A series of novel self-assembled hyaluronic acid derivatives (HA-C18) grafted with hydrophobic octadecyl moiety and further dual targeting folic acid-conjugated HA-C18 (FA-HA-C18) were synthesized. With the increase in the degree of substitution of octadecyl group from 12.7% to 19.3%, the critical micellar concentration of HA-C18 copolymers decreased from 37.3 to 10.0 μg/mL. Paclitaxel (PTX) was successfully encapsulated into the hydrophobic cores of the HA-C18 and FA-HA-C18 micelles, with encapsulation efficiency as high as 97.3%. The physicochemical properties of the polymeric micelles were measured by DLS, TEM and DSC. Moreover, in vitro release behavior of PTX was investigated by dialysis bag method and PTX was released from micelles in a near zero-order sustained manner. In vitro antitumor activity tests suggested PTX-loaded HA-C18 and FA-HA-C18 micelles exhibited significantly higher cytotoxic activity against MCF-7 and A549 cells compared to Taxol at a lower PTX concentration. The cellular uptake experiments were conducted by quantitative assay of PTX cellular accumulation and confocal laser scanning microscopy imaging of coumarin-6 labeled HA-C18 and FA-HA-C18 micelles in folate receptor overexpressing MCF-7 cells. Folate and CD44 receptor competitive inhibition studies performed by fluorescence microscopy imaging suggested intracellular delivery of HA-C18 and FA-HA-C18 micelles were efficiently taken up via CD44 receptor-mediated endocytosis. The folate receptor-mediated endocytosis further enhanced internalized amounts of FA-HA-C18 micelles in MCF-7 cells, as compared with HA-C18 micelles. The internalization pathways of PTX-loaded HA-C18 and FA-HA-C18 micelles might include clathrin-mediated endocytosis, caveolae-mediated endocytosis and macropinocytosis. Therefore, the present study suggested that HA-C18 and FA-HA-C18 copolymers as biodegradable, biocompatible and cell-specific targetable nanostructure carriers, are promising nanosystems for cellular and intracellular targeting delivery of hydrophobic anticancer drugs.