| 2-methoxyestradiol was first found in the urine of pregnant women as theestrogen metabolites, but it almost does not have the estrogen-like effects. In recentyears, many studies have shown that this compound is a high specificity and lowtoxicity anti-cancer drug, it can inhibit kinds of malignancies. However, owing to itspoor water solubility, oral absorption without the law, short half-life (about12minintravenous), domestic and international researchers prepared2-ME to solidnano-liposomes, phospholipid complex, suspensions and so on to overcome itsshortcomings, but the effect is still not ideal. In this study, we explore an innovativecompound2-ME sustained-release implants which is composed by2-ME andgefitinib and polylactic acid-glycolic acid (PLGA) as a carrier. The preparation wasinjected directly into the lesion site and drug directly play a role, and can achieve thesustained release effect via slowly release drugs.The effects of2-ME and gefitinib alone and their combination on MCF-7,A549,4T-1cells proliferation were tested by SRB assay, the IC50values of threetypes of cells treated by2-ME for72h were0.96μmol·L-1,0.866μmol·L-1,0.692μmol·L-1. Cytotoxicity experiments in vitro showed that,2-ME and gefitinib hadgood anti-tumor effect, and presented the concentration and dose-dependent manner.Moreover, when2-ME and gefitinib used in combination, the combination index wasless than1of A549cell for72h, showing synergistic effect at low concentrations.Combined with the combination index, we finally determine the best proporation of2-ME and gefitinib was2.1:1, which is used for preparing the sustained-releaseimplants.We established the HPLC methods for the determination of2-ME and gefitinibin2-ME compound preparation. The Chromatographic conditions were as follows:mobile phase consisted of methanol and water (75:25,v/v); flow rate0.1ml min-1;column temperature25℃; UV-detector at285nm. The regression equation of2-MEand gefitinib were: A=0.2599C+0.0488(r=0.9998), A=0.4004C-0.1305(r=0.9999).Under the chromatographic conditions, the precision in days and between days of 2-ME were1.57%and1.38%, the average recovery was99.87%(RSD=0.90%, n=9);the precision in days and between days of gefitinib were0.93%and1.08%, theaverage recovery was99.55%(RSD=0.67%, n=9). The result indicated that this methodwas fitted for the demand of sample analysis and detection.By measuring the solubility of2-ME and gefitinib in different release media,finally determine the in vitro release medium was1%SDS-PBS. In this study,through the single factor method, we investigate if the different types of PLGA,different drug loading, different preparation methods have an effect on drug releasebehavior. Identified PLGA (75:25) as the carrier; the amount of drug formulation was60%; the solvent evaporation method was used. In vitro release of the implants, after6days,16days,30days,2-ME cumulative release were33.9%,56.6%,73.1%respectively.In vivo release experiment of compound2-ME sustained-release implants werecarried out in mice. The effects shows that after13days,45days,54days, thecumulative release percentage reached30%,50%,72.8%, indicating that the agenthas a sustained release effect. The drug release for the implants were best fitted withRitger-Peppas model. According to the Ritger-Peppas, the predominant releasemechanism was drug diffusion and erosion of the skeleton.In this study, the mice tumor model was established to assess the anti-tumoreffect of in the compound2-ME release implants by subcutaneous injection the S180cell. The results indicated that the compound2-ME release implants have anti-tumoreffect. Relative tumor appreciation rate of each group shown below: low-dose ofcompound2-ME release implants was49.13%, medium-dose of implants was38.96%(<40%), high-dose of implants is38.55%(<40%), blank PLGA implantgroup was82.29%. |
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