Detection of Minimal Residual Disease in Childhood Acute Lymphoblastic Leukemia After Termination of Therapy

Using 1gH and TCRy gene rearrangements as gene markers, we detected minimal residual disease (MRD) by means of the polymerase chain reaction (PCR) and restriction analysis. Of 18 children with acute lymphoblastic leukemia (ALL), MRDs were detected in 9 patients after termination of therapy. All 18 patients had been followed for 1.5 to 102 months after detection. Three of the nine MRD-positive patients relapsed within 3 to 6 months; none of the nine MRD-negative patients relapsed. We suggest that MRD negativity at the end of therapy might be an important factor for long-term disease-free survival, because the negative cases had a very low risk of relapse. Because the outcome for MRD-positive cases is more difficult to evaluate, patients with MRD after termination of therapy should be monitored.     

Minimal Residual Disease and Childhood Leukemia: Standard of Care Recommendations From the Pediatric Oncology Group of Ontario MRD Working Group

Minimal residual disease (MRD) is an independent predictor of relapse risk in children with leukemia and is