流式细胞术检测骨髓微量神经母细胞瘤细胞在临床中的应用  

The detection of minimal residual disease with flow cytometry and the relationship between prognosis and minimal residual disease in bone marrow in advanced neuroblastoma

作　　者：蔡娇阳[1] 汤燕静[1] 蒋黎敏[1] 宋得莲[1] 潘慈[1] 陈静[1] 董璐[1] 周敏[1] 薛惠良[1] 顾龙君[1] 汤静燕[1] 

机构地区：[1]上海儿童医学中心血液肿瘤科,上海200127

出　　处：《中国循证儿科杂志》2006年第3期193-198,共6页Chinese Journal of Evidence Based Pediatrics

摘　　要：目的建立神经母细胞瘤（NB）患儿骨髓（BM）微量肿瘤病灶（MRD）的检测方法，分析其结果与预后的关系。方法应用CD45^FITC／CD81^PE／CD56^PECy5单抗组合通过流式细胞术（FCM）检测人NBTGW细胞株的免疫表型，并确定本实验室FCM的敏感度；比较Ⅲ～Ⅳ期NB患儿治疗前及治疗中BM细胞形态学观察与FCM检测BM中的CD45^-／CD81^＋／CD56^＋细胞阳性率，分析病程中MRD检测与预后的关系。结果①TGW细胞株表达CD56和CD81，不表达CD45，本实验室FCM敏感度为1×10^-4。②58例患儿共144份标本中BM细胞形态学观察分析提示BM转移者23份，FCM检测显示表达CD45^-／CD81^＋／CD56^＋细胞的标本数62份，BM细胞形态学观察和FCM法阳性检出率差异有统计学意义（P〈0．01）。③比较平均化疗4个疗程后BMMRD结果与预后关系：31例患儿初诊时BMMRD检测阳性，化疗4个疗程后11例患儿BMMRD转阴，随访至今无复发及进展，无瘤生存率（DFS）中位时间23个月；另20例患儿化疗4个疗程后BMMRD仍为阳性，其中11例患儿复发或进展，包括1例死亡，两组差异有统计学意义（P〈0．01）。④化疗后移植患儿外周血干细胞（PBSC）采集前BMMRD结果与预后的相关性：共有19例患儿接受化疗后行PBSC移植，14例患儿采集PBSC时BMMRD为阴性，其中2例患儿复发，自移植后始DFS中位时间9个月；5例患儿采集PBSC时BMMRD为阳性，均复发，两组差异有统计学意义（P〈0．05）。结论运用FCM检测NB患儿BMMRD敏感度高、特异性强。可协助NB患儿的诊断和评估临床疗效，BMMRD残留与NB预后相关。Objective To establish a method to investigate whether flow cytometry detection of minimal residual disease （MRD） in bone marrow （BM） could predict prognosis and clinically guide the risk-adjusted therapy for neuroblastoma （NB） patients. Methods Human TGW NB cell line was used to detect the immunophenotype of NB cells by flow cytometry （FCM）. The FCM sensitivity of the cocktail of CD45^FITC/CD81^PE/CD56^PECy5＋ Was tested in our lab. The results of MRD with FCM were used to evaluate neuroblastoma metastasis in BM at diagnosis and during chemotherapy. Results ①The TGW cell line expressed CD56 and CD81 antigen and did not express CD45. The FCM sensitivity in our lab was 1/10^4. ②In 144 samples from 61 patients, NB cells were found in 23 of them morphologically（ BM smear）. All of those 23 samples were CD45^-/ CD81^＋/ CD56^＋ positive by FCM. Thirty-nine BM samples were cytologically negative by BM smear but CD45^-/CD81^＋/CD56^＋ positive by FCM. There was statistically significant difference between the two methods （P 〈 0.01）. ③ BM samples from 31 patients were positive for neuroblastoma by FCM at diagnosis, eleven of them became negative after average 4 courses of chemotherapy. All of those 11 patients remained alive without evidence of disease （median23mos,range8-38 months）. In contrast, twenty patients whose BM samples were still positive, when they received the same chemotherapy, eleven of them were relapsed and 1 patient was died of disease（ medianl7.5 months ,range6-48 months）. There was statistically significant difference in disease-free survival between the two groups（ P 〈 0. 01 ）. ④MRD in BM was tested by FCM before PBSC harvest for 19 advanced neuroblastoma patients. Fourtheen was negative, two of them were relapsed. Eleven patients remained alive without evidence of disease （ median 9 months after transplant,range 4-30 months）. In contrast, other 5 patients whose BM remained positive before PBSC harvest, all of them were relapsed（median 10. 5mo

关 键 词：神经母细胞瘤 流式细胞术 微量残留病灶 预后 

分 类 号：R739.4[医药卫生—肿瘤]