Mutations And Expression Of FMS-Like Tyrosine Kinase 3 Gene In Patients With Acute Promyelocytic Leukemia And Its Clinical Significance

Background and objective:The hypothesis of "two hit" model suggests that at least two classes of gene mutations are responsible for the leukemogenesis.ClassⅠmutation involves mostly tyrosine kinase genes such as FLT3,C-Kit and BCR/ABL,which confer proliferative and/or survival advantage to hematopoietic cells.On the other hand, ClassⅡmutation involves most transcription factors such as PML/RARα,AML1/ETO and C/EBPα,which serve to impair differentiation of hematopoietic cells.Apparently,the different gene may have different influence on the clinical manifestation,the symptoms and the responses to treatment.At present,a lot of researches on the FLT3 mutation in acute myeloid leukemia and PML/RARαgene in acute promyelocytic leukemia(APL)have been done.But the effects of FLT3 gene mutation and FLT3 gene mutation accompanied the expression of PML/RARαfusion gene in APL patients on the clinical manifestation,the symptoms, the responses to treatment are still unknown in detail.Therefore,in the study,the mutations and expression of FLT3 gene,the expression of PML/RARαfusion gene in APL patients,as well as the relationship between the gene mutation and clinical manifestation,symptom, therapeutic effects were systematically studied.Methods:Fresh bone marrows from 34 APL patients who had been admitted to Hematology Department of Xiangya Hospital from April 2007 to March 2008 were aspirated.Genomic DNA and total RNA of white blood cells were then isolated followed by analyzing the gene mutations of FLT3/ITD,FLT3 D835 and gene expression of FLT3 and PML/RARαfusion gene mRNA by the methods of polymerase chain reaction(PCR),reverse transcription PCR(RT-PCR),enzyme digestion and nested RT-PCR,respectively.Based on these results,the relationship between FLT3 gene mutation,the expression of PML/RARαfusion gene in APL patients and the patient' s clinical manifestation,symptom and therapeutic effect were analyzed.Results:In 34 APL patients,4 patients(11.8%)showed FLT3/ITD mutation,and FLT3 D835 point mutation was found in 4 patients(11.8%). Among these patients,both FLT3 ITD and D835 mutations were co-existed in 2 patients.RT-PCR results indicated that 30 of 34 patients had the expression of FLT3 gene mRNA,the expression positive rate was 88.3%,and 22 out of 34 patients exhibited the expression of PML/RARαfusion gene mRNA,the expression positive rate was 64.7%,of which 15 patients expressed the long-subtype(L-type)of PML/RARαfusion gene,the remain seven patients expressed short-subtype(S-type)of PML/RARαfusion gene.In 4 patients with FLT3/ITD mutation,3 patients expressed S-type of PML/RARαfusion gene,and the remain one patient expressed L-type of PML/RARαfusion gene.In another 4 patients with FLT3 D835 point mutation,one patient showed L-type of PML/RARαfusion gene expression,2 patients expressed S-type of PML/RARαfusion gene,and the remain one patient expressed neither L-type nor S-type of PML/RARαfusion gene.Statistical analysis results showed that the positive rate of patients with FLT3/ITD mutation who had higher initial peripheral white blood cell(WBC)count(WBC≥10×10~9/L)were higher than those that of patients without FLT3/ITD mutation,whose the differences was significant(P＜0.05).The positive rate of patients with FLT3 gene mutation accompanied PML/RARαfusion gene expression who had higher initial peripheral WBC count were higher than those that of patients with PML/RARαfusion gene expression alone,which had statistical differences(P＜0.05).However,the positive rate of patients who had higher percentage of blast cells plus premyelocyte cells in bone marrow(blast cells plus premyelocyte cells in bone marrow≥80%), hemorrhage symptom B-grade,and complete remission rate had no statistical differences between the patients with FLT3/ITD mutation or the patients with FLT3/ITD mutation accompanied PML/RARαfusion gene expression and the patients without FLT3/ITD mutation or the patients with PML/RARαfusion gene expression alone(P＞0.05).Furthermore, the positive rate of patients who had initial peripheral WBC count,higher percentage of blast cells plus premyelocyte cells in bone marrow, hemorrhage symptom B-grade,and complete remission rate had no statistical differences between the patients with FLT3 D835 mutation and the patients without FLT3 D835 mutation(P＞0.05)Conclusion:FLT3 mutations could be frequently identified in patients with APL,the incidence rate of FLT3/ITD and FLT3 D835 point mutation is similar.FLT3/ITD and FLT3 D835 mutation can exist in a same patient with APL.Meanwhile,majority of patients with APL can express FLT3 mRNA.Additionally,most APL patients with FLT3/ITD gene mutation and APL patients with FLT3 gene mutation accompanied the mRNA expression of PML/RARαfusion gene usually present higher initial peripheral WBC count,but there is no statistical differences in the percentage of bone marrow blast cells plus premyelocyte cells, hemorrhage symptom and complete remission rate between the APL patients with FLT3/ITD gene mutation or APL patients with FLT3 gene mutation accompanied the mRNA expression of PML/RARαfusion gene and the patients without FLT3/ITD mutation or the patients with PML/RARαexpression alone.Similarly,there is no statistical differences in initial peripheral WBC count,the percentage of bone marrow blast cells plus premyelocyte cells,hemorrhage symptom and complete remission rate between the APL patients with FLT3 D835 mutation and APL patients without FLT3 D835 mutation.