Betulinic acid is a eukaryotic topoisomerase I inhibitor, with an IC50 of 5 μM, and prevents topoisomerase I-DNA interaction. Betulinic acid (10, 20, 40, 80, and 160 µM) significantly suppresses MDA-MB-231 cell viability in a time- and dose-dependent manner after treatment for 24 or 48 h. Betulinic acid (20, 40 µM) causes decrease in Bcl-2 expression of MDA-MB-231 cells. Betulinic acid also induces morphological changes of MDA-MB-231 cells at 20 µM, and leads to ultrastructure changes of MDA-MB-231 cells at 40 µM. Betulinic acid shows anti-HIV activities, with an EC50 of 1.4 µM in acutely infected H9 lymphocytes.