Preparation Of A Doxorubicin-loaded Small Molecular Hydrogel And An Anticancer Study In Vitro

The doxorubicin is an anti-cancer drug widely used to treat liver cancer、cervical cancer and stomach cancer. Nanocarriers such as polymer micelles、polymer nanoparticles and liposome have been used to increase its water solubility and reduce its side effects to the heart.Supramolecular hydrogels are formed by the self-assembly of small molecules(hydrogelators) that are biocompatible and easily degradable. They are promising soft materials for three dimensional(3D) cell culture, controlled delivery of therapeutic agents and proteins, and regenerative medicine. This paper aimed to increase the water solubility 、 loading efficiencies and anti-cancer activity of doxorubicin by preparing doxorubicin-loaded small molecular hydrogels which may as new durg delivery system being widely used.Methods 1.Synthesis and characterize of small molecular peptide: solid phase peptide synthesis was used to obtain negative charged peptide:Nap GFFYGRGD; RP-HPLC was used to purify the small molecular peptide; LC-MS and H NMR were used to confirm the structure of the small molecular peptide.2.Preparation and characterize of a doxorubicin-loaded small molecular hydrogels: heat-cooling method was used to prepare a doxorubicin-loaded small molecular hydrogel; rheology test was performed to characterize the mechanical properties of resulting gels with different amounts of doxorubicin;Transmission electron microscopy(TEM) was then used to characterize the morphology of self-assembled nanostructures in the peptide solution and the doxorubicin-loaded small molecular hydrogels; Zeta potential was performed to characterize the influence of doxorubicin on peptide nanofibers; the release behaviour of doxorubicin in vitro was performed to characterize the effect of mechanical properties for release rate.3. Anti-cancer study in vitro: MTT cell viability test was used to determine the anti-cancer cells activities of doxorubicin-loaded peptide nanofibers 、 free doxorubicin and peptide nanofibers for pancreatic cancer( Pac-2)、cervical cancer(Hela) and liver cancer(MCF-7).Results 1. The crude product yield of Nap GFFYGRGD was up to 90% by using a solid phase peptide synthesis and the purity of Nap GFFYGRGD was up to 95% after RP-HPLC.2. Doxorubicin-loaded small molecular hydrogels containing different amounts of doxorubicin were obtained by heat-cooling method; rheology results indicated that the mechanical properties of doxorubicin-loaded small molecular hydrogels were stronger when the amount of doxorubicin was higher; TEM results demonstrated that the addition of doxorubicin to the nanofiber solution led to the growth of nanospheres at the surface of nanofibers which served as the cross-linkers to increase the inter-fiber interactions, leading to the formation of stable hydrogels; release profile of doxorubicin suggested that higher percentages of doxorubicin got released from gels with less amounts of doxorubicin, suggesting that doxorubicin would release more rapidly from mechanically weak gels.3. Results of MTT assay suggested that the cytotoxicity of free doxorubicin for pancreatic cancer( Pac-2)、cervical cancer(Hela) and liver cancer(MCF-7) was improved by using doxorubicin-loading peptide nanofibers, especially for pancreatic cancer( Pac-2) whose IC50 value being reduced dramatically.Conclusion 1. Doxorubicin-loaded small molecular hydrogels could be formed by the positive-negative charge interaction between peptide and doxorubicin,which greatly improved the solubility and loading efficiency of doxorubicin.2. Small molecular peptide we designed and synthesized possessed better biocompability, and the anti-cancer effect of doxorubicin could be improved by using doxorubicin-loaded peptide nanofibers which was expected to become a kind of smart and targeted carrier for doxorubicin.