The Study Of DNMT3A Mutation In Acute Myeloid Leukemia

Objective:1. To explore the clinical characteristics and prognostic significance of DNMT3 A mutations of Chromosome normal(CN)-AML patients and the role of allogeneic hematopoietic stem cell transplantation(HSCT) in treatment of DNMT3A+ CN-AML patients.2. To study the efficacy of demethylating drugs Decitabine in the treatment of DNMT3A+refractory or relapsed patients.3. To explore the significance of DNMT3 A mutation in clonal hematopoiesis.Methods:1. We retrospectively analyzed the prognostic impact and prognostic value of DNMT3 A mutations in 397 CN-AML patients who received consolidation of intensive chemotherapy or allo-HSCT in our center(Part 1: 308 CN-AML patients from March 2005 to May 2014; Part 2: 53 refractory and relapsed patients who received Decitabine as therapy from April 2011 to October 2014; Part3: 173 CN-AML patients from March 2005 to May 2015).2. The gene mutations were detected by PCR amplification of the entire or a portion of the coding region followed by direct bidirectional DNA sequencing.3. In addition, next-generation sequencing of 700 tumor associated genes were performed to explore new potential target gene and the leukemogenic effect of DNMT3 A.Results:1. The median age was 40(16-68)years old, and the median blast in bone marrow was 56.12% and the median white blood cells was 25.22(0.5-355.9)*109/L. In the whole cohort, 63 patients(20.5%) were identified and R882 H was the most frequent(n=36). DNMT3 Amut patients had shorter overall survival(3-year OS: 31.9% vs. 52.0%, P = 0.009) and disease-free survival(3-year DFS: 21.8% vs. 40.1%, P=0.004) compared with DNMT3 Awt patients. Based on FLT3/NPM1/CEBPA mutations, 308 CN-AML patients were divided into favourable/intermediate risk group(n=262) and unfavorable group(n=46). There were no significant differences in overall survival between DNMT3 Amut and DNMT3 Awt patients in both favourable/intermediate and unfavorable groups. Additionally, in multivariate analysis DNMT3 A mutation remained an independent adverse prognostic factor for the survival. In the DNMT3 Amut cohort reaching CR, 23 patients received allo-HSCT consolidation and 32 patients received chemotherapy consolidation, dramatic differences were observed in overall survival(3-year OS: 51.7% vs. 28.9%, P = 0.048) and disease free survival(3-year DFS: 41.6% vs. 14.9%, P = 0.024) between allo-HSCT group and chemotherapy group. Allo-HSCT could reverse the poor outcome of DNMT3A+ AML patients.2. We retrospectively collected treatment data from 53 refractory or relapsed CN-AML patients receiving re-induction therapy including decitabine combined with CAG and CAG- like regimen. There were 24 patients with DNMT3 A mutations while 29 patients without DNMT3 A mutations. The CR rate of DNMT3A+ patients was 54.17% and ORR was 62.50% with one course of treatment, andthe CR and ORR rate of DNMT3 Agroup was 37.93% and 48.28% respectively. No differences were observed between these two groups. Interestingly, DNMT3A+/FLT3-ITD+ CN-AML patients had higher ORR and CR rates than DNMT3A-/FLT3-ITD+CN-AML patients(P=0.040 and 0.042, respectively). Moreover, there were no differences between DNMT3A+ CN-AML and DNMT3ACN-AML patients on overall survival.3. In 173 CN-AML patients, 117 were with DNMT3 A mutation, 81 patients(81/117,69.2%) finally reached CR after induction therapy, 48 patients(48/81, 59.3%) were detected as DNMT3 A mutation positive when achieved CR. 42 patients received HSCT and 39 received chemotherapy. At the last follow-up, 40 patients remained CR and 8 CR patients(9.9%) were still detected with DNMT3 A mutation, the median follow-up time of 8 patients was 21(12-107)months. FLT3-ITD and NPM1 mutation could disappear before DNMT3 A mutation. We further detected the possible gene aberrations in 12 de novo and 2 relapse samples by second generation sequencing technology, NPM1(5/12, 41.7%), FLT3-ITD(5/12, 41.7%) and CEBPA mutations(4/12, 33.3%) were the more frequent co-existed mutations. In the relapse samples, additional genes aberrations could be observed and some of them were never reported in AML patients. The 2-year overall survival(2-OS) for 81 DNMT3 A mutated CN-AML patients was 39.0%. No differences in 2-OS(38.2% vs 41.6%, P=0.226) and 2-year disease free survival(2-DFS: 28.5% vs 34.3%, P=0.183) between patients with negative DNMT3Amut(n=33) group and positive DNMT3Amut(n=48) group at the first remission.Conclusion:1. DNMT3 A mutation was a poor prognostic factor for CN-AML patients. Allo-HSCT could improve survival of cytogenetically normal acute myeloid leukemia patients with DNMT3 A mutations.2. Decitabine likely offered better CR and ORR rate for FLT3-ITD+/DNMT3A+ refractory or relapsed CN-AML patients.3. DNMT3 A mutations burden could persistently exist in adult DNMT3 A mutated CN-AML patients with long-term remission, DNMT3 A mutation was the early events in development of leukemic cells.