Characterization and integrated analysis of extrachromosomal DNA amplification in hematological malignancies.

The study of extrachromosomal DNA (ecDNA), an element existing beyond classical chromosomes, contributes to creating a more comprehensive map of the cancer genome. In hematological malignancies, research on ecDNA has lacked comprehensive investigation into its frequency, structure, function, and mechanisms of formation. We re-analyzed WGS data from 208 hematological cancer samples across 11 types, focusing on ecDNA characteristics. Amplification of ecDNA was observed in 7 of these cancer types, with no instances found in normal blood cells. Patients with leukemia carrying ecDNA showed a low induction therapy remission rate (<30 %), a high relapse rate (75 %) among those who achieved complete remission, and a significantly lower survival rate compared to the general leukemia population, even those with complex chromosomal karyotypes. Among the 55 identified ecDNA amplicons, 268 genes were detected, of which 38 are known cancer-related genes exhibiting significantly increased copy numbers. By integrating RNA-Seq data, we discovered that the increased copy number, resulting in a higher amount of available DNA templates, indeed leads to the elevated expression of genes encoded on ecDNA. Additionally, through the integration of H3K4me3/H3K27ac chromatin immunoprecipitation sequencing, assay for transposase-accessible chromatin with sequencing, and high-throughput chromosome conformation capture data, we identified that ecDNA amplifications can also facilitate efficient, copy number-independent amplification of oncogenes. This process is linked to active histone modifications, improved chromatin accessibility, and enhancer hijacking, all of which are effects of ecDNA amplification. Mechanistically, chromothripsis and dysfunction of the DNA repair pathway can, to some extent, explain the origin of ecDNA.

Identification of mast cells as a candidate significant target of immunotherapy for acute myeloid leukemia.

OBJECTION: Immunotherapy based on T cells is a new therapy for Acute myeloid leukemia (AML). However, there has not been considerable improvement compared with traditional chemotherapeutics. This study aimed to identify important immune cells, genes, and drugs associated with the immunotherapy of AML. METHODS: The gene expression profile and clinical data of patients with AML were downloaded from TCGA database, and the abundance ratio of immune cells was obtained via CIBERSORT. Kaplan-Meier (KM) survival analysis was used to assess the relationship between immune cells and survival time of patients with AML. Differentially expressed genes (DEGs) analysis was conducted to obtained DEGs related to mast cells. Then, protein-protein interaction (PPI) analysis and enrichment analysis were performed to explore the hub genes. Finally, Connectivity Map (CMap) database was utilized to predicts potential drugs that may reverse or induce the mast cell-related gene expression. RESULTS: Our study showed that mast cell was correlated with survival time of patients with AML, and 135 genes were screened to be related with mast cells. 6 hub genes were identified via PPI network, and 3 potential small molecule drugs were screened to be related to regulating the mast cell-related gene expression via CMap database. CONCLUSION: The hub genes and drugs have high research value and clinical application in AML therapy. Our study not only provides gene targets and small molecule drugs for AML immunotherapy concerning mast cells but also provides new ideas for researchers to explore immunotherapy targets of other tumors.

Higher red blood cell distribution width at diagnose is a simple negative prognostic factor in chronic phase-chronic myeloid leukemia patients treated with tyrosine kinase inhibitors: A retrospective study.

ABSTRACT: The aim of this study was to evaluate the ability of the red blood cell distribution width (RDW) to predict prognosis and treatment response in chronic myeloid leukemia (CML)-chronic phase (CP) patients treated with tyrosine kinase inhibitor (TKIs).We retrospectively enrolled 93 newly diagnosed CML-CP patients treated with TKIs from 2009 to 2018 at the First Hospital of Lanzhou University. Patients were divided into 2 groups using an RDW of 18.65% determined by receiver operating characteristic curve analysis. We analyzed the correlation of treatment responses and the RDW compared to common scoring systems, as well as the correlation of the RDW with disease outcome, including overall survival (OS) and progression-free survival (PFS), and demographic and laboratory factors affecting outcome. Univariate analysis and Cox regression analysis were used.The median age of patients was 40 years, and 51 patients (54.8%) were men. A high RDW could predict treatment response at 3 months (P = .03) and 6 months (P = .02). The RDW was significantly lower in patients who achieved molecular response by 3 months (P < .001) and complete cytogenetic response by 6 months (P = .001) than in those who did not respond. Patients with a high RDW (>18.65%, n = 35) had significantly worse 5-year OS (77.1% vs 96.6%; P = .008) and PFS (80.0% vs 98.3%; P = .002) than those with a low RDW (≤18.65%, n = 58). Multivariate analysis demonstrated that a high RDW was an adverse predictor of OS (P = .005, HR (hazard ratio) = 9.741) and PFS (P = .009, HR = 16.735).The RDW is a readily available prognostic marker of outcome in patients with CML-CP and can predict treatment response to TKIs. Further larger and prospective studies are required.

