Study On SMAD4 Expression And Gene Mutation In Epithelial Ovarian Cancer

Objective Detection of SMAD4 expression and the somatic mutations of exon 2,8,9,11 in epithelial ovarian cancer(EOC).Study the characteristics of their expression and genetic changes.Aimed to explore its role in the occurrence and development of EOC and provide a theoretical basis for the precise targeting therapy of EOC.Methods 1.107 cases of EOC,33 cases of benign ovarian tumors,and 35 cases of normal ovarian paraffin-embedded tissues were collected as research subjects.The two-step immunohistochemistry method was used to detect the expression of SMAD4 in three groups of ovarian tissues.The expression level of the SMAD4 in different ovarian tissues and the relationship with clinicopathological parameters of ovarian cancer were statistically analyzed by IBM SPSS Statistics 20.2.Extracted genomic DNA from 107 cases of EOC paraffin tissue specimens as the research subjects.The exon 2,8,9 and 11 of the SMAD4 gene were analyzed by Sanger sequencing after amplification by polymerase chain reaction(PCR).According to the results of sequencing,Bioedit and DNAMAN software were used to screen the gene mutation or gene polymorphism to found genetic changes.Results 1.The expression rate of SMAD4 was 79.44% in ovarian cancer,18.18% in ovarian benign tumor and 16.67% in normal ovary.There was a significant difference(?2=67.298,P<0.01).2.The expression rate of SMAD4 was 64.86% in ovarian serous cancer and 60.61% in non-serous cancer.The difference wasn't significant(?2=0.485,P>0.05).The tissue classification was 60.0% in G1 group and 82.1% in G2~G3 group.The differences were significant(?2=6.326,P<0.05).57.14% of I~II stage and 79.27% of Ill~IV stage were significantly different(?2=6.043,P<0.05).3.Exon sequencing revealed a single nucleotide polymorphism(SNP)marked as rs77389132,whose allele was A/G,located in the intron region of exon 2 and exon 3 of SMAD4 gene.No mutations were founded in exon 2,8,9 and 11,and SMAD4 gene mutation may not be the main reason in the pathogenesis of EOC.It's implied that the study of mutations in the SMAD4 gene for preliminary screening of susceptible individuals in ovarian cancer still needs further investigation.Conclusion 1.The expression of SMAD4 protein in EOC was related to clinical stage and histological grade,but not to histological type.The later of the clinical stage,the worse histological of differentiation,and the higher the positive expression rate.There was no significant correlation between the high expression of SMAD4 protein and its genetic alterations.2.SMAD4 gene somatic mutation is rare in EOC,which may be the inherent characteristics of EOC.The SNPs or mutations of somatic SMAD4 gene in EOC were demonstrated randomization and non-hot mutation characteristics,which may require we improve the gene sequencing method,and expand the sample size and use more accurate two generation sequencing to test and verify.3.The mechanism of the induction of ovarian cancer by SMAD4 gene mutation is still not clear.The discovery of SNPs provides a new possibility for exploring the changes in the molecular level of the gene in the development of ovarian cancer,and it will help us find new treatments for ovarian cancer.