Small Molecule Inhibitors Targeting FOXM1 Inhibit The Proliferation Of Ovarian Cancer Cells

Research Background:Ovarian cancer is one of the three malignant tumors of female reproductive organs;The mortality rate is the highest among the three malignant tumors of female reproductive organs.Ovarian cancer is more secretive and hidden,Because it’s not easy to be found early.So about 75%of patients are late at the time of diagnosis,And it has poor survival and poor prognosis.Relapse is one of the leading causes of failure of ovarian cancer treatment and patient death.At present,the treatment method of ovarian cancer mainly adopts the standard plan of combination of tumor cell and platinum-type chemotherapy;But so far,drug resistance has brought a great challenge for cancer treatment.In recent years,with the rapid development of medical molecular biology theory and its biological technology,small molecular targeted therapy for ovarian cancer has become a focus of medical attention.The global treatment of ovarian cancer drugs has been developed and has remarkable therapeutic effect.In Georgia and Armenia,for example,small molecule drug ginsenoside has been registered.The small molecular compound can not only significantly inhibit the rapid growth of ovarian cancer cells,But also induce apoptosis of cancer cells.And that molecular targeted therapy has lots of advantages,such as high specificity,high efficiency and low toxicity.Therefore,small molecular targeted therapy has good application prospect.The FOXM1 molecule is a star of the 2010 year;It’s a member of the Fox transcription factor family.It contains a conservative DNA domain.The human transcription factor FOXM1 is located on the 12p13.3 chromosome and it contains 10 exons.According to different ways of connection between the Ⅴa and Ⅶa,There are three FOXM1 molecular subtypes,They are FOXM1a,FOMX1b,FOXM1c respectively With the deepening study of FOXM1 molecules,It is found that the gene of FOXM1 is closely related to cell proliferation,and it is involved in biological processes such as cell differentiation,cell aging,stem cell growth,cell metabolism,and tumor formation.In recent years,it has been found that FOXM1 is unusually high expression in many tumors such as leukemia,breast cancer,gastric cancer,lung cancer and other solid tumors.Also,a large number of biochips and experimental studies have shown that the FOXM1 gene is highly expressed in ovarian cancer.The expression level of FOXM1 in ovarian cancer is 4-6 times that of normal tissues,The increase rate of FOXM1 is increased in patients with advanced disease.FOXM1 is closely related to the development and prognosis of ovarian cancer.As the research progresses,the 3d crystal structure of FOXM1 protein is reported,It is possible to make specific drug screening based on molecular structure.Therefore,FOXM1 is expected to be a potential new target for tumor therapy.To sum up,this subject is based on the three-dimensional spatial structure of FOXM1 protein,We selected small molecules capable of targeting FOXM1,The molecular mechanism of inhibiting the growth of ovarian cancer cells was preliminarily studied.Research contents:1.The virtual screening of small molecular compounds targeting FOXM1 and the determination of the compound XST-20.We first download the FOXM1 protein from the PDB database(3G73),The Protein was processed by the Prepare Protein module in Discovery Studio3.0.,Looking for small molecules that might bind to the target protein,and calculate the binding structure of the ligands and proteins,the potential ’key amino acids were found.The two hundred thousand molecules in the SPECS database are performed in sequence by Libdock and CDOCKER virtual docking calculations.By scoring the small molecules,we remove the molecules that were easy to take off.The pharmacokinetic properties are then predicted by ADMET and finally 27 small molecule compounds are obtained.The biological activity of 27 compounds are tested by MTT proliferation assay.The results showed that 8 small molecules were screened from 27 compounds.The effect of the genes downstream of target protein are detected by qRT-PCR、Western blotting.The results show that XST-20 could inhibit the expression of CCNB1 a downstream target of FOXM1 in 8 small molecules.We teste the ability of compound XST-20 to bind FOXM1 by SPR assay and and dual luciferase reporter assay.The results show that XST-20 is able to bind FOXM1 significantly.2.The molecular mechanism of small molecule compound XST-20 inhibits the growth of human ovarian cancer cells.we first to confirm that whether XST-20 has biological activity in the second part of the work.We evaluate the effect of XST-20 on the proliferation of ovarian cancer cells using MTT assay and plate clone formation assay in vitro.The results show that XST-20 can significantly inhibit the growth of ovarian cancer cells.Further,MTT results show that the small molecule compound XST-20 inhibit the tumor cell activity is universal.The safety assessment of small molecule compounds is as important as pharmacodynamics and the potential druggability of candidate small molecule compounds needs to be assessed by assessing toxicity.The results of virulence experiments show that the toxicity of small molecule compound XST-20 to FTE-187 cells is significantly lower than that of tumor cells.The effect of XST-20 on cell cycle.apoptosis and autophagy in ovarian cancer cells is detected by flow cytometry and Western blotting.The results show that the small molecule compound XST-20 can induce ovarian cancer cells G1 arrest,It is the concentration gradient and time-dependent.The results show that the expression of Bax arnd autophagy-related genes LC3-II,Atg7 and Beclin increased after treatment of ovarian cancer cell line XST-20,So small molecule compound XST-20 can induce apoptosis and autopaghy in ovarian cancer cells.Research conclusions:1.Small molecular compounds are screened by computer virtual screening technology.The affinity of small molecule compound with FOXM1 protein is verified by SPR technique.2.XST-20 can significantly inhibit the growth of human ovarian cancer cells and show time and dose dependence.3.A preliminary study of the toxicity of XST-20 on normal fallopian tube epithelial FTE-187 cells with a significantly lower virulence than ovarian cancer cells4.XST-20 can significantly induce G1 arrest of ovarian cancer cells in a time and dose-dependent manner.and we discuss that the XST-20 could induce autophagy and apoptosis significantly in ovarian cancer cells.