The Clinical Significance And Mechanism Of BCAT1 In Acute Myeloid Leukemia

BackgroundAcute myeloid leukemia(AML)is a kind of malignant desease originating from hematopoietic stem cells and precursor cells in the bone marrow.It is the most common hematologic malignancy in adults,with rapid progression and poor prognosis.The5-year survival rate of AML is only about 20%.With the development of science and accumulation of clinical data,there are some targeted drugs for common mutation sites in AML,such as FLT3-ITD inhibitors,IDH1 inhibitors,IDH2 inhibitors,etc.However,AML is highly heterogeneous,the current prognostic assessment system still needs to be improved,and the development of new molecular markers and therapeutics is important.Currently,with the rise of metabolomics research,key molecules related to metabolic abnormalities and reprogramming in AML are expected to be potential targets for prognostic assessment and stratified treatment of AML.Branched-chain amino acid transferase 1(BCAT1)is a key enzyme that catalyzes the catabolism of branched-chain amino acids and plays an important role in regulating intracellular metabolic homeostasis.BCAT1 expression has been reported to be closely associated with the development and progression of a variety of tumors and significantly affects the prognosis of patients.However,the mechanism of BCAT1 in AML is not yet clear.In this study,we will focus on the clinical significance and mechanism of BCAT1 in AML.Objective and method(1)Using q-PCR technology to detect the BCAT1 m RNA expression level of bone marrow mononuclear cells in cytogenetically normal AML patients.Analyze the relationship between BCAT1 m RNA expression level and clinical characteristics such as age,gender,peripheral blood white blood cell count,hemoglobin amount,platelet count,bone marrow blasts,gene mutation type,treatment plan,treatment effect,survival time and other clinical characteristics of AML patients at the time of onset;The Kaplan-Meier method and log-rank test were used to compare the survival time of AML patients in the BCAT1 high and low expression groups.(2)Construct BCAT1 knockout/overexpression AML cell,study the effect of BCAT1 expression level on cell proliferation,cycle,apoptosis and other biological behaviors,and Western blot to detect changes in related protein levels.(3)BCAT1 knockout/overexpression/control AML cell lines were injected into the tail vein of mice to construct xenograft models,study the effect of BCAT1 expression level on the tumor burden and survival time of mice.(4)To study the effect of BCAT1 expression level on DNA damage and histone methylation level of AML cells.(5)Explore the changes in the sensitivity of BCAT1 high-expressing AML cells to PARP inhibitor BMN673 and BCL-2 inhibitor Venetoclax.(6)To study the combined effect of BMN673 and Venetoclax in vitro: use BMN673 and Venetoclax as a single agent or combination to AML cell lines and primary patient cells to detect their growth inhibitory effects and calculate the combined index.(7)To study the therapeutic effect of BMN673 combined with Ventoclax on AML animal models: mice were injected with the THP-1-luc cell line overexpressing BCAT1 through the tail vein to construct a mouse xenograft model,and the mice were randomly divided into groups after tumor burden.Placebo,BMN673,Venetoclax monotherapy and BMN673 combined with Venetoclax treatment.Observe the tumor burden,survival time,and weight change of mice.Result(1)The overall survival(OS)time and event-free survival(EFS)time of cytogenetically normal AML patients in BCAT1 high expression group were significantly shorter than those in BCAT1 low expression group;high expression of BCAT1 was an independent prognostic factor forcytogenetically normal AML patients.(2)Knockout of BCAT1 inhibits the proliferation of AML cells,affects the clonogenic ability of AML cells,down-regulates the expression of G0/G1 phase checkpoint proteins CDK2,CDK4,CDK6 and cyclin D1,and blocks cells in G0/G1phase;vice versa,Overexpression of BCAT1 promotes the proliferation of AML cells and enhances the cloning ability of AML cells.(3)Knockout of BCAT1 reduces the tumor burden of mice and prolongs the overall survival time of mice;overexpression of BCAT1 increases the tumor burden of mice and shortens the survival time of mice.(4)High expression of BCAT1 increases the baseline level of DNA double-strand damage in AML cells and the level of histone methylation.(5)AML cells with high expression of BCAT1 increases cells’ sensitivity to PARP inhibitor BMN673 and BCL-2 inhibitor Venetoclax.(6)BMN673 combined with Venetoclax can significantly inhibit the growth of AML cells,and the combined effect is more significant in BCAT1 high-expressing cell lines/primary cells.(7)Compared with single drug,BMN673 combined with Venetoclax can significantly reduce the tumor burden of AML mice and prolong the survival time of mice.ConclusionThe OS and EFS of AML patients in the BCAT1 high expression group were significantly shorter than those in the low expression group.High BCAT1 expression is an independent risk factor for poor prognosis in cytogenetically normal AML patients;BCAT1 functions an oncogene in AML;high BCAT1 expression impair intracellular aKG level,affects the level of DNA damage and histone methylation of AML,thereby enhancing the sensitivity of AML cells to the PARP inhibitor BMN673 and the BCL-2inhibitor Venetoclax;in vivo and in vitro experiments,BMN673 combined with Venetoclax can significantly inhibit the growth of AML cells with high expression of BCAT1 and can be used as an effective treatment strategy for patients with high expression of BCAT1.