荧光原位杂交法检测造血干细胞移植后骨髓嵌合状态和微量残留病灶

目的 探讨荧光原位杂交（FISH）法在异基因造血干细胞移植（allo-HSCT）后检测供、受者骨髓嵌合状态和微量残留病灶（MRD）的作用。方法 2001年2月至2005年6月对83例患者进行allo-HSCT。对供者为异性的64例受者实施骨髓细胞性染色体着丝粒探针FISH法检测，评价供、受者骨髓嵌合状态及其动态改变；对供、受者性别相同而有特殊染色体异常的19例受者，应用相应的基因探针（BCR／ABL、AMIL1／ETO和MLL）对骨髓细胞行FISH法检测，评价微量残留病灶的发生。结果 64例异性allo-HSCT的受者中，50例供、受者骨髓嵌合度在99％以上；7例早期嵌合度偏低（96．2％～98．7％），在随访过程中逐渐上升至99％以上，移植后均未复发原病；另外7例嵌合度在随访过程中呈进行性下降，其中3例微量残留病灶在10％以上，均同时出现血液学复发；4例患者微量残留病灶在2％～5％之间，其中2例经快速停用免疫抑制剂后出现严重的移植物抗宿主病（GVHD）；1例在免疫抑制剂快速减量后骨髓嵌合度逐步上升至99．9％，目前仍处于完全缓解（CR）状态中；1例持续处于CR状态。19例供、受者性别相同的allo-HSCT受者中，16例移植后未检测到原来的异常核型；1例检测到10％的微量残留病灶，经免疫抑制剂减量4个月后降为1％，移植后1年仍在完全缓解中；2例分别在移植后1个月和4个月时出现原来的染色体异常。复查骨髓为原病复发，再次化疗未缓解。结论 应用FISH法检测allo-HSCT后骨髓嵌合状态和微量残留病灶，对判断植入、复发和指导早期干预性免疫治疗有重要意义。

Fluorescence in situ hybridization detected minimal residual disease and chimerism in patients with hematologic malignancies after allogeneic hematopoietic stem cell transplantation

Objective To explore the minimal residual disease (MRD) and cellular chimerism in patients with hematopoietic malignancies after allogeneic hematopoietic stem cell transplantation (allo-HSCT).Methods From May 2001 to June 2005,83 patients received allo-HSCT,including 55 males and 28 females. Of them 49 patients received sibling HLA-matched bone marrow transplantation (BMT),3 HLA-matched peripheral blood stem cell transplantation,8 un-related BMT,9 nonmyeloablative stem cell transplantation (NST) and 14 related haploidentical transplantation. Among them,49 patients were diagnosed as having CML,16 having AML,16 having ALL,one having multiple myeloma and one having malignant lymphoma. Chimerism and MRD were monitored by fluorescence in situ hybridization (FISH) using X and Y specific centromeric probes or gene probes for BCR/ABL,MLL and AML1/ETO. 1000 cells were analyzed for each sample.Results Among 19 patients receiving sex-matched transplant,the former chromosome rearrangements were not found in 16 patients after transplantation,MRD was detected in 10 % of cells in one patient and 1 % of cells having MRD in 4th month after the reduction of immunotherapy,and the patients were still in remission one year after transplantation. Two patients were found having the former chromosomal rearrangement 1 and 4 months after transplantation,respectively,who did not achieve remission after chemotherapy. Over 99 % donor chimerisms were found in 50 patients on day 25,donor cells were at a low level ( 96.2 % ~ 98.7 % ) in 7 patients on day 25,and increased over 99 % later. They were in remission without relapse. The donor chimerisms were decreased gradually in other 7 patients,of them 3 patients with the host cells above 10 % showed hematologic relapse. Four patients with the host cells between 2 %~5 % had different outcomes: 2 patients died of severe GVHD after the reduction of cyclosporine A,one patient got a donor chimerism over 99 % after reduction of immunotherapy,and one patient was still in complete remission.Conclusions FISH could play a pivotal role in the detection of MRD and chimerism. It is helpful to the evaluation of graft and relapse and to the guide of intervention of early immunotherapy.