The Preparation Of Tanshinone ⅡA Solid Dispersion And Its Mechanism Of Inhibiting Colon Cancer Cell Proliferation

Objective:(1)In order to improve the bioavailability of tanshinone ⅡA(Tanshinone ⅡA,Tan ⅡA)in vitro and in vivo,the preparation and process optimization of tanshinone ⅡA solid dispersion were completed.(2)Using network pharmacology to sort out the mapping targets of tanshinone ⅡA and colorectal cancer(CRC),and analyze its regulatory effect on the biological processes and signaling pathways of colorectal cancer cells.(3)To reveal the specific molecular mechanism of tanshinone ⅡA anti-colorectal cancer activity and inhibiting the proliferation of colorectal cancer cells.Methods:(1)High performance liquid chromatography was used to establish the content detection method of tanshinone ⅡA and carry out methodological investigation.The formulation of tanshinone ⅡA solid dispersion was optimized by single factor investigation,and then the quality was evaluated by characterization experiment.(2)Retrieval of component and disease database to mine tanshinone ⅡA and colorectal cancer-related targets.The "component-target" network of the two overlapping targets was drawn;the core targets in the network were used in the DAVID database for GO biological annotation and KEGG pathway enrichment analysis.(3)The inhibitory effect of tanshinone ⅡA on the viability of two colorectal cancer cells was detected by CCK-8 assay.Cell cycle and apoptosis were assessed by flow cytometry and morphological observation.Western blotting was used to detect the expression levels of cyclins,gene MOF and their downstream targets.Crystal violet staining and growth curves were performed to evaluate the growth and proliferation ability of colorectal cancer cells in vitro.The activation of MOF gene was detected by immunofluorescence,and the distribution of MOF in colorectal cancer cells was further observed by the separation of cytoplasm and nuclear protein.At the same time,the RNA transcription levels of MOF and P53 were detected by PCR.Results:(1)The results of the methodological experiments were all good.Single factor investigation determined the optimal formulation of solid dispersion Tan ⅡA-F68-SD(m Tan ⅡA/m F68=1:7),the cumulative dissolution rate reached 88.20% at 120 min,and the saturated solubility was significantly higher than that of the API.Characterization experiments show the presence of hydrogen bonds in the solid dispersion,in a eutectic state.(2)Network pharmacology analysis found that tanshinone ⅡA overlapped with 125 targets in colorectal cancer,and participated in the regulation of cell cycle,apoptosis,proliferation and other pathways.(3)CCK-8 assay confirmed that tanshinone ⅡA inhibited cell viability and proliferation of SW480 and SW620.Flow cytometry showed that tanshinone ⅡA blocked colorectal cancer cell cycle in G2/M phase and promoted cell apoptosis.The results of western blotting indicated that the expression of cell cycle-related proteins Cyclin D1,CDK2,and CDK4 was decreased,and the proliferation and colony formation ability of two colorectal cancer cells were inhibited in a dose-dependent manner.Tanshinone ⅡA can also reduce the expression level of MOF and prevent it from entering the nucleus of SW480 and SW620 cells,and down-regulate the expression level of histone H4k16 ac.Compared with the control group,the RNA transcript levels of MOF and P53 were significantly decreased in colorectal cancer cells receiving tanshinone ⅡA.Conclusion:Tanshinone ⅡA solid dispersion with higher saturation solubility and cumulative dissolution was prepared.Network pharmacology summary Tanshinone ⅡA regulates multiple pathways related to the pathogenesis of colorectal cancer,including cell cycle,cell proliferation and apoptosis.In vitro experiments confirmed that tanshinone ⅡA inhibited the proliferation and growth cycle of colorectal cancer cells in vitro.This was due to changes in the levels of cell cycle-related proteins,MOF,and H4k16 ac,as well as significant inhibition of RNA transcription levels of MOF and P53.These findings provide a theoretical basis and basis for in-depth study of the molecular mechanism of tanshinone ⅡA in the treatment of colorectal cancer.