Peptide Self-assembly Systems For Drug Delivery

Self-assembling peptides have attracted extensive attention in recent years.They have wide applications in cell culture,drug release,tissue engineering and immobilization of enzymes.Self-assembled supramolecular nanostructures have proven to be good candidates for effective drug carriers.Based on this concept,we designed a series of peptides and studied their self-assembly behaviors,cytotoxicity and drug-loading and delivery properties both in vitro and in vivo.The main research contents and conclusions are as follows:（1）Four peptide molecules with different charges and compositions were designed and synthesized,which are Fmoc-Cha Cha-GK-NH₂（GK）,Fmoc-Cha Cha-GKK-NH₂（GKK）,Fmoc-Cha Cha-GD-NH₂（GD）,Fmoc-Cha Cha-GDD-NH₂（GDD）.GK and GKK are positively charged,while GD and GDD are negatively charged.The charge number is varied by changing the solution p H,thereby adjusting the self-assembly behavior.It was found that GK and GKK molecules gradually agglomerated with increasing p H.GK of 0.5 m M formed a helical structure at p H 3.0 and 7.0,while GKK formed nanofibers.While for GD and GDD,the p H decrease induced higher aggregation propensity.With further experiments to probe into the drug loading efficiencies,we found that GK exhibited the best drug-loading efficiency,which can load Nile Red,a model drug molecule,at an efficiency of 1.22%.However,GK gave relatively higher cytotoxicity.The survival ratios of 293E and Hepg2 cells were below 50%at a GK concentration of 50μM.GD and GDD gave relatively lower cytotoxicity.The survival ratios of all cells were above 80%at a concentration of 50μM.（2）Another four amphiphilic peptide molecules that respond to MMP7 were designed,which are Nap-IIGPLGLAGRRRR-NH₂（II-R₄）,Nap-IIIIGPLGLAGRRRR-NH₂（I₄-R₄）,Nap-IIIIIIGPLGLAGRRRR-NH₂（I₆-R₄）and Nap-FFGPLGLARRRR-NH₂（FF-R₄）,respectively.They contain a hydrophobic motif with multiple benzene rings for promoting self-assembly,a hydrophilic group of four repeated arginine residues for providing positive charges,and the-GPLGLA-fragment for MMP7-sensitive cleavage.The self-assembly results showed that,with increasing the number of isoleucine,the I_x-R₄ molecules gave a secondary structure change from random coil toβ-sheet.Simultaneously the self-assembled structures changed from fibers to short rods.For II-R₄ and FF-R₄,they have the same number of hydrophobic amino acids but different side chain groups.Their secondary structures wereβ-sheet,and their self-assembled structures were similar in morphology to be nanofibers.The ability of these structures to load Nile Red was also studied.I₆-R₄ had the highest drug-loading capacity with a loading ratio of～9.87%.However,due to its poor biocompatibility and toxicity to some cells,no further studies were conducted.This provides an important reference for the development of new drug carriers.（3）The MMP7-sensitive peptide Nap-FFGPLGLARKRK-NH₂（RKRK）was applied for animal experiments.RKRK self-assembled fibrils can efficiently load the drug molecule of doxorubicin（DOX）at a loading efficiency of 23.75%.Upon treatment with MMP7,the loaded DOX can be efficiently released at a ratio of 80%.The cytotoxicity experiments showed that the RKRK/DOX complexes had almost no cytotoxicity to the normal cells of COS7 and 293E.The cell viability remained above 75%at a RKRK/DOX concentration of 40μg/m L.However,the complexes showed higher cytotoxicity towards cancer cells.The survival ratios of Hela,Hepg2 and A549 dropped below 30%at a RKRK/DOX concentration of only 20μg/m L.Further,the RKRK/DOX complexes were applied for in vivo animal experiments.For the RKRK/DOX groups,the mice body weight remained to be about 16 g during the whole experimental course.The life length was also significantly increased.Moreover,comparing to the control groups the tumor growth of the RKRK/DOX groups was significantly inhibited.The results indicated that the RKRK/DOX complexes have low toxicity and can effectively inhibit the tumor growth to give excellent therapeutic efficiency.Further pathological study showed that the RKRK/DOX complexes can also effectively inhibit the tumor metastasis.By having low toxicity and high cancer killing efficiency,the RKRK/DOX complexes are highly promising for applications in tumor therapy.