| We have got a new kind of photosensitizer by connecting5-ALA to C60throughthe effective combination of chemical synthesis methods in this article.5-ALA is anone-phototoxicity prodrug and it can generate PpIX in vivo which can canselectively kill cancer cells,what’s more it’s also a kind of drug in endogenousphotodynamic therapy without damage to normal cells.5-ALA-PDT has theadvantages of simple method、high selectivity and low toxicity, etc.But its highlywater-soluble making it difficult to enter the cell through the cell membrane.WhileC60is an ideal drug carrier and it has made great achievements in anti-cancerresearch.However, the seriously hydrophobic of C60has hinder its better applicationand development.So we use C60as a carrier combining with5-ALA to form a kind ofwater-soluble nano-system and this system can strengthen the phototherapy effect of5-ALA under the condition of reducing the amount of5-ALA and enhance it’s passivetargeting and cell penetration according to the EPR effect.The main content of this paper is divided into two parts（:1）、The killing effect of5-ALA on B16-F10cells in vitro;（2）、The antitumor effect of C60-5-ALA on mousemelanoma model in vivo.The killing effect of C60-5-ALA on B16-F10cells is divided into twoparts:uptake experiments in vitro and anti-tumor activity in vitro.①、B16-F10cellswere selected as a cell model to study the cellular uptake of the drug to study thegeneration of PpIX after the effect of the drug to the cells.Cells seeded underfluorescent microscopy showed that C60-5-ALA group at the time of2h generated redfluorescence,suggesting that PpIX has generated and and the red fluorescenceintensity was strong at4h which can initially infer that PpIX has increased when theeffect time was longer.While the influence of the effect time of the drug to the cellsand the concentration of C60-5-ALA to the content of PpIX in cells were detected byHPLC.The results showed that the effect time of the drug to the cells was2h and theoptimum concentration of C60-5-ALA is200μg/ml.②、The anti-tumor activity in vitro was studied by SRB to determinate the influence to cell viability of differentconcentration of5-ALA and C60-5-ALA under650nm(20J/cm2）laser irradiation. Theresults showed when the cells were affected by different concentrations of5-ALAunder650nm(20J/cm2）laser irradiation,their survival rate were all above75%,whilethe cells were affected by different concentrations of C60-5-ALA under650nm(20J/cm2) laser irradiation, their survival rate decreased when the theconcentration of C60-5-ALA was improved,and it showed a concentration-dependentmanner.The cells survival rate was45.44±5.22%When the concentration ofC60-5-ALA was200μg/ml(include5-ALA90μg/ml) and the significant inhibitoryeffect was produced under those conditions.It’s showed that C60-5-ALA hassignificant antitumor activity by comparing with5-ALA(P <0.05).The anti-tumor effect in vivo studies of C60-5-ALA on mouse melanoma modelincludes the establishment of mouse melanoma model、antitumor effect study invivo、frozen sections and the generation and determination of PpIX in the organizationand pathological study.①、The tumor formation rate was97.37%in the establish of amouse model of melanoma tumor;②、 The study of antitumor effect in miceexperiments in vivo has mainly investigated the living conditions and the changes ofvolume over time, showing the tumor appreciation rate was7.41%under dose ofC60-5-ALA(include5-ALA30mg/kg) and the tumor appreciation rate was39.93%under dose of30mg/kg of5-ALA, and there is significant difference between them(P<0.05), shows that the effect of C60-5-ALA was better than5-ALA.③、In the studyof determination of frozen sections and the content of PpIX in the organization,thefrozen sections of the light illumination group of C60-5-ALA and5-ALA were made.Itcan be visually observed that fluorescent of tumor part in C60-5-ALA group,whilefluorescent of tumor part in5-ALA group was not very obvious.And it showed thatPpIX was produced in the tumor part in C60-5-ALA group.While the quantitativedetection of PpIX determined the content of PpIX of heart、liver、spleen、lung、kidneyand tumor tissue after the main injection via the tail vein of mice of C60-5-ALA and5-ALA.The results showed that the content of PpIX of tumors was the most and theother organs was fewer,while the least in heart and spleen in the C60-5-ALAgroup.The content of PpIX in the tumor tissue was fewer in the5-ALA group than C60-5-ALA group.It showed that the content of PpIX gathered mainly in the tumorparts after the effect of C60-5-ALA.④、The results of pathology showed that tumorcells were necrotic in large area,while smaller cell damage of other tissues.It showedthat the content of PpIX gathering in the tumor parts was more than any otherparts,PpIX has played a pharmacodynamic effects. |
PDF Full Text Request