A novel mutation in SPTA1 identified by whole exome sequencing in a Chinese family for hereditary elliptocytosis presenting with hyperbilirubinemia: A case report.

RATIONALE: Hereditary elliptocytosis is an inherited disorder characterized by the elliptical red blood cells (RBCs) on the peripheral blood smear and related hemolysis, mainly results from a heterozygous mutation in the genes that encode protein 4.1, α-spectrin, ß-spectrin. Mutations of SPTA1 are the most common. PATIENT CONCERNS: A 21-year-old female presented with left epigastric pain and jaundice with numerous elliptical RBCs on blood film. The family history review discovered jaundice in her sibling. DIAGNOSIS: A novel heterozygous mutation of SPTA1 was detected in the proband, her brother and father, c.7220_7221del:p.Tyr2407* in exon 52. Bioinformatics analysis indicated that this mutation was likely pathogenic and results in early termination of transcription and production of defective protein. INTERVENTIONS: The proband underwent splenectomy and cholecystectomy due to symptomatic splenomegaly and gallstone. OUTCOMES: After surgery, the bilirubin levels decreased to normal (i.e., total bilirubin 16.4 µmol/L; indirect bilirubin 12.3 µmol/L), and the pain and uncomfortableness in the upper abdomen relieved completely. LESSONS: We suggest that simultaneous whole exome sequencing of causative genes of all family members is a useful strategy to identify pathogenetic mutations for hereditary RBC membrane disorders, mainly in cases with an ambiguous phenotype.

[Clinical Characteristics of 223 Chinese Patients with Hemophilia in A Medical Center of Gansu Province in China].

OBJECTIVE: To analyze the clinical characteristics, diagonsis and treatment of patients with hemophilia in Gansu province of China. METHODS: The clinical data of 223 cases of hemophilia in our center between January 2010 and May 2015 were collected and analyzed retrospectively, these 223 cases of hemophilia were from 14 cities in Gansu and neighboring provinces, including 203 cases of hemophili A (HA) and 20 cases of hemophili B (HB), among them 222 cases were male, only 1 female(HA), 177 cases were from Rural areas (79.4%). RESULTS: The median age of first bleeding was 2 years old, and the average age of confirmed as hemophilia was 5.6±6.5 years, the delayed time of diagnoses of HA and HB was 2.50±4.91 and 2.07±4.76 years, respetively, among all the patients 168 caese complicated with joint hemorrhage (75.3%), 123 cases with joint deformities (55.2%). 91.6% of the patients were treated according to demand, the HBV and HCV infection rates were 1.7% and 6.2% respectively. The first-visited hospital of 86.9% patients was hospitalized below 3 grade of level, only 15.9% of these patients were considered to diagnose as hemophili. CONCLUSION: The accurate level of diagnosis rate for hemophiliacs in Gansu province is low, the delay time of diagnosis is longer, the ratios of complicated joint hemorrhage, total accumulative joint deformity were high, HCV infection rate is also high.

[Effect of Emodin Combined with AZT on the Proliferation and the Expression of BCL-2, NF-κB, TGF-ß in the Leukemia Stem Cells-KG-1a cells].

OBJECTIVE: To investigate the effect of Emodin combined with 3'-azido-3'-deoxythymidine (AZT) on the proliferation and apoptosis of concentrated leukemia stem cells (CLSC)-human acute myeloid leukemia KG-la cells and expression of BCL-2, NF-κB and TGF-ß. METHODS: The tumor stem cell-like subpopulation in human leukemia cell line KG-1a was enriched with 5-fluorouracil (5-FU). The CD34⁺ CD38⁻ subpopulation in the KG-1a cells was detected with flow cytometry, the cell proliferation was detected by MTT method to study the of Emodin and AZT in the CLSC. The cell apoptosis was analyzed by flow cytometry. The expression of NF-κB, BCL-2 and TGF-ß mRNA and proteins were measured with RT-PCR and Western blot respectively. RESULTS: As compared with cells treated with mentioned above drugs alone, the inhibition of proliferation potential and apoptosis rate of cells in combination group markedly increase with time and concentration dependent member (P < 0.01), the expression of NF-κB, BCL-2 and TGF-ß mRNA and proteins decreased. CONCLUSION: Emodin combined AZT can synergistically inhibit the proliferation, induce cell apoptosis, and down regulate the expression of NF-κB, BCL-2 and TGF-ß mRNA and proteins in the CLSC, the possible mechanism of synergistic effect may be associated with inhibiton of BCL-2 activation and down-regulation of the expression of NF-κB, and TGF-ß.

[Relationship of MPO and NQO1 gene polymorphisms with susceptibility to acute leukemia].

