Mechanisms And Therapeutic Studies Of Mutant P53 In The Development Of Esophageal Squamous Cell Carcinoma

Esophageal cancer is one of the deadly malignancies that ranks sixth in the cause of overall cancer mortality worldwide.Esophageal squamous cell carcinoma(ESCC),the major histopathological type of esophageal cancer,accounts for approximately 90%of all cases.ESCC is also the main subtype of esophageal cancer in China,and the patients with ESCC usually have a high malignancy and poor prognosis.Therefore,identifying the critical drivers of ESCC and revealing their functional mechanism are important for the establisnent of new anticancer strategies.Firstly,we performed whole-genome sequencing of 663 pairs of ESCC tumor tissues and paired normal tissues.The results showed that TP53 is the most frequent mutated gene,accounts for 78%,and ESCC samples from our cohort have more dispersed distribution of TP53 mutants than European and American ESCC samples in the International Agency for Research on Cancer(IARC)database.TP53 is the most important tumor surpressor gene in the body,and more than half of human cancers have TP53 mutations.The common mutation types of TP53 include missense mutation and nonsense mutation.Several p53 mutants not only lose the tumor suppressive roles of wild-type p53,but also acquire a series of oncogenic roles.p53 mutations promote cancer cell development,survival and metastasis,and lead to resistance to drug therapy and poor prognosis.Currently,no drugs targeting oncogenic p53 mutants have been approved for cancer treatment,so elucidating the oncogenic mechanism of p53 mutants is critical to developing cancer treatment strategies targeting p53 mutants.In current study,we sought to identify the function and potential mechanism of p53 mutations and explore the targeted therapeutic strategies in ESCC.At first,we selected the missense mutations and nonsense mutations,the mutations with high frequency in our ESCC cohort,and constructed cell lines expressing p53 wild-type and mutant.The roles of p53 mutations in ESCC were determined by performing Transwell assays,clonogenic assays and CCK-8 assays.The results indicated that the majority of p53 mutations exerted oncogenic roles and disrupted the inhibition of proliferation,migration and invasion mediated by wild-type p53.Mechanistically,p53 mutations alter its protein subcellular localization and stability.The p53 mutation G245S(p53-G245S)interacted with heterogeneous nuclear ribonucleoprotein A2B1(hnRNPA2B 1)to increase protein translation of phosphatidylinositol-dependent Arf GAP(AGAP1)by promoting AGAP1 mRNA stability.AGAP1 promotes cancer cell proliferation and metastasis by enhancing exosome formation.Furthermore,we demonstrated that a combination of the HSP90 inhibitor HSP90i and the AGAP1 inhibitor QS11 inhibits ESCC cell proliferation and metastasis.In addition to the above studies,based on our p53 mutation data in the ESCC cohort and combined with the latest research results,we also actively explore other potential and feasible therapeutic strategies targeting p53 mutation.We first used existing public software and websites to predict the harmfulness of p53 mutations;Prediction results showed that most of the mutations were deleterious mutations,which would affect the normal function of the proteins in the cell.The latest research results show that AS atoms can bind specifically to structural p53 mutations to save structural p53 mutations.Therefore,combining with our p53 mutation data of esophageal squamous cell carcinoma,we reconstructed structural p53 mutation cell lines that may be sensitive to As2O3(ATO),then explored the function of the mutation and sensitivity to ATO,and confirmed that the cell lines with structural p53 mutation were generally more sensitive to ATO,and some structural p53 mutation cells were extremely sensitive to ATO.Mechanistically,ATO can restore the wild-type conformation of mutant p53 and activate the reaction element of wildtype p53.Finally,we explored the effect of ATO combined with inhibitors of activated downstream signal PI3K/AKT pathway in the treatment of mutant p53,providing a potential strategy for targeting mutant p53 in the treatment of ESCC.In conclusion,our study identified the interaction between p53-G245S and hnRNPA2B1 for the first time,and further revealed that p53-G245S/hnRNPA2B1/AGAP1 regulatory axis mediated exosome formation and promoted ESCC cell proliferation and metastasis.We also proposed the combination of HSP90 inhibitor and AGAP1 inhibitor as a potential therapeutic strategy to inhibit the progression of ESCC.Finally,we found that some structural p53 mutations in ESCC are extremely sensitive to ATO,and inhibitors of PI3K/AKT signaling pathway can enhance the therapeutic effect of ATO on structural p53 mutations.