Targeted sequencing analysis of hyper-eosinophilic syndrome and chronic eosinophilic leukemia / 中华血液学杂志

Objective@#Analysis of the molecular characteristics of eosinophilia. @*Methods@#Targeting sequence to 24 patients with chronic eosinophilic leukemia (CEL) with rearrangement of PDGFRA, PDGFRB, or FGFR1 and 62 patients with hyper-eosinophilic syndrome (HES). Mutation annotation and analysis of amino acid mutation using authoritative databases to speculate on possible pathogenic mutation. @*Results@#Thirty-seven kinds of clonal variant were detected from 17 patients with CEL, no recurrent mutation site and hot spot region were found. No pathogenic mutation was detected in 19 patients with PDGFRA rearrangement, but pathogenic mutations of ASXL1, RUNX1 and NRAS were detected from 2 patients with FGFR1 rearrangement who progressed to acute myeloid leukemia and 1 patient with PDGFRB rearrangement who progressed to T lymphoblastic lymphoma, respectively. One hundred and two kinds of clonal abnormalities were detected in 49 patients with HES. The main hot spot mutation regions included: CEBPA Exon1, TET2 Exon3, ASXL1 Exon12, IDH1 Y208C, and FGFR3 L164V. CRRLF2 P224L and PDGFRB R370C point mutations were detected separately in 2 patients with HES who treated with imatinib monotherapy and achieved hematologic remission. @*Conclusion@#The pathogenesis of CEL with PDGFRA, PDGFRB or FGFR1 rearrangement is usually single, and the progression of the disease may involve other driver mutation. A variety of genes with hot mutation regions may be involved in the pathogenesis of HES, and some mutation sites are sensitive to tyrosine kinase inhibitors.

Spectrum of somatic mutations and their prognostic significance in adult patients with B cell acute lymphoblastic leukemia / 中华血液学杂志

Objective@#To investigate the spectrum of gene mutations in adult patients with B-acute lymphoblastic leukemia (B-ALL), and to analyze the influences of different gene mutations on prognosis.@*Methods@#DNA samples from 113 adult B-ALL patients who administered from June 2009 to September 2015 were collected. Target-specific next generation sequencing (NGS) approach was used to analyze the mutations of 112 genes (focused on the specific mutational hotspots) and all putative mutations were compared against multiple databases to calculate the frequency spectrum. The impact of gene mutation on the patients’ overall survival (OS) and recurrence free survival (RFS) was analyzed by the putative mutations through Kaplan-Meier, and Cox regression methods.@*Results@#Of the 113 patients, 103 (92.0%) harbored at least one mutation and 29 (25.6%) harbored more than 3 genes mutation. The five most frequently mutated genes in B-ALL are SF1, FAT1, MPL, PTPN11 and NRAS. Gene mutations are different between Ph+ B-ALL and Ph- B-ALL patients. Ph- B-ALL patients with JAK-STAT signal pathway related gene mutation, such as JAK1/JAK2 mutation showed a poor prognosis compared to the patients without mutation (OS: P=0.011, 0.001; RFS: P=0.014,<0.001). Patients with PTPN11 mutation showed better survival than those without mutation, but the difference was not statistically significant (P value > 0.05). Besides, in Ph+ B-ALL patients whose epigenetic modifications related signaling pathway genes were affected, they had a worse prognosis (OS: P=0.038; RFS: P=0.047).@*Conclusion@#Gene mutations are common in adult ALL patients, a variety of signaling pathways are involved. The frequency and spectrum are varied in different types of B-ALL. JAK family gene mutation usually indicates poor prognosis. The co-occurrence of somatic mutations in adult B-ALL patients indicate the genetic complex and instability of adult B-ALL patients.

