Preparation And Evaluation Of Solid Dispersion System For Tumor Targeted Celecoxib Prodrug Based On Electrostatic Spray Technology

Celecoxib（CXB）is a COX-2 inhibitor,which is mainly used for analgesia,anti-inflammatory and anti-rheumatism.Recently,the researchers found that CXB also has an anti-colorectal cancer effect.However,celecoxib with low water solubility limited dosage form of the development and application,poor targeting resulted in high side effects.In this paper,the structure of celecoxib was modified to obtain p H-sensitive prodrug,which can improve the targeting of tumor cells.The prodrug was prepared into solid dispersions by electrostatic spraying technology to improve its solubility,which provides possibilities for the development of CXB injection.The prodrug of celecoxib was prepared,the prodrug was characterized and its purity,stability and physicochemical properties were investigated.The celecoxib prodrug solid dispersion（CXB-m PEG-SD）was prepared by electrostatic spray technology,the optimal carrier and drug load ratio were selected by single factor and the optimal process was selected by orthogonal experiment,the MTT assay was used to evaluate the biosafety of the carrier and the bioactivity of the prodrug solid dispersion,and the cell uptake of the prodrug solid dispersion was evaluated at p H 6.0 and p H 7.4.Finally,the in vivo analysis method of celecoxib was established and verified by HPLC.Pharmacokinetic experiment was conducted on SD rats to fit the pharmacokinetic parameters.Tissue distribution and in vivo imaging experiments were conducted on Balb/c mice to evaluate the distribution of prodrug and prodrug solid dispersions in vivo.Pharmacodynamic experiments were conducted in Balb/c mice to evaluate the antitumor effect of celecoxib and prodrug solid dispersion.The main conclusions are as follows:（1）In this paper,celecoxib prodrug（CXB-m PEG）was prepared by a two-step reaction.The first step is to obtain the intermediate m PEG-NPC and the successful synthesis of m PEG-NPC is proved by ¹H-NMR and infrared.In the second step,the prodrug（CXB-m PEG）with carbamate bond was generated.The successful synthesis of CXB-m PEG was demonstrated by ¹H-NMR and infrared spectroscopy,The prodrug was stable in water,methanol,and p H 7.4 PBS（simulated the p H surrounding normal cells in the body）,and unstable in p H 6.0 PBS（simulated the p H of the environment around the tumor cells）.The purity of prodrug was 98.21%,the content of CXB in prodrug was 15.5%.The solubility of CXB in water was 3.03μg/m L,and the solubility of prodrug in water was 156.77μg/m L,The oil/water partition coefficient of CXB was1.96,and that of prodrug was 0.80.（2）Using water:methanol:DMF=2:1:1 as solvent,the optimal prescription process of electrostatic spray was determined as follows:The prodrug:PVP K30 was1:3,the positive voltage was+16 k V,the negative voltage was-1.5 k V,the flow rate was 0.05 mm/min,the receiving distance was 18 cm,the needle diameter was 0.34 mm.The results of XRD and DSC showed that the prodrug solid dispersion was successfully prepared and the dissolution rate of the prodrug solid dispersion was higher than that of celecoxib and prodrug.At the maximum vector concentration of 200μg/m L,the cell survival rate still reached 90%and the carrier has good biological safety.CXB-m PEG-SD can inhibit tumor cells in a time-dependent and concentration-dependent manner at both p H 7.4 and p H 6.0,and CXB-m PEG-SD has a significantly higher cell inhibition rate at p H 6.0 than p H 7.4.Cell uptake experiments showed that CXB-m PEG-SD was more likely to release CXB at p H 6.0 and cell uptake increased.（3）In vivo methodology of celecoxib was specific,the precision,stability and recovery rate met the requirements.The pharmacokinetic results showed that CXB was eliminated quickly.The C_max and T_max of CXB and CXB-m PEG-SD were almost the same,but the half-life of CXB and CXB-m PEG-SD increased from 5.48 h to 9.44 h,about 1.7 times,and the average retention time increased from 6.33 h to 9.79 h.The circulation time of the drug in vivo can be prolonged and the drug has a certain slow-release effect when the drug is prepared into the solid dispersion of prodrug by electrostatic spraying technology.The tissue distribution results showed that CXB was more distributed in the liver and spleen and less distributed in the tumor after caudal vein injection,but less in the tumor,the content of CXB-m PEG-SD group gradually decreased in all tissues.CXB-m PEG-SD group accumulated slowly in the body,and its content in the tumor site was significantly higher than that of CXB group at the same time.In vivo imaging results showed that Cy7@CXB 4 h will produce accumulation in the tumor site,and also distributed in other organs,Cy7@CXB-m PEG-SD 8 h in the tumor site of a large amount of accumulation,less distribution in the heart,lung,kidney,has a good tumor targeting ability.The pharmacodynamic results showed that the tumor volume in the prodrug solid dispersion group was much smaller than that in the normal PBS group（P<0.001）.The inhibitory rate was 59%in the prodrug solid dispersion group and 33%in the celecoxib group.According to the results of tumor volume,tumor volume change curve,weight change curve and tumor inhibitory rate,the results showed that both celecoxib and prodrug solid dispersion inhibited the growth of tumor cells,but the prodrug solid dispersion had higher inhibitory rate and better effect.CEA immunohistochemical results showed that CEA was highly expressed in PBS group,but CEA had relatively low expression in celecoxib and prodrug solid dispersion.Both CXB and CXB-m PEG-SD had inhibitory effect on tumor cells.H&E staining was performed on tumor tissue and mouse heart,liver,spleen,lung and kidney.The results showed that the prodrug solid dispersion was more destructive to tumor tissue,the space between cells was increased,and the organs of mice were not damaged,indicating that prodrug solid dispersion of safety was high and could prolong the survival time of tumor-bearing mice.