Dissertation > Excellent graduate degree dissertation topics show

Experimental and Clinical Research of FLT3, NPM1 and C-KIT Gene in Acute Myeloid Leukemia by Bone Marrow Slides

Author: PanYing
Tutor: GongWuXing
School: Jinan University
Course: Internal Medicine
Keywords: Bone marrow smears FLT3-ITD gene NPM1 gene C-KIT gene Acute myeloid leukemia Gene mutation Prognosis
CLC: R733.7
Type: Master's thesis
Year: 2011
Downloads: 66
Quote: 0
Read: Download Dissertation

Abstract

Research background and purpose of acute myeloid leukemia (Acute myeloid leukemia, AML) is a group of hematological malignancies with different biological characteristics. In recent years, with the progress of the study of molecular genetics, the majority of AML can be detected abnormal gene level. FLT3 gene has an important role in the proliferation and differentiation of hematopoietic stem / progenitor cells and precursor B cells. FLT3 gene mutations lead to the proliferation and differentiation of hematopoietic cells and abnormal apoptosis, caused by the occurrence of leukemia. NPM1 gene encoding nucleolar phosphoprotein (nucleophosmin, NPM, NO38, B23 or NPM1 protein) expressed in the nucleolus, shuttling between the nucleolus and cytoplasm, involved in the transport and synthesis of ribosomal precursor centrosome replication, maintaining genome stability and DNA polymerase a activity regulation, regulating cell cycle progression and proliferation and development. In addition, NPM1 protein and P53, P19 protein interactions, inhibition of tumor plays an important role. NPM1 gene mutations can cause it to shift to the cytoplasm (alternate-reading-frame protein, Arf) inactivation of the tumor suppressor protein within the nucleolus, dependent and non-dependent p53 pathway promote leukemia cells. MW 145kD C-KIT gene encoding a transmembrane tyrosine kinase receptor, its ligand stem cell factor (stem cell factor, SCF), the SCF is one of the hematopoietic growth factors important synergy with other cytokines to promote hematopoietic stem cell proliferation and differentiation. C-KIT gene mutation leads to the C-KIT does not depend on the the spontaneous ligand receptor dimerization, caused sustained activation of C-KIT receptor, resulting in hematopoietic cell proliferation or apoptosis inhibition caused leukemia. 2011 U.S. National Comprehensive Cancer Network (National Comprehensive Cancer Network, NCCN) sent FLT3 gene, the NPM1 gene C-KIT gene mutation as the the important molecular genetics flag AML risk stratification that gene mutations in AML occurred Development, prognosis and efficacy are closely related. The topic by extracting the DNA stored bone marrow smears FLT3 gene in AML, NPM1 gene and C-KIT gene mutation analysis, to explore the relationship between the three gene mutations in AML clinical features of AML clinical stratification The prognosis of minimal residual disease detection and molecular targeted therapy provide more scientific experiments and theoretical basis. Method 1. Stored bone marrow smears of DNA using a modified phenol: chloroform: isoamyl alcohol extraction. Using the polymerase chain reaction (polymerase chain reaction, PCR) technology and agarose gel electrophoresis methods 55 patients with AML FLT3 internal tandem duplication (internal tandem duplication, ITD) mutation detection. 3 of 55 patients with AML NPM1 gene mutation detection and analysis using PCR, DNA sequencing and molecular cloning methods. 4 C-KIT gene mutation detection and analysis of 55 patients with AML using PCR, DNA sequencing and molecular cloning methods. Results 55 -20 ℃ cryopreservation without Wright's stained bone marrow smears and 10 cases and stored at room temperature Wright's stained bone marrow smears can be modified phenol: chloroform: iso-amyl alcohol method successfully extracted DNA and The extracted DNA can be used for PCR, and direct sequencing and molecular cloning and sequencing analysis. 55 AML patients detected 10 patients with FLT3-ITD positive (mutant) patients, including 9 cases of heterozygous mutations and homozygous mutations, the positive rate was 18.2%; FLT3-ITD-positive patients to M5 the majority, but the distribution of FAB subtypes showed no statistical significance (P gt; 0.05); FLT3-ITD-positive group than negative group initial remission induction treatment of complete remission (complete remission, CR) rate is low, and event-free survival (event -free survival, EFS) time and overall survival (overall survival rate, OS) time is short, and the difference was statistically significant (P lt; 0.05) between the two groups of three indicators. 