A conjugate of the antitumor drug, methotrexate (MTX), with a diblock methoxypoly(ethylene glycol)–poly(caprolactone) (mPEG–PCL) copolymer was synthesized by the reaction of the mPEG–PCL copolymer with MTX in the presence of dicyclohexylcarbodiimide and dimethylaminopyridine. The conjugated copolymer was characterized in vitro using ¹H NMR, FTIR, DSC and GPC techniques. Then, the MTX conjugated mPEG–PCL was self-assembled into micelles in aqueous solution. The resulting micelles were characterized further by various techniques such as dynamic light scattering (DLS) and atomic force microscopy (AFM). The cytotoxicity of the MTX, mPEG–PCL copolymer and MTX-conjugated mPEG–PCL micelles were compared to each other by the MTT assay of the treated MCF-7 cell lines. The results revealed that the micelles formed by the MTX-conjugated mPEG–PCL have a spherical structure with an average size of 72.31 nm. The release behavior of the conjugated micelles was studied in different media. The in vitro release study showed that the release rate of the micelles consisting of the conjugated copolymer was dependent on the pH as it was higher at lower pH than under neutral conditions. An in vitro cytotoxicity assay showed that the mPEG–PCL copolymer did not affect the growth of MCF-7 cells. The cytotoxic activity of the MTX-conjugated mPEG–PCL micelles against MCF-7 cells was greater than that of free MTX. The kinetics of drug release from methotrexate-conjugated micelles under different values of pH was also investigated by different kinetic models such as zero order, first-order, Hill equation, Hixson–Crowell, Korsmeyer–Peppas, and Weibull.