Chemo-resistance Mechanism Study Of TP53 Mutations In Relapsed Childhood ALL And Treatment Strategy Exploration

Resistance to chemotherapy remains the leading cause of treatment failure and death in relapsed childhood acute lymphoblastic leukemia(ALL).Genomic and clinical analysis shows that TP53 mutations play a key role in driving chemo-resistance and relapse in childhood ALL.In our study,we integrated clinical analysis with functional study to investigate the role of TP53 mutations in chemoresistance and the molecular mechanisms by which different TP53 mutations contribute to chemotherapy resistance in relapsed childhood B-cell ALL(B-ALL).And we tried to exploit the potentially effective treatment strategies for relapsed childhood ALL with TP53 mutations.Case data on 53 relapsed childhood B-ALL patients with TP53 mutations were reviewed in this study.We performed clinical analysis and found that TP53 hot spot mutations,different from non-hot spot mutations,are associated with on-treatment relapse and non/poor-response to chemotherapy in childhood B-ALL.Utilizing TP53 knockout and TP53 mutant re-expression B-ALL cell model,we demonstrated that replacing wild-type p53 with hot spot mutants and the majority of non-hot spot mutants causes resistance to multiple chemotherapy drugs used in the ALL treatment,but reexpression B-ALL cells with several non-hot spot mutants are still sensitive to all or some chemotherapy drugs.Topoisomerase Ⅱ inhibitor idarubicin induced DNA double-strand breaks(DSBs)of all p53 mutant cells,induced cell cycle arrest and apoptosis in cells over-expressing several p53 non-hot spot mutants,but failed to induce cell cycle arrest and apoptosis in cells overexpressing p53 hot spot mutants and the majority of non-hot spot mutants.The high-throughput sequencing of chromatin immunoprecipitated DNA(Ch IP-seq)results showed that p53 hot spot mutants and the non-hot spot mutant R337 C are deficient in binding to the p53’s consensus element,but the non-hot spot mutant R110 H is still able to binding to the p53’s consensus element.In addition,mutant p53 reactivator PRIMA-1 rescued the induction of p21 and PUMA in TP53 hot spot mutant cells and reversed their resistance to idarubicin.In a word,these findings suggest loss of p53 tumor suppressive function in hot spot mutants and most non-hot spot mutants as a primary driver of chemotherapy resistance in relapsed childhood B-ALL and provide a potential therapeutic strategy to overcome chemo-resistance.It was reported that AICAr(5-amino-4-imidazolecarboxamide riboside,AICAr,also called Acadisine)might be a valuable agent for the treatment of relapsed/refractory chronic lymphoblastic leukemia(CLL)patients in clinical trial and AICAr induces apoptosis independent of p53 in CLL cells.We further assessed the role of wild-type p53 function in the cytotoxic effects of AICAr in childhood ALL cells,and studied the mechanisms by which AICAr inhibits ALL cell proliferation.We demonstrated that AICAr-induced proliferation inhibition was dependent on p53 but independent of AMPK activation.AICAr induced the NTP and dNTP pool imbalances between purine and pyrimidine(a decrease in pyrimidine biosynthesis),which resulted in perturbed r RNA transcription and DNA replication stress,DNA DSBs,and p53 activation.Finally,proliferation inhibition and cell death were induced by AICAr in ALL cells.Our data indicate that RNA transcription inhibition and DNA replication stress induced by NTP and dNTP pool imbalances might play a key role in AICAr-mediated cytotoxic effects on ALL cells,suggesting a potential clinical application of AICAr in future ALL therapy.