In this study, bio-responsive polymeric MoS nanocomposites were prepared for use as a drug carrier for cancer therapy. Herein, we report the synthesis and demonstrate the self-assembly of pluronic F127 (PF127) on a cystamine-glutathione-MoS (CYS-GSH-MoS) system, which can be used for GSH-triggered drug release under biological reducing conditions. The reduction-sensitive disulfide bond containing CYS was incorporated between the amphiphilic copolymer PF127 and GSH-MoS to achieve feasible drug release. Percent drug loading capacity and encapsulation efficiency were 51.3% and 56%, respectively. In addition, when the MoS-GSH-CYS-PF127 nanocomposite was incubated in a GSH environment, the morphology of the nanocomposite tended to change, ultimately leading to drug release. The drug-loaded PF127-CYS-GSH-MoS polymeric nanocomposites efficiently released 52% of their drug content after 72 h of incubation in a GSH reduction environment. The HeLa cells treated with DOX loaded MoS-GSH-CYS-PF127 showed 38% toxicity at drug concentration of 40 μg, which indicated that the successfully released of drug from carrier and caused the cell death. Further, fluorescence microscopy images of HeLa cells revealed the potential behavior of the MoS-GSH-CYS-PF12 nanocomposite during the 2- and 4 h incubation periods; the nanocomposite was only found in the cytoplasm of HeLa cells. Interestingly, after 6 h of incubation, the drug was slowly released from the nanocomposite and could enter the nucleus as confirmed by fluorescence imaging of HeLa cells. Altogether, our synthesized PF127-coated MoS nanocomposite could be effectively adopted in the near future as a GSH-sensitive drug carrier.