The Study Of Clinical And Genetic Features In Adult Acute Lymphoblastic Leukemia Patients With T(1;19)(q23;p13)/E2A-PBX1

Objective1.To study the clinical characteristics of adult acute lymphoblastic leukemia patients with t(1;19)(q23;p13)/E2A-PBX1 and explore prognostic significance of different treatments.2.To analyze the genetic features of adult acute lymphoblastic leukemia patients with t(1;19)(q23;p13)/E2A-PBX1 and research its pathogenetic mechanism.Methods1.In this study,we retrospectively analyzed 43 t(1;19)(q23;p13)/E2A-PBX1 positive B-ALL patients' clinical characteristics,treatment response and survival.2.23 t(1;19)(q23;pl3)/E2A-PBX1 positive acute lymphoblastic leukemia patients'genomic DNA samples from bone marrow were collected.The patients' DNA were detected by next generation sequencing(NGS)and whole exome sequencing(WES)in order to analyze their gene expression profiles.PyClone and ClonEvol were used to reveal possible clonal evolution of disease pathogenesis and progression.Results1.There were 20 males and 23 females in this study.The median age for the entire study population was 26 years(range,16-85 years).9 patients had only t(1;19)(q23;p13),whereas the remaining 23(64%)showed abnormal karyotypes.5 patients had complex karyotype and 3 patients had 13q-.These patients had significantly higher bone marrow blast level(P=0.025)and lower PLT level(P=0.022)than control group at diagnosis.The 3-year overall survival(OS)and diease-free survival(DFS)estimates for the entire patient cohort were 30.2%(13/43)and 20.9%(9/43),respectively.69.8%patients(30/43)relapsed,including 12 patients relapsed twice and 3 patients relapsed 3 times,which was dramatically higher than control group(P=0.008).Also,their median complete remission time(4.0 months;range,0.5-15 months)was significantly shorter than control group(P=0.009).39.5%patients(17/43)underwent HSCT,while the other 60.5%patients(26/43)had chemotherapy(chemotherapy-alone group).HSCT could significantly improve these patients' 3-year OS,DFS and relapse rate(HSCT group vs.chemotherapy-alone group,OS:P=0.002,DFS:P=0.007,relapse:P=0.018).Additionally,patients whose MRD level at 30 days after induction chemotherapy were lower than 2.4×1014 had dramatically better 3-year OS,DFS and relapse rate(OS:P=0.002,DFS:P=0.012,relapse:P=0.005).By multivariate analysis,we found that higher MRD level at 30 days after inducing chemotherapy(? 2.4 × 10-4)could independently predict adult t(1;19)(q23;p13)/E2A-PBX1 positive ALL patients' OS and relapse(OS:HR=2.409,95%CI 1.048-5.540,P=0.039;relapse:HR=2.95,95%Cl 1.299-6.702,P=0.010).Meanwhile,3 patients received anti-CD 19 CAR-T in this study,one of them relapsed and died after transfusion and another patient died from cytokine release syndrome(CRS).2.NGS was performed on 23 t(1;19)(q23;p13)/E2A-PBX1 positive ALL patients'genomic DNA,including 18 DNA samples at diagnosis and 7 DNA samples at relapse.Mutations were detected in 16 of the 18(88.9%)E2A-PBX1-positive ALL patients at diagnosis.A total of 103 genomic aberrations,involving 45 genes,were recorded.The most frequently mutated genes were histone methyltransferase genes(14/103,13.6%),including KMT2D,KMT2C,KMT2B and WHSC1.We found that these mutations were grouped in chromatin-modifying genes(18/103,17.5%),signaling genes(16/103,15.5%),tumor suppressor genes(13/103,12.6%),transcription-factor genes(10/103,9.7%)and DNA-methylation-related genes(2/103,1.9%).44 genomic aberrations were detected in 7 relapsed ALL patients with E2A-PBX1.30 genes were involved in these molecular events.7 genes were first detected at relapse,while BCL6,DICER1,FOXO1 and PTPN13 were the most common lesions.There was no statistic difference in all aberrated genes between diagnosis samples and relapse samples.We performed WES in 9 cases,and found that 55.6%patients had abnormalities in genes involved in ubiquitin-proteasome system(CDC27,USP40,USP5).Structural modeling was performed to explore the possible biological effect of USP5 mutations.We found that c.272delC(p.Pro91Hisfs)could lead to the loss of function of USP5 while c.2222C>A(p.Ala741Asp)could weakened its ubiquitin binding ability.Results of RT-PCR showed decreased expression of USP5,P53 and 21 in 697 and KASUMI cell lines,which were E2A-PBX1 fusion positive.We further analyzed WES results and identified clonal evolution in 2 pairs of samples at diagnosis and relapse.One patient had a subclone of the founding clone,which survived from initial therapy,gained additional mutations and expanded at the second relapse.In the other patient,one subclone acquired additional gene mutations affecting DNA repairing,ubiquitination and tumor suppressor gene,which made it evolved to the dominant clone at relapse.Their founding clone remained high proportion during the process of the disease.Conclusion1.t(1;19)(q23;p13)/E2A-PBX1 positive B-ALL patients tended to relapse in the short term,resulted in poor prognosis.HSCT could improve these patients' prognosis.Higher MRD level at 30 days after induction chemotherapy could independently predict adult t(1;19)(q23;p13)/E2A-PBX1 positive ALL patients' OS and relapse.2.The most frequently mutated genes in t(1;19)(q23;p13)/E2A-PBX1 positive B-ALL patients at diagnosis were histone methyltransferase genes,including KMT2D,KMT2C,KMT2B and ARID1A,while BCL6,DICER1,FOXO1 and PTPN13 were the most frequently mutated genes at relapse.WES test showed that 22.2%patients had reproducible USP5 mutation,and our preliminary results demonstrated that USP5 might participate in the pathogenesis of t(1;19)(q23;p13)/E2A-PBX1 positive B-ALL.Clonal evolution analysis showed that new gene mutations might participate in the disease progresses.