus. Chemical analysis combined with mass spectrometry of AM/Ai identified the compounds Betulinic acid, Kaempferol, Hederagenin, and formononetin, all major mediators of leukemia inhibition in culture and in vivo. Docking analysis demonstrated binding of these four compounds to FLI1, resulting in downregulation of its targets, induction of apoptosis, differentiation, and suppression of cell proliferation. Chemical composition analysis identified other compounds previously known having anti-tumor activity independent of the FLI1 blockade. Among these, Astragaloside-A (As-A) has marginal effect on cells in culture, but strongly inhibits leukemogenesis in vivo, likely through improvement of anti-tumor immunity. Indeed, both IDO1 and TDO2 were identified as targets of As-A, leading to suppression of tryptophane-mediated Kyn production and leukemia suppression. Moreover, As-A interacts with histamine decarboxylase (HDC), leading to suppression of anti-inflammatory genes TNF, IL1B/IL1A, TNFAIP3, and CXCR2, but not IL6. These results implicate HDC as a novel immune checkpoint mediator, induced in the tumor microenvironment to promote leukemia. Functional analysis of AM components may allow development of combination therapy with optimal anti-leukemia effect.