Synthesis And Properties Of Biotin-modified Galactosylated Chitosan Nanomaterials

Liver cancer is a kind of tumor that occurs in the liver.It has a high incidence and great harm,particularly among the Chinese population.At present,most chemotherapeutic drugs used in the treatment of liver cancer have poor targeting,resulting in low bioavailability.Also,these drugs have been associated with some serious side effects.Therefore,it is urgent to explore new effective chemotherapeutic drugs for liver cancer.As a drug carrier,chitosan（CS）nanomaterials offer slow drug release,biodegradability,and surface modification.In order to improve the targeting of liver cancer,in previous studies,galactose was grafted onto CS to synthesize galactosylated chitosan（GC）,and GC/5-FU nanoparticles were synthesized with 5-fluorouracil（5-FU）.After treating the orthotopic liver cancer transplantation model with GC/5-FU nanoparticles,the survival time of mice was significantly prolonged,but the concentration of 5-FU in normal liver tissue was still high;thus,there is still room for improvement in the targeting of liver cancer.Therefore,how to modify GC nanomaterials,further improving the targeting efficiency of drugs to liver cancer cells and reducing the enrichment of drugs in normal liver tissue is the key to increase the bioavailability of drugs and improve the therapeutic effect.Considering the high expression of the biotin（Bio）receptor on liver cancer cells,Bio was grafted into the GC system to synthesize the galactosylated chitosan modified by biotin（Bio-GC）to optimize the liver cancer-targeting efficiency of the GC system.Bio-GC was compounded with 5-FU and DNA to form corresponding nanoparticles.The liver cancer-targeting ability of Bio-GC nanoparticles was measured at the cellular and animal levels.The therapeutic effect of Bio-GC/5-FU nanoparticles on liver cancer was explored using an orthotopic liver cancer transplantation model in mice,and its mechanism was further discussed.The main research contents and results were as follows:（1）Synthesis and characterization of Bio-GC materials.Bio-GC nanomaterials were synthesized by orthogonal experiment and verified by infrared spectrum and nuclear magnetic resonance hydrogen spectrum.Bio-GC/5-FU and Bio-GC/plasmid DNA nanoparticles were synthesized by the ion coagulation method.Transmission electron microscopy showed that the synthesized materials had a smooth surface,good dispersion,and a good sustained-release effect.At the same time,Bio-GC nanomaterials had a protective effect on DNA,and the synthetic materials had good biosafety.（2）The hepatoma targeting of Bio-GC nanomaterials was verified at the cellular level.Cell proliferation and cell migration experiments were used to verify the effects of Bio-GC/5-FU nanoparticles on the proliferation,migration,and intracellular 5-FU concentration of hepatoma cells.The liver cancer-targeting ability of Bio-GC nanomaterials was verified by gene transfection and laser confocal imaging.The results showed that Bio-GC/5-FU nanoparticles inhibited the proliferation and migration of hepatoma cells in a time-and dose-dependent manner.Meanwhile,Bio-GC nanomaterials could significantly increase the concentration of 5-FU in hepatoma cells,improve the transfection efficiency of genes into hepatoma cells,and promote the endocytosis of hepatoma cells.Compared with GC,Bio-GC significantly improved liver cancer targeting.（3）The liver cancer targeting of Bio-GC nanomaterials was verified at the animal level.Small animal in vivo imaging was used to explore the dynamic distribution of Bio-GC and its drug compound system in the mice orthotopic liver cancer transplantation model.The results showed that compared with CS and GC groups,Bio-GC nanomaterials had the highest liver cancer/liver fluorescence intensity ratio.Also,the concentration of 5-FU in the Bio-GC/5-FU group was 6 times higher than that in the 5-FU group and 2.5 times higher than that in the GC/5-FU group.This data further proved that Bio-GC has better liver cancer targeting than GC.（4）The inhibition of Bio-GC/5-FU nanoparticles on liver cancer in vivo and its mechanism were evaluated in a mouse tumor-bearing model.After the tumor-bearing mouse model was treated with Bio-GC/5-FU,the morphology,apoptosis rate,and cell cycle of liver cancer cells were detected by scanning electron microscope and flow cytometry and so on.The results showed that Bio-GC/5-FU nanoparticles could significantly prolong the survival time and reduce the weight of the orthotopic liver cancer transplantation model in mice.The volume of hepatoma cells in tumor tissue decreased,nuclear pyknosis and apoptotic body formation were also observed.Cell cycle detection showed that the ratio of G₀/G₁ significantly decreased,and the apoptosis rate significantly increased.The expression of tumor apoptosis-related signal pathway genes was detected by RT-PCR and Western bolt,revealing the inhibitory mechanism of Bio-GC/5-FU on liver cancer,which was related to the apoptosis caused by the 5-FU activating BAD/Bcl-x L signal pathway.Compared with GC,Bio-GC can significantly enhance the tumor-killing effect of 5-FU and enhance the therapeutic ability of tumor-bearing mice.To sum up,by discussing the liver cancer targeting effect of Bio-GC and the tumor treatment ability after drug loading,and evaluating the application potential of Bio-GC in liver cancer treatment,these data provide new ideas for the design of new anti-tumor nano targeted drugs and new strategies for the treatment of liver cancer.