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pH-sensitive poly(ethylene glycol)-poly(L-histidine)-poly(L-lactide) (PEG-PH-PLLA) nanoparticles were prepared and used as carriers for anti-tumor drug delivery. The morphology and properties of the nanoparticles such as pH sensitivity, zeta potential and mean diameters were investigated. The cytotoxicity of PEG-PH-PLLA nanoparticles was evaluated. Doxorubicin (DOX) was encapsulated in the nanoparticles to explore the release profile. The drug-loaded nanoparticles were incubated with HepG2 cells to study the in vitro anti-tumor effect. The results showed the sizes of both blank nanoparticles and drug-loaded nanoparticles in pH 7.4 were smaller than those of nanoparticles in pH 5.0, and the mean diameter of drug-loaded nanoparticles was much bigger than that of blank nanoparticles. The PEG-PH-PLLA nanoparticles were nontoxic to both NIH 3T3 fibroblasts and HepG2 cells. The release profile showed that the release of DOX in pH 5.0 was much faster than that in pH 7.4. The in vitro experiments demonstrated that the anti-tumor effect of drug-loaded nanoparticles was preferable to free doxorubicin. The pH-sensitive PEG-PH-PLLA nanoparticles are promising carriers for anti-tumor drug delivery.