Objective To prepare a pH-sensitive dual-targeted doxorubicin-loaded nanoparticle and evaluate its antitumor activity in vitro.Methods The folatc and cRGD were conjugated to the nanoparticle using heparin as backbone.Doxorubicin was loaded in the nanoparticle through hydrazine linkage.The size and zeta potential of the nanoparticle were detected by Dynamic Light Scattering (DLS), while the morphology was visualized by Transmission electron microscope (TEM) and drug loading efficiency was evaluated using UV-spectrophotometry.The release of doxorubicin from nanoparticle was estimated by in vitro release experiment in different pH environment.Cellular uptake of the nanoparticle was evaluated by confocal microscopy and flow cytometry.And the cytotoxicity was evaluated by MTT experiment in MCF-7 breast cancer cells.Results The size of the nanoparticle was (81± 14.5)nm with PDI of 0.12, while the zeta potential was about-28.3 mV.It presents uniform spherical morphology visualized by TEM and the drug loading content was 8.9%.The release rate of doxorubicin was 80% at pH5.0, which was significantly higher than that of 80% at pH 7.4.The pH-insensitive dual-targeted doxorubicin-loaded nanoparticle was mainly visualized in cytoplasm while the pH-sensitive dual-targeted doxorubicin-loaded nanoparticle could efficiently penetrate into the nuclear.It was also confirmed by flow cytometry (P<0.05).The cytotoxicity experiment showed that the anticancer ability of pH-sensitive dual-targeted doxorubicin-loaded nanoparticle was significantly higher than that of pH-insensitive dual-targeted doxorubicin-loaded nanoparticle and free doxorubicin (P<0.05) . Conclusion A pH-sensitive dual-targeted doxorubicin-loaded nanoparticle was successfully synthesized.It exhibited the characteristic of pH-responsive and efficient cellular uptake and cytotoxicity to MCF-7 cells.