Neutrophil Elastase Promotes APL Progression Via Proteolytic Cleavage Of The Transcription Factor P200CUX1

Objectives:Neutrophil elastase?NE?is a serine protease encoded by the ELANE gene.The present study aimed to elucidate the specific role of NE in the proliferation and differentiation of APL cells,its effects on the oncogenicity of APL-NOD/SCID mice in vivo,and the underlying molecular mechanisms.Methods:NE expression was detected in bone marrow samples from APL patients and analyzed in the BloodSpot database.Recombinant lentiviral-transfected NB4 and U937 cells were used to investigate the role of NE.CCK-8 assay and flow cytometry were used to assess cell proliferation and cell cycle distribution,respectively.The co-localization and interaction of NE and p200 cut-like homeobox 1?CUX1?were evaluated by immunofluorescence,co-immunoprecipitation,and Duolink?in situ proximity ligation assay.The expression levels of proliferation and differentiation markers,PCNA,Ki-67,CD11b,C/EBP?,C/EBP?,CyclinD1,Cyclin E1,Cyclin A2,P21^Waf1/Cip1,and P27^Kip1,were measured by qRT-PCR and western blot.Finally,the general appearances,body weight changes,survival time,bone marrow and peripheral blood cell morphology,hepatosplenomegaly and hepatosplenic leukocyte infiltration were observed in NB4-NE and NB4-NC NOD/SCID mice to evaluate the effect of NE on the oncogenicity of APL cells in vivo.Results:NE was highly expressed in APL bone marrow and blood neutrophils.NE overexpression promoted the proliferation and inhibited the differentiation of NB4 cells,whereas NE knockdown and its specific inhibitor achieved the opposite results in U937 cells.Mechanistically,NE interacted with and effectively hydrolyzed the transcription factor p200CUX1.Rescue experiments revealed that p200CUX1 upregulation reversed the functional influence of NE on NB4 cell proliferation and differentiation.Furthermore,NE enhanced the oncogenicity of APL cells in NOD/SCID mice and shortened the survival time.Conclusions:Neutrophil elastase regulates the proliferation and differentiation of APL cells by hydrolyzing the transcription factor p200CUX1,thus promoting the progression of the disease.NE is a novel and promising therapeutic target for APL treatment.