The clinical application of gambogic acid, a natural component with promising antitumor activity, is limited due to its extremely poor aqueous solubility, short half-life in blood, and severe systemic toxicity. To solve these problems, an amphiphilic polymer-drug conjugate was prepared by attachment of low molecular weight (ie, 2 kDa) methoxy poly(ethylene glycol) methyl ether (mPEG) to gambogic acid (GA-mPEG₂₀₀₀) through an ester linkage and characterized by (1)H nuclear magnetic resonance. The GA-mPEG₂₀₀₀ conjugates self-assembled to form nanosized micelles, with mean diameters of less than 50 nm, and a very narrow particle size distribution. The properties of the GA-mPEG₂₀₀₀ micelles, including morphology, stability, molecular modeling, and drug release profile, were evaluated. MTT (3-(4,5-dimethylthiazo l-2-yl)-2,5 diphenyl tetrazolium bromide) tests demonstrated that the GA-mPEG₂₀₀₀ micelle formulation had obvious cytotoxicity to tumor cells and human umbilical vein endothelial cells. Further, GA-mPEG₂₀₀₀ micelles were effective in inhibiting tumor growth and prolonged survival in subcutaneous B16-F10 and C26 tumor models. Our findings suggest that GA-mPEG₂₀₀₀ micelles may have promising applications in tumor therapy.