| Chemotherapy is a main therapeutic strategy for the treatment of malignant tumors.However,many first-line anticancer drugs such as 5-fluorouracil(5FU)and doxorubicin(DOX)have poor tumor selectivity,resulting in insignificant treatment effects and serious toxic side effects.In recent years,with the continuous development and in-depth research of nano drug delivery systems,small molecular nanometer prodrugs with good biocompatibility have become the research hotspots of cancer therapy.The small molecule prodrug not only retains the advantages of traditional physical embedding nano drug carrier and polymer prodrug,but also has the advantages of clear structure,simple preparation,high drug loading and clear metabolism,so as to achieve precise regulation of drugs and enhance chemotherapy effect.However,at present,small molecule nanoprodrugs are still not clinically applied.The reason is that there is a physiological barrier at the tumor site,and the ideal drug concentration cannot be obtained at the tumor site only by the high permeation and long retention(EPR)effect of the tumor,and After being ingested by tumor cells,small molecule prodrugs cannot release drugs quickly in a short period of time,affecting the ability to kill tumor cells.In view of this,it is urgent to develop a novel small molecule prodrug nano-transport system,combined with the advantages of synergistic drug administration,significantly improve the drug's ability to target enrichment,cell uptake and intracellular controlled release of drugs at the tumor site.Due to the abnormal metabolism of tumor cells,the micro-environment of tumor tissue is different from normal tissue.Therefore,the tumor tissue-specific microenvironment can be utilized to achieve targeted enrichment and controllable delivery of the drug at the tumor site.The pH-sensitive drug delivery system specifically recognizes the tumor micro-acid environment due to its acid-responsive groups or chemical bonds.Therefore,the selection of a suitable acid responsive group or chemical bond,and through further structural optimization,is expected to construct an ideal pH-sensitive small molecule prodrug delivery system.Studies have shown that under the same micro-acid conditions,the hydrolysis capacity of the orthoester bond is 1-4 grades faster than the acetal,ketal,and hydrazine linkages.Therefore,the orthoester bond can be selected to construct a pH-sensitive small molecule prodrug that combines tumor-targeted enrichment with controlled,high-efficiency drug delivery.In this thesis,a pH-sensitive small molecule prodrug delivery system was constructed based on the orthoester bond,5-fluorouracil and stearyl alcohol.A pH-sensitive small molecule prodrug was prepared by embedding the hydrophobic chemotherapy drug doxorubicin(DOX).Synergistic delivery system finally through a series of anti-tumor experiments in vitro and in vivo to evaluate its chemotherapy effect.The specific research contents are as follows:(1)Transesterification of stearyl alcohol with 2,2,2-trifluoro-N-(2-methoxy-[1,3]dipentane-4-methylene)acetamide(OE),the product reacted with 5-fluorouracil-l-ace-tic acid to prepare a pH-sensitive small molecule prodrug(C18-oe-fu).As a control,stearyl alcohol was directly reacted with 5-fluorouracil-l-acetic acid to form no reaction.The 5-fluorouracil small molecule prodrug(C18-5fu)of the orthoester bond was confirmed to be correctly synthesized by 1HNMR,13CNMR and mass spectrometry,respectively.(2)Two small molecule prodrugs were prepared into 5-fluorouracil nanoprodrug nano transport system(NP1)and pH-sensitive 5-fluorouracil nanoprodrug transport system(NP2)by oil-in-water single-layer emulsion evaporation method.The particle size of the two prodrug nano-transport systems was determined to be within 200 nm by transmission electron microscopy(TEM)and nano-particle size analyzer(DLS),and the morphology was regular and spherical.The orthoester NMR and particle size degradation experiments show that NP2 has no degradation of the orthoester bond in the neutral environment,and the particle size does not change.The original acid ester bond degrades and the particle size decreases gradually at pH 5.0,while NP1 is at pH 7.At the time of pH7.4 and pH5.0,the orthoester bond was not degraded and the particle size was unchanged.By comparing the in vitro degradation behavior of NP1 and NP2,the results show that the NP2 bonded to the orthoester bond has the intracellular pH degradability of the tumor.(3)Another antitumor drug,doxorubicin,was packaged in two small molecule prodrug nanotransport systems to construct a small molecule prodrug synergistic delivery system(NP/DOX),and the particle size was measured at about 200 nm.The particle size is relatively small and the drug loading efficiency is relatively high(specific values are required).In vitro drug release experiments showed that 5-fluorouracil and doxorubicin in NP1/DOX released slowly at pH7.4 and pH5.0,and released about 20%after 120 hours,while 5-fluorouracil and doxorubicin in NP2/DOX The release was slow at pH 7.4,but the release accelerated at pH5.0 and about 80%after 120 hours.The results of laser confocal microscopy and flow cytometry showed that the two nanoprodrugs could be efficiently taken up by A549,HepG-2 and H22 cells,and the uptake ability of NP2/DOX was stronger than NP1/DOX.It may be due to the fact that the pH-sensitive nanoprodrug synergistic delivery system is more easily degraded and released in the cells after being taken up by the tumor cells.In vitro cytotoxicity experiments showed that pH-sensitive small-molecule prodrug delivery systems showed stronger ability to kill tumor cells,24 or 48 hours after co-culture with three cancer cells,regardless of blank or doxorubicin-containing prodrugs.This is closely related to its faster intracellular acid responsiveness and drug release capacity.In addition,the results of apoptosis experiments also showed that the pH-sensitive nanoprodrug delivery system has a more effective ability to promote apoptosis.(4)The drug distribution experiment of small molecule prodrug nano-transport system in mice showed that the pH-sensitive small molecule prodrug delivery system can significantly enhance the drug concentration in blood and tumor sites,and can reduce the distribution of drugs in normal tissues.In vivo anti-tumor experiments showed that the pH-sensitive small molecule prodrug nano-cooperative delivery system has stronger anti-tumor effect and can significantly prolong the life of negative tumor mice.In summary,the pH-sensitive small molecule prodrug synergistic delivery system with clear structure and simple preparation method can significantly reduce toxic side effects and enhance anti-tumor effect.Through further functional modification and structural optimization,a pH-sensitive small molecule prodrug synergistic delivery system based on orthoester linkage,5-fluorouracil and doxorubicin is expected to be applied in clinical research. |
PDF Full Text Request