Screening of adult Ph-like acute lymphoblastic leukemia by multiplex real-time quantitative PCR / 中华血液学杂志

Objective@#To investigate the feasibility of multiplex real-time RT-PCR with fluorescent probes in early screening of Ph-like acute lymphoblastic leukemia (ALL) and analyze the clinical feature and prognos.@*Method@#A total of 118 adult B-ALL patients diagnosed between October 2010 and March 2016 were enrolled in this study. Multiplex RT-PCR was used to detect the Ph-like ALL related fusion gene and CRLF2 expression in 58 BCR-ABL and MLL rearrangement negative patients. The clinical features, treatment response and prognosis were analyzed in Ph-like fusion gene positive and/or CRLF2 over-expression patients.@*Result@#Among 58 patients, 9 patients (9/58, 15.5%) showed Ph-like ALL related fusion genes positive and 10 patients (10/58, 17.2%) showed CRLF2 over-expression. There were statistical differences in age, WBC count, immunophenotypes, cytogenetics and risk stratification among Ph-like fusion gene positive or CRLF2 over-expression patients, Ph+ patients, MLL+ patients and B-other patients. The 2-year overall survival rates were 65%, 47%, 64% and 74% respectively among these four groups (P=0.043) . The 2-year relapse free survival rates were 51%, 39%, 62% and 70% respectively among these four groups (P=0.010) .@*Conclusion@#Routine screening of Ph-like ALL by multiplex RTPCR is feasible.

Study of the effects and mechanism of all-trans retinoic acid on leukemic cell line U937 cells with NPM1 mutation / 中华血液学杂志

Objective@#To investigate the effect and mechanism of all-trans retinoic acid (ATRA) on leukemic cell line U937 cells with NPM1 mutation.@*Methods@#Human acute myeloid leukemia cell line U937 was explored, NPM1 mutated (A type) plasmids were transfected into U937 to form stable clones A1 and A2, which were identified by Western blot and Co-immunoprecipitation. The cell proliferation was measured by methylthiazolyl tetrazolium bromide (MTT) ; cell cycle and cell apoptosis were explored by flow cytometric; cell colony formation was measured by microscope count, the molecular pathways related to cell proliferation were measured by Western blot.@*Results@#①The cell proliferations of mutant A1 and A2 were inhibited significantly by 52.6% and 35.8% (P<0.05) , respectively under ATRA exposure. ②The percentages of G0/G1 stage of mutant A1 and A2 increased by 20.1% and 35.8%, respectively under ATRA exposure. ③All the U937 leukemic cells were inhibited under ATRA exposure; the decreased percentages of vector, wild-type and mutant NPM1 cells were 32.7%, 57.9% and 90.9% respectively. ④p-ERK decreased obviously after ATRA exposure in NPM1 mutated leukemic cells. ⑤More mutant NPM1 cells inclined to apoptosis under the exposure of ATRA and cytotoxic drugs than cytotoxic drugs alone, meanwhile more cells apoptosis occurred when ATRA was administrated after cytotoxic drugs exposure.@*Conclusions@#ATRA could inhibit cell proliferation and colony formation, blocked the cell cycle in the G0/G1 stage accompanied by the significant reduction of p-ERK in U937 leukemic cells with NPM1 mutation. Besides, ATRA could synergize with drugs to suppress the leukemic cells survival more effectively when ATRA was administered after the cytotoxic drugs exposure in U937 leukemic cells with NPM1 mutation.

Gene mutation and clonal evolution in patients with myelodysplastic syndromes after treatment of azacitidine / 白血病·淋巴瘤

At the 58th American Society of Hematology Annual Meeting,there were many reports on clonal evolution related with blood disease.Gene mutation and clonal evolution after treatment of azacitidine (AZA) in myelodysplastic syndromes (MDS) were summarized to explore the relationship between clonal evolution and treatment response and clinical process in MDS,and to provide reference for clinical treatment decision.

Application of next generation sequencing in the study of hematologic neoplasms: reports from the 56th American Society of Hematology annual meeting / 白血病·淋巴瘤

In comparison with the American Society of Hematology (ASH) annual meeting last year,more studies of hematologic malignancies using next generation sequencing (NGS) are published in the 56th ASH meeting this year.NGS was mainly used in the diagnosis and classification,risk stratification,MRD detection,clonal evolution,prognosis and therapy guidance.This short report will address the application of NGS in the study of hematologic malignancies by summarizing several representative publications from the 56th ASH meeting.