Idh1 Mutations In Acute Myeloid Leukemia Analysis Sp1 Interact With Aml1 - Eto Role On The Pathogenesis Of Leukemia

Background:Acute myeloid leukemia (AML) presents great heterogeneity in phenotypes and gene profile. The genetic diversity of AML results mainly from mutant genes that are causally implicated in the pathogenesis of AML. Somatic mutations in the IDH1gene encoding cytosolic NADP+-dependent isocitrate dehydrogenase have been shown in majority of gliomas. These mutations occur at a single amino acid residue of IDH1, arginine132. Recently, IDH1R132mutations were found to be recurrent gene alterations in AML. However, the clinical and biological features of IDH1-mutated AML and how the mutations impact the prognosis have not been identified.Objective:To detect the IDH1R132mutations in adult AML patients, characterize the clinical and biological features of IDH1-mutated AML, and reveal how the IDH1R132mutations impact the prognosis of AMLMethods:Bone marrow samples of410newly diagnose AML patients from Oct.2004to Mar.2010were collected. We isolated the mononuclear cells and extracted the genome DNA. The hotspot mutations in IDH1were prescreened by endonuclease-based detection and identified by cloning into vectors and sequencing. FLT3-ITD was detected by PCR and appearance of abnormally larger segment(s). The clinical information was collected and the datas were analyzed with SPSS software.Results:In the total410AML patients, we found24patients with IDH1R132mutations(5.85%). R132H and R132G were predominant in four mutation types. Besides, we found one homozygous mutant which has never been reported. IDH1mutation had no association with any subtype of FAB and any distict karyotype. IDH1-mutated and wild-typed patients had no significant difference in blood cell counts, percentage of bone marrow blasts, expression of immunophenotypes and the complete remission rate following the first induction. We discovered the existences of IDHl mutations in M2b and M3subtypes carrying distinct fusion genes which were not found in other studies. The frequency of IDH1mutation in M2b was12%and higher than5%of wild-typed patients, although the difference was not statistically significant. The frequency of FLT-ITD3in IDH1-mutated patients was higher than that of wild-typed patients, also, the difference was not statistically significant. However, this result was different form other studies in which FLT3-ITD occurred less frequently in IDH1-mutated patients. Therefore, in this study, IDH1-mutated AML presented no distict clinical and biological characteristics. Background:AML1-ETO fusion protein is observed in approximately12%of acute myeloid leukemia. AML1upregulates a number of target genes critical to normal hematopoiesis, whereas the AML1-ETO fusion protein interferes with this trans-activation. Also, AML1-ETO can interact with other transcription factors involved in hematopoietic cell differentiation, such as C/EBPa and PU.1. AML1-ETO was found to inhibit Spl which is a transcription factor involved in apoptosis and hematopoietic cell differentiation, and bind to Spl through its RUNT domain. It was proposed that the inhibition of Spl transactivity promotes leukemogenesis by blocking cell differentiation and apoptosis.Objective:Overexpress Spl in leukemic cell line expressing fusion protein AML1-ETO and overcome the inhibition of AML1-ETO in order to reveal how Spl impacts cells and how the inhibition of Spl by AML1-ETO impacts cells.Methods:Recombinant lentiviral vectors pCDH-Sp1containing the open reading frame (ORF) of Sp1was constructed to transfer Sp1into leukemic cells and overcome the inhibition of AML1-ETO. After infection with pseudovirus, the cells were collected and stained at exact time points to detect the apoptosis and differentiation status.Results:Utilizing pCDH lentivirus system, we succeeded in generating pCDH-Spl. Cells expressing Green fluorescence were observed after infection. And the overexpression of Spl protein induced apoptosis and promoted differentiation of Kasumi-1cell in vitro.