55 patients with AML were amplified by PCR the reverse direct sequencing and cloning, there were nine cases of co mutant NPM1 gene heterozygosity mutation rate of 16.4% for all type A mutation in the 960 to 961 nucleoside acid inserted between TCTG (reverse complementary CAGA); NPM1 gene mutations in M2 and M5 is only found in this experiment; NPM1 gene mutation group than in the wild group newly diagnosed peripheral white blood cell count and bone marrow blasts high (P lt; 0.05 ), gender, age, hemoglobin, platelets, lactate dehydrogenase, and initial remission induction treatment the CR rate differences were not statistically significance (P gt; 0.05); EFS rate than the wild group of NPM1 mutation group of less than 10 months, 19 months of NPM1 mutation group accompanied by three cases FLT3-ITD-positive rate than the wild group OS (P lt; 0.05); 9 cases of patients with NPM1 mutations of NPM1 / FLT3-ITD. NPM1 / FLT3-ITD-, NPM1 / FLT3-ITD and NPM1 / FLT3-ITD-group of newly diagnosed peripheral blood white blood cell count to compare results: NPM1 / FLT3-ITD gt; NPM1 / FLT3-ITD-gt; NPM1-/FLT3-ITD gt; NPM1- / FLT3-ITD-(P lt; 0.05); four sets of initial remission induction treatment CR rate in order were NPM1 / FLT3-ITD-gt; NPM1-/FLT3-ITD- gt; NPM1-/FLT3-ITD gt ; NPM1 / FLT3-ITD (P lt; 0.05): four groups the proportion of bone marrow blasts and lactate dehydrogenase difference had no statistical significance (P gt; 0.05); comprehensive evaluation of six prognostic factors using Cox regression including age, bone marrow blast cell ratio NPM1 / FLT3-ITD NPM1 / FLT3-ITD-NPM1-/FLT3-ITD and NPM1-/FLT3-ITD- on AML survival of the results suggest NPM1-/FLT3-ITD (P = 0.005, RR = 1.250) risk factors affecting OS. Found 55 patients specimens outside No. 17 C-KIT gene was amplified forward sequencing exon 2 cases of C-KIT gene mutations (3.6%), found only in the experimental M2a and M3 were D816H mutant and D816V mutant; Logistic Regression Analysis on the influencing factors and AML patients in first remission induction therapy achieved CR relationship OR = 66.940, FLT3-ITD mutations as risk factors; C-KIT gene mutation in patients with FLT3, FLT3-ITD mutation positive-ITD and NPM1 mutations in patients found no overlap. Conclusion 1. Improved phenol: chloroform: isoamyl alcohol method to extract bone marrow smears DNA more successful, suitable for cryopreservation, without Wright's stain Wright's stained bone marrow smears and stored at room temperature, and extracted DNA can be used for PCR direct sequencing and molecular cloning and sequencing analysis. 2.FLT3-ITD mutations in AML patients a higher frequency of occurrence; FLT3-ITD positive patients with initial remission induction treatment CR rate, EFS and OS short time, prompted FLT3-ITD mutation poor prognosis molecular markers. The 3.NPM1 gene mutations and FLT3-ITD mutations are chance similar; the NPM1 gene A mutant of the more common type; NPM1 mutation group has a high white blood cell count, the clinical features of the proportion of bone marrow blast cells; NPM1 mutation group EFS rate in 10 months NPM1 mutations OS rate within 19 months, suggesting that NPM1 gene mutation may be a better prognostic factors. 4 The experiment found three cases of patients with NPM1 gene and gene mutation of the FLT3-ITD; combinations of two genes initial remission induction treatment CR rate is the highest for NPM1 / FLT3-ITD-group, the lowest for NPM1 / FLT3-ITD group. Cox regression analysis results for NPM1-/FLT3-ITD risk factors affect the OS time. 5.C-KIT gene mutation in a low incidence of AML in C-KIT gene have seen with the FLT3 gene overlap, NPM1 gene mutations. 6. Logistic regression analysis was prompted FLT3-ITD mutations affect AML patients in first remission induction treatment achieved CR risk factors NPM1 gene mutations and C-KIT gene mutation does not affect the CR rate.

Related Dissertations

CLC: > Medicine, health > Oncology > Hematopoietic and lymphoid neoplasms > Leukemia
© 2012 www.DissertationTopic.Net Mobile