The Pathological Significance Of RNF43 Gene Mutation In Colorectal Cancer

Colorectal cancer(CRC) has been one of the most common gastrointestinal malignancies in the world. It will be of great practical significance to study the pathogenesis of colorectal cancer. The activation of Wnt signal pathway occurs in the early stage of adenoma-carcinoma sequence, plays an important role in the development of CRC.RNF43(Ring finger protein 43) is an important negative regulation factor of Wnt signal pathway. RNF43 promotes Wnt receptor turnover, and inhibits Wnt signaling downstream by sequestering TCF4 to the nuclear membrane. The abnormal expression and mutation of RNF43 have great influence on the activation of Wnt signaling pathway, may be of great significance in clinical practice. Studies have reported that RNF43 mutations occur frequently in high frequency microsatellite instability tumors. The majority of truncating mutations of RNF43 occurred in MSI-H cases and rarely occur simultaneously with APC mutations, suggesting that RNF43 inactivation may be the key part of colorectal cancer development of MSI type. Analysis of RNF43 gene changes/clinical parameters/prognoses in patients with different MSI types will do help to find new prognostic markers.Both the mutation status of KRAS/BRAF and microsatellite instability status can be used as a predictor for prognosis and drug resistance, so we also analysed whether the RNF43 mutation is associated with BRAF/KRAS mutations.This study selected 446 cases of sporadic colorectal cancer samples. Samples were taken from formalin fixed-paraffin embedded tissues. There are some microsatellite loci in the exon region of RNF43, these loci may have a high mutation rate according to the correlation between the RNF43 mutation and microsatellite instability. So we designed primers for these RNF43 microsatellite loci. We extracted the genomic DNA from the samples. The microsatellite-like loci in RNF43 were amplified by PCR method, and the sequence informations were determined by Sanger sequencing method. SPSS software was used to analyze the RNF43 mutation status, KRAS/BRAF mutation status and MSI status(determined by pyrosequencing and fluorescent multiplex PCR-capillary electrophoresis), pathological parameters(Sex, age, tumor location, depth of invasion, gross type, histological type, lymph node metastasis, distant metastasis and TNM stage.), and survival time. The effect of RNF43 mutation on the expression of tumor tissue was analyzed by immunohistochemistry. We want to analyze the interrelationship between these changes and clinical parameters, and to analyze the effect of these changes on the prognosis.This study find that there are 4 kinds of RNF43 mutations in colorectal cancer in China, they are p.Gly659fs、p.Arg117fs、c.350G> A、c.816 T>C. The overall mutation frequency is 9.2%,1.1%,54.5%,0.91%.The mutation rate of p.Gly659fs is 28.6% in the MSI-H state and is 3.1% in the MSI-L/MSS type. The mutation rate of p. Arg117fs is 3.6% in the MSI-H type, and is not found in the MSI-L/MSS state. It shows no difference in the mutation rate of c.350G> A in different microsatellite states.p.Gly659fs is related with tumor in colon, tumor in right side, mucinous adenocarcinoma, low histological differentiation. C.350G> A is related with Adenocarcinoma, high histological differentiation, and these associations are present only in the case of a homozygous mutation state at this site. The five year survival rate of the patients with BRAF mutations is lower when these patients carrying the p.Gly659fs mutation. No association was found between the RNF43 mutation state and the RNF43 protein expression.The present research can draw the following conclusions:1:RNF43 is frequently mutated in CRC population;2:p.Gly659fs、p. Arg117fs mutations are associated with microsatellite instability. Their mutation rates are higher in MSI-H type than in MSI-L or MSS type;3:p.Gly659fs mutation of RNF43 gene is a protective factor for colorectal cancer metastasis;4:For those patients carrying RNF43 p.Gly659fs mutation, BRAF gene mutation will lead to a decreased five-year survival rate.