多色流式细胞术检测微小残留病变在CD19-CAR-T桥接allo-HSCT治疗B-ALL患者监测的意义和价值  

Significance of multicolor flow cytometry in the detection of minimal residual disease in monitoring CD19-CAR-T cell bridging allo-HSCT treatment of B-ALL patients

作　　者：陈曼 赵玮 傅旻婧 王爱先 张改玲 邱丽媛 王榕 赵艳丽 张弦 王卉 Chen Man;Zhao Wei;Fu Minjing;Wang Aixian;Zhang Gailing;Qiu Liyuan;Wang Rong;Zhao Yanli;Zhang Xuan;Wang Hui(Department of Laboratory Medicine,Hebei Yanda Lu Daopei Hospital,Langfang 065201,China;Department of Pathology and Medical Laboratory,Beijing Lu Daopei Hospital,BeiJing 100176,China;Department of Clinical Microbiology and Immunology,School of Medical Laboratory,Guizhou Medical University,Guiyang 550025,China)

机构地区：[1]河北燕达陆道培医院检验科,廊坊065201 [2]北京陆道培医院病理与医学检验科,北京100176 [3]贵州医科大学检验医学院临床微生物与免疫学教研室,贵阳550025

出　　处：《中华检验医学杂志》2022年第8期813-819,共7页Chinese Journal of Laboratory Medicine

基　　金：河北省科技厅青年基金(20210890);河北省医学科学研究课题(20220963)。

摘　　要：目的探讨多色流式细胞术(MFC)监测微小残留病变(MRD)在CD19嵌合抗原受体T细胞(CD19-CAR-T细胞)治疗后桥接异基因造血干细胞移植(allo-HSCT)治疗难治、复发急性B淋巴细胞白血病(r/r B-ALL)患者过程中的意义和价值。方法收集2019年1月至7月河北燕达陆道培医院收治的r/r B-ALL患者37例,年龄15(6,19)岁,其中男24例,女13例,均为经CD19-CAR-T细胞免疫治疗桥接allo-HSCT治疗的患者。采用多色流式细胞技术以胞浆CD79a抗体设B细胞门监测患者在CART治疗前第0天、治疗后第15天、第28天及移植后骨髓、脑脊液样本、组织样本的MRD值评估肿瘤细胞残存以及肿瘤累及位置,监测CD19阳性B细胞在移植前恢复情况及CD19阳性B细胞恢复距离CAR-T细胞回输天数评估CAR-T细胞实际杀伤效果,同时进行患者各时间点外周血CAR-T细胞计数等。采用Kaplan-Meier法及Log-rank检验分析单因素累积生存差异。结果(1)37例患者中8例死亡,29例存活。5例患者移植后复发,其中复发患者4例死亡,1例存活。(2)死亡组患者在CAR-T细胞免疫治疗后到桥接移植前MFC-MRD阴性缓解率为5/8,低于存活组28/29(χ^(2)=7.540,P=0.006);回输第15天阴性缓解率为3/8,低于存活组24/29(χ^(2)=6.512,P=0.011);回输第28天阴性缓解率为3/8,低于存活组26/29(χ^(2)=10.065,P=0.002)。而死亡组伴髓外MFC-MRD阳性肿瘤浸润率为(7/8)高于存活组(14/29)(χ^(2)=3.931,P=0.047)。CAR-T细胞免疫治疗后,死亡组CD19阳性细胞恢复时间即CAR-T细胞可对CD19阳性细胞杀伤时间42.00(30.00,49.00)d短于生存组55.00(41.50,73.50)d(Z=0.022,P=0.020)。结论利用多色流式细胞术检测B-ALL患者微小残留病变发现,在CD19-CAR-T细胞免疫治疗后到桥接allo-HSCT前、回输第15天、回输第28天等时间点,阳性结果均提示预后不良。这一结果为临床进行对症治疗提供一定指导意义。Objective To investigate the significance of multicolor flow cytometry(MFC)monitoring of minimal residual disease(MRD)in the course of allogeneic hematopoietic stem cell transplantation(allo-HSCT)after CD19-chimeric antigen receptor(CAR)-T cell immunotherapy for patients with refractory,relapsed B-cell acute lymphoblastic leukemia(r/r B-ALL).Methods 37 patients with r/r B-ALL admitted to Hebei Yanda Lu Daopei Hospital from January to July 2019,aged 15(6,19)years old,including 24 males and 13 females,were treated with CD19-CAR-T cell immunotherapy bridging allo-HSCT.MFC with cytoplasmic CD79a antibody to set up B-cell gates was used to monitor patients′bone marrow(BM),cerebrospinal fluid(CSF),and tissue samples on day 0(prior to the CAR-T cell immunotherapy),day 15,day 28 post CAR-T cell immunotherapy,and post transplantation.The MRD values of these samples were analyzed to evaluate the residual tumor cells and metastasis.The killing effect of the CAR-T cells was evaluated by the recovery of CD19+B cells before transplantation and the period between the timepoint when CD19+B cells was recovered and the timepoint when CAR-T cells were infused.Peripheral blood CAR-T cells were counted at different time points.Statistic analysis was performed by Kaplan-Meie assay and Log-rank test to analyze the difference of univariate cumulative survival.Results(1)Among the 37 patients,8 died and 29 survived.5 patients relapsed after transplantation,of which 4 relapsed patients died and 1 survived.(2)MFC MRD negative remission rate of the death group was lower than that of the survival group at the following time points:post-CAR-T therapy and prior to transplantation(5/8 vs.28/29,χ^(2)=7.540,P=0.006);day 15 of the CAR-T cell reinfusion(3/8 vs.24/29,χ^(2)=6.512,P=0.011);day 28 of the reinfusion(3/8 vs.276/29,χ^(2)=10.065,P=0.002).The probability of extramedullary MFC MRD positive tumor infiltration in the death group was higher than that in the survival group(7/8 vs.14/29,χ^(2)=3.931,P=0.047).After CAR-T cell immunotherapy,the

关 键 词：白血病 B细胞 嵌合抗原受体T细胞 异基因造血干细胞移植 微小残留病变 多色流式细胞技术 

分 类 号：R733.71[医药卫生—肿瘤]