The past few years have witnessed major developments in nanotechnology with great potential in powering new therapeutic tools for cancer management. Our goal in this study was to develop a biocompatible nanoshuttle for the efficient delivery of 5FU in colorectal cancer patients. Silk fibroin/Poly(ethylene glycol) nanoparticles (SF/PEG NPs) were obtained and further loaded with 5FU. These nanoshuttles were characterized in terms of: morphological properties, size and size-distribution, drug uptake and release potential as well as in vitro cytotoxicity potential screening. The SF/PEG + 5FU NPs cytotoxicity was determined on HT-29 cells after determination of the lethal dose 50 and targeted the evaluation of the cells viability, proliferation potential and migration and invasion potential. The inflammatory profile of RAW 264.7 macrophage cells was also detrmined by flowcytometry. The basic cytotoxicity screening revealed that the pristine SF/PEG NPs displayed a good biocompatibility while the 5FU loaded NPs induced cytotoxic effects on HT-29 cells. More, the 5FU loaded SF/PEG NPs significantly reduced the migration and invasion processes as compared with the unloaded NPs. Lastly, we observed that the cytokines inflammatory profile was significantly altered after the treatment with the 5FU loaded SF/PEG NPs as compared with the unloaded nanoshuttles.