Silk Fibroin Nanoparticle Functionalization with Arg-Gly-Asp Cyclopentapeptide Promotes Active Targeting for Tumor Site-Specific Delivery

SIMPLE SUMMARY: Many tumor cell types overexpress integrins, a glycoprotein, on their cell membranes. The tripeptide motif Arg-Gly-Asp (RGD) is well-known for being recognized by the integrin superfamily members and can thus be used to actively target nanoparticles containing cytotoxic drugs directly to the tumor cells. According to this strategy, the antitumor activity is boosted, and healthy organs are spared. In this paper, silk fibroin, a naturally derived protein, has been used to prepare nanoparticles (SFNs) functionalized on their surface with RGD. In vitro experiments revealed that functionalization of SFNs with RGD provided active internalization by tumor cells overexpressing integrin receptors. Therefore, RGD-SFNs may be used for tumor-specific delivery of anticancer drugs. ABSTRACT: Arg-Gly-Asp (RGD)-based cyclopentapeptides (cRGDs) have a high affinity towards integrin αvβ3 and αvβ5, which are overexpressed by many tumor cells. Here, curcumin-loaded silk fibroin nanoparticles (SFNs) have been functionalized on the surface with cRGD to provide active targeting towards tumor cells; a “click reaction” between the RGD-based cyclopentapeptide carrying an azide group and triple-bond-functionalized nanoparticles has been exploited. Both naked and functionalized SFNs were less than 200 nm in diameter and showed a round-shaped morphology but, after functionalization, SFNs increased in size and protein molecular weight. The functionalization of SFNs’ surfaces with cRGD provided active internalization by cells overexpressing integrin receptors. At the lowest concentration tested (0.01 mg/mL), functionalized SFNs showed more effective uptake with respect to the naked by tumor cells that overexpress integrin receptors (but not for non-overexpressing ones). In contrast, at higher concentrations, the non-specific cell membrane protein–particle interactions are promoted and coupled to specific and target mediated uptake. Visual observations by fluorescence microscopy suggested that SFNs bind to integrin receptors on the cell surface and are then internalized by endocytosis. Overall, SFN functionalization provided in vitro active targeting for site-specific delivery of anticancer drugs, boosting activity and sparing healthy organs.