Gambogic acid (GA), a natural product, exhibits high potency in inhibiting cancer cell growth through the effective induction of apoptosis. In order to investigate the structure-activity relationships of GA derivatives, 11 oxidized derivatives of GA were synthesized. Some of them showed strong inhibitory effects on HT-29, Bel-7402, BGC-823, A549, and SKOV 3 cell lines. Moreover, in this paper the cellular growth inhibitor 39-hydroxy-6-methoxy-gambogic acid methyl ester (10) was identified as a HepG2 cell apoptosis inhibitor through Annexin-V/PI double staining assay and the expression of the related apoptotic proteins (Bax and Bcl-2). Compound 10 may serve as a potential lead compound for the development of new anticancer drugs. Further SAR studies of GA derivatives indicated that modification of carbon-carbon double bond at C-32/33 or C-37/38 and of the methyl groups at C-39/C-35 can improve antitumor activity.