【摘要】 目的 探讨细胞周期蛋白依赖性激酶4/6(CDK4/6)抑制剂对多发性骨髓瘤(MM)细胞免疫治疗相关分子B细胞成熟抗原(BCMA)、程序性死亡受体配体1(PD-L1)表达及MM细胞和正常微环境细胞凋亡的影响。方法 对数生长期H929细胞分为阿贝西利组、帕博西尼组、DMSO组，于细胞培养基中分别加入2μmol/L阿贝西利、5μmol/L帕博西尼和等体积DMSO处理，处理1、3、5 d时，采用流式细胞术检测3组细胞BCMA、PD-L1相对平均荧光强度。2020年1月—2023年6月中国医学科学院血液病医院诊治MM患者3例，采集患者骨髓，采用淋巴细胞分离液分离骨髓单个核细胞，分为阿贝西利组、帕博西尼组、DMSO组，分别给予1μmol/L阿贝西利、10μmol/L帕博西尼和等体积DMSO处理，采用流式细胞术检测CD138~+肿瘤细胞和CD138~-正常微环境细胞相对细胞存活率。结果 帕博西尼组处理1、3、5 d时BCMA相对平均荧光强度(1.37±0.04、2.36±0.11、3.99±0.17)均高于阿贝西利组(0.69±0.02、1.03±0.05、1.69±0.06)(P<0.05),处理3、5 d时均高于DMSO组(1.00±0.02、1.00±0.05)(P<0.05),处理1 d时与DMSO组比较差异无统计学意义(P>0.05);阿贝西利组处理5 d时BCMA相对平均荧光强度高于DMSO组(P<0.05),处理1、3 d时与DMSO组比较差异均无统计学意义(P>0.05);阿贝西利组、帕博西尼组处理1、3、5 d时BCMA相对平均荧光强度均依次增高(P<0.05)。处理1、3、5 d时，阿贝西利组、帕博西尼组PD-L1相对平均荧光强度(1.30±0.02、3.21±0.10、2.28±0.17,1.28±0.12、2.81±0.38、2.00±0.16)均高于DMSO组(1.00±0.09、1.00±0.06、1.00±0.14)(P<0.05),阿贝西利组与帕博西尼组比较差异均无统计学意义(P>0.05);阿贝西利组、帕博西尼组处理3 d时均高于处理1、5 d时(P<0.05),处理5 d时高于处理1 d时(P<0.05)。处理24、48 h时，阿贝西利组、帕博西尼组骨髓单个核细胞CD138~+相对细胞存活率均低于DMSO组(P<0.05),帕博西尼组均低于阿贝西利组(P<0.05)。处理48 h时，帕博西尼组骨髓单个核细胞CD138~-相对细胞存活率低于DMSO组、阿贝西利组(P<0.05),处理24 h时与DMSO组、阿贝西利组比较差异均无统计学意义(P>0.05);阿贝西利组处理24、48 h时骨髓单个核细胞CD138~-相对细胞存活率与DMSO组比较差异均无统计学意义(P>0.05)。结论 CDK4/6抑制剂上调MM细胞BCMA、PD-L1表达，对MM细胞有选择性杀伤作用，对正常微环境细胞无明显影响。更多还原

【Abstract】 Objective To investigate the influences of cyclin-dependent kinase 4/6(CDK4/6) inhibitors on the expressions of multiple myeloma(MM) cellular immunotherapy-associated molecules B-cell maturation antigen(BCMA) and programmed death-ligand 1(PD-L1) and on the apoptosis of MM cells and cells in normal microenvironment.Methods H929 cells in logarithmic growth phase were divided into abemaciclib,palbociclib and DMSO groups,and treated with 2 μmol/L abemaciclib,5 μmol/L palbociclib and equivalent volume of DMSO in cell culture medium,respectively.The relative mean fluorescence intensities of BCMA and PD-L1 were detected by flow cytometry on day 1,3 and 5 of culture in three groups.The bone marrow was collected from 3 MM patients diagnosed and treated in Blood Diseases Hospital of Chinese Academy of Medical Sciences from January,2020 to June,2023,and the bone marrow mononuclear cells were separated by using lymphocyte isolation solution,and divided into abemaciclib,palbociclib and DMSO groups,which were treated with 1 μmol/L abemaciclib,10 μmol/L palbociclib,and equivalent volume of DMSO respectively.Flow cytometry was used to detect the relative cell survival rates of CD138~+ tumor cells and GD138 cells in normal microenvironment.Results The relative mean fluorescence intensities of BCMA were higher in palbociclib group(1.37±0.04,2.36±0.11,3.99±0.17) than those in abemaciclib group(0.69±0.02,1.03±0.05,1.69±0.06) on day 1,3 and 5 of culture(P<0.05),and were higher in palbociclib group than those in DMSO group(1.00±0.02,1.00±0.05) on day 3 and 5(P<0.05),while there was no significant difference between palbociclib group and DMSO group on day 1(P>0.05),In abemaciclib group,the relative mean fluorescence intensity of BCMA was higher than that in DMSO group on day 5(P<0.05),and showed no significant difference on day 1 and 3compared with DMSO group(P>0.05).The relative mean fluorescence intensities of BCMA increased sequentially in abemaciclib group and palbociclib group on day 1,3 and 5(P<0.05).On day 1,3 and 5,the relative mean fluorescence intensities of PD-L1 were higher in abemaciclib group(1.30±0.02,3.21±0.10,2.28±0.17) and palbociclib group(1.28±0.12,2.81±0.38,2.00±0.16) than those in DMSO group(1.00±0.09,1.00±0.06,1.00±0.14)(P<0.05),and showed no significant differences between abemaciclib group and palbociclib group(P>0.05).The relative mean fluorescence intensities of PD-L1 were higher on day 3 in both abemaciclib and palbociclib groups than those on day 1 and 5(P<0.05),and were higher on day 5 than those on day 1(P<0.05).After 24 and 48 h of treatment,the relative cell survival rates of CD138~+ were lower in abemaciclib group and palbociclib group than those in DMSO group(P<0.05),and lower in palbociclib group than those in abemaciclib group(P<0.05).The relative cell survival rate of CD138 was lower in palbociclib group than those in DMSO group and abemaciclib group after treatment for 48 h(P<0.05),and showed no significant difference in palbociclib group after treatment for 24 h compared with DMSO group and abemaciclib group(P>0.05).Conclusion CDK4/6 inhibitors up-regulate BCMA and PD-L1 expressions in MM cells and selectively kill MM cells,with no significant influence on the cells in normal microenvironment.更多还原