Hydrophilic Anticancer Drug-loaded RSF Nanospheres

Lymphatic chemotherapy utilizes particles as anticancer drug carriers to delivery drug to the targeted site by local interstitial administration. Drug-loaded particles are easy to be entrapped by lymph node tissues during physical filtration and thus achieve long time to release the drug towards cancer tissues.In this way, the side effects of anti-cancer drugs are predominantly reduced. Regenerated silk fibroin (RSF) from Bombyx mori. silk is one of the fibrous proteins that has been widely used in biomedical and pharmaceutical fields. RSF shows not only prominent mechanical properties but also unique advantages of low cost, biocompatibility, biodegradability and low inflammatory responses. We adopt RSF nanospheres to load two traditional anti-cancer drugs i.e. floxuridine (FUDR) and 5-fluorouracil (5-Fu) for a future application of lymphatic chemotherapy.As RSF shows intrinsic characteristics of hydrophobic and hydrophilic multi-blocks copolymer structure and conformational transition mechanism, we prepared the FUDR-loaded RSF nanospheres by adding a small amount of ethanol into the FUDR/RSF mixed aqueous solution and then freezing the mixture. The drug-loaded nanospheres were in a fine spherical shape and without any aggregation. The process was simple and did not involve any toxic organic solvents. We found the re-dispersed capability and the stability of the nanospheres increased after introducing mPEG-NH2 onto the surface of nanospheres via the electrostatic force.The controllable size (210-510 nm) of nanospheres and drug loading capacity were depended on the silk fibroin concentration and initial FUDR/RSF ratio. The released property of FUDR-loaded RSF nanospheres were enhanced with increasing the silk fibroin concentration but with little change while varying the initial FUDR/RSF ratio. When the RSF concentration was set to 40 mg/mL, the maximum drug loading (6.8%) was obtained and the release time of such a kind of nanospheres was about 2 days. The in vitro cell experiment showed that the viability of both normal cells (293T) and cancer cells (Hela cells) was over 80% even incubated with pristine RSF nanospheres with high concentration (1.0 mg/mL), demonstrating the cytotoxicity of RSF nanospheres themselves was quite small. The FITC-labled RSF nanospheres were found to be able to adhere to Hela cells easily by confocal miroscope observation. The FUDR loaded in the RSF nanospheres exhibits the similar curative effect to kill or inhibit Hela cells to free FUDR.Due to the severe side effects of 5-Fu in conventional intravenous chemotherapy, FUDR is adopted to replace 5-Fu in some clinical cases. However, if we can reduce the side effects of 5-Fu by using drug carrier, the curative effect will be more direct since avoiding the complicated conversion of FUDR to 5-Fu in vivo, thus making better use of this traditional and cheap drug. Similarly, we applied the above self-assembly method and the 5-Fu-loaded RSF nanospheres randing from 290-460 nm were produced by changing silk fibroin concentration and initial 5-Fu/RSF ratio. The maxmuim drug loading was over 8% and the release time was over 2 days. The 5-Fu-loaded RSF nanospheres showed curative effect to the Hela cells, which was the same as the FUDR-loaded RSF nanospheres mentioned in the earlier part.All these results imply that such a kind of biomacromolecule based anti-cancer drug nanocarrier, which has suitable size and efficient curative effect to cancer cells, has great potential in lymphatic chemotherapy in clinical applications.