In this study, a series of poly(amino acid)s materials with amphiphilic properties were synthesized by ring-opening polymerization. In specific solvents, macromolecules can self-assemble to form rod-shape/cubic or spherical morphologies. Drug-loaded nanoparticles were formed with the model drug doxorubicin via physical embedding and chemical bonding, with most of them presenting regular spherical shapes with the particle size less than 200 nm, which meets the size requirements for sustained drug release. CCK-8 experiments proved that the obtained material was non-toxic to normal cells, and because of the presence of cholesterol in the structure with good biocompatibility, the uptake of drug-loaded nanoparticles by HeLa cells was facilitated. In vitro release experiments showed that the prepared nanoparticles exhibited dual responsiveness to pH and temperature, and the kinetics of the drug release was investigated.