Nanodelivery System Targeting Fibroblast Activating Protein For Combined Treatment Of Oral Leukoplakia

Objective:Oral leukoplakia(OLK)is one of the most common oral potentially malignant disorders(OPMDs)in the oral mucosa,with a malignancy rate of 0.13-34.0%.The currently existing therapeutic approaches are somewhat flawed.A new approach to the treatment of OLK can be provided by combining pharmacotherapy and photothermal therapy(PTT)using highly expressed receptors as targets.Fibroblast Activation Protein(FAP)has been shown to be highly expressed in OLK.Therefore,the aim of this study was to prepare CXB@NGO-PEG-FAP targeting peptide,a nano-drug delivery system for OLK,and to investigate its therapeutic feasibility through in vitro and in vivo experiments.Methods:1.Verification of FAP high expression in oral leukoplakia and DOK by immunohistochemistry experiments and cellular immunochemical experiments.2.The nano-drug delivery system CXB@NGO-PEG-FAP targeting peptide was constructed and the prepared materials were characterized by UV spectrophotometer,Fourier transform infrared spectrometer,transmission electron microscope and dynamic light scattering instrument.3.To study the biocompatibility of NGO,NGO-PEG,NGO-PEG-FAP targeting peptides and CXB@NGO-PEG-FAP targeting peptides and to observe their stability in aqueous and FBS-containing solutions.4.The drug loading rate of nanoprobe NGO-PEG-FAP targeted peptide was studied,and the release of drug CXB from CXB@NGO-PEG-FAP targeted peptide in PBS solution at different pH was observed.5.Study the photothermal properties of different materials6.Toxicity and targeting of materials are investigated through in vitro experiments.The experimental group was DOK cells,and HOK was the control group for cytotoxicity test and cell uptake test.7.In vivo verification of the targeting,efficacy and biological safety of nano-drug delivery system on oral leukoplakia.Results:1.Immunohistochemistry experiments verified FAP high expression in OLK,and cellular immunochemical experiments verified FAP high expression in DOK cells.2.The successful preparation of a nanodrug delivery system was demonstrated by the utilization of UV spectrophotometer,Fourier infrared measuring instrument,transmission electron microscopy,zeta potential,and dynamic light scattering analyzer.Fluorescence quenching and recovery experiments show that when the NGO concentration is 0.1mg/ml,the fluorescence of the targeted peptide loading band will be completely quenched,and after encountering FAP,the FAP-targeted peptide can be specifically cleaved by FAP,and part of it will fall off from the CXB@NGO-PEG-FAP targeted peptide,and the fluorescence will be restored and play a targeted role.3.The stability test results show that the stability and biocompatibility of NGOs have been significantly improved after being modified by PEG.4.At a pH of 5.5,the experiments on drug release revealed that the rate and total amount of drug released were greater than those of the other two groups..5.Photothermal performance test studies have shown that the nano-drug carrier system has good photothermal performance and can be used for treatment.6.The results of cytotoxicity experiments showed that NGO-PEG was basically non-toxic.Cell viability in the CXB@NGO-PEG-FAP targeting peptide group was augmented when CXB concentration was equal,yet when the medium’s pH dropped,the drug was released and cell viability decreased.This shows that the CXB@NGO-PEG-FAP targeting peptide can kill DOK cells well under low pH conditions.The results of cell uptake experiments showed that the nano-drug delivery system can observe fluorescence signals in DOK cells,but not in HOK cells.7.The mouse model of oral leukoplakia was successfully constructed by4 NQO drinking method,and the in vivo experimental results showed that the nano-drug carrier system could target OLK well,and had good efficacy on oral leukoplakia,as well as good biological safety.Conclusion:1.This experiment successfully verified the high expression of FAP in OLK and DOK.2.In this experiment,a nanodrug-loading system was successfully constructed CXB@NGO-PEG-FAP targeted peptide.3.pH response experiments and cytotoxicity experiments confirmed that CXB@NGO-PEG-FAP targeted peptide has inhibitory effect on DOK cells and can reduce the toxic side effects on normal groups.4.In vitro uptake experiments confirmed that CXB@NGO-PEG-FAP-targeted peptides can target DOK cells.5.In vivo experiments have confirmed that CXB@NGO-PEG-FAP targeted peptide can target OLK well and has a good effect on OLK.