The aim of this study was to investigate the relationship of the gene polymorphisms of myeloperoxidase (MPO) and NAD (P) H: quinone oxidoreductase 1 (NQO1) with the susceptibility to acute leukemia (AL) in Chinese Gansu population. A 1:1 paired case-control study of 150 patients with acute leukemia and 150 cancer-free inpatients as a control was conducted to detect the polymorphisms of MPO and NQO1 by LDR techniques. The results showed that the MPO-463A genotype frequency in patient group was lower than that in control group, and there was significant difference of MPO (G-463A) genotype between patient group and control group (χ(2) = 11.828, P < 0.05, OR = 0.368, 95%CI = 0.205 - 0.610). The NQO1-609T genotype frequency in patient group was higher than that in control group, and there was significant difference of NQO1 (C-609T) genotype between patient group and control group (χ(2) = 17.931, P < 0.05, OR = 1.428, 95%CI = 1.237 - 3.339). The combined gene analysis showed that the AML risk in patients carrying the wild genotypes of MPO and NQO1 was dropped to 33.6%. It is concluded that the MPO and NQO1 gene polymorphisms are associated with susceptibility to AL. The AL risk may decrease in patients carrying MPO (G-463A) mutant gene (GA/AA), while the AL risk may increase in patients carrying NQO1 (C-609T) mutant gene (TC/TT). The combined effect of MPO and NQO1 wild genotypes may further decrease AL risk.

[JAK2V617F mutation in the patients with myeloproliferative disorder and its relation with clinical characteristics].

This study was aimed to investigate the incidence of JAK2V617F mutation in BCR-ABL negative patients with myeloproliferative disorders (MPD) and its relation with clinical characteristics of MPD. The sensitive and specific test for JAK2V617F mutation was established for improving diagnosis level in Gansu province. 47 BCR/ABL negative MPD patients and 12 healthy people were enrolled in this study. Allele specific polymerase chain reaction (AS-PCR) was used to amplify the exon 12 of JAK2 gene which harbours V617F mutation. The PCR products were identified by DNA sequencing. And its relation with clinical characteristics of MPD was analyzed also. The results indicated that the incidence of JAK2V617F positive mutation in 47 patients with BCR-ABL negative MPD was 74.5 % (35/47), including 83.9 %(26/31) in patients with polycythemia vera (PV), 60 % (9/15) in patients with essential thrombocythemia (ET), only in one patient with idiopathic myelofibrosis (IMF). In PV group, the patients with JAK2V617F positive mutation had higher counts of WBC and Plt than patients with JAK2V617F negative mutation. In ET group, the patients with JAK2V617F positive mutation had higher WBC count and Hb level than those in the patients with JAK2V617F negative mutation with tendency of suffering from complications such as hepatosplenomegaly, haemorrhage and thrombosis. It is concluded that JAK2V617F mutation is more frequent in BCR-ABL negative patients with MPD, the AS-PCR method is sensitive and specific for detection of the mutation and may successfully use in clinical examination.

[Relation of GSTP1 and CYP2E1 polymorphisms with susceptibility to acute leukemia].

This study was aimed to investigate the relation of glutathione S-transferase pI (GSTP1) and cytochrome P450 enzyme 2E1 (CYP2E1) gene polymorphisms with the susceptibility to acute leukemia (AL) in Chinese population. The GSFP1 and CTP2E1 gene polymorphisms in 150 patients with AL and 150 patients with non-hematological diseases or non-tumor as controls were detected by means of case-control paired 1:1 method and ligase detection reaction (LDR) techniques. The results indicated that the frequently of G allele and Ile/Val + Val/Val of GSTP1 gene (26.7%and 44% respectively) in AL group were higher than those in control group (10% and 16% respectively); the AL risk for persons with Ile/Val + Val/Val was 3.260-fold (95%CI = 1.527 - 5.236) of persons with Ile/Ile. The further stratified analysis showed the frequency of Ile/Val + Val/Val in AML group was higher than that in control group (55% vs 16%, p < 0.05); the AML risk for persons with Ile/Val + Val/Val was 2.214-fold (95% CI = 1.009-3.260) as persons with Ile/Ile. The frequencies of C2 allele (16.7%) and C1C2/C2C2 of CYP2E1 gene (30%) in AL group seemed higher than those in control groups (13.9% and 26%), but the difference between them was not statistical significant (p > 0.05). The further stratified analysis showed that C1C2/C2C2 of CYP2E1 gene occurred more frequently in AML group (36%) than that in control group (32%), but there was no statistical difference between them (p > 0.05). Combined genotype analysis showed that the AML risk for persons in combination of lle/Val + Val/Val of GSTP1 gene with C1C2 + C2C2 of CYP2E1 gene increased by 3.208-fold. It is concluded that the GSTP1 gene is related with susceptibility to AML, the AL risk for persons with lle/Val + Val/Val of GSTP1 gene decreased, while CYP2E1 gene is not related with susceptibility to AL, the AML risk for persons in combination of GSTP1 wildtype with CYP2E1 hybrid and mutant genotype can be further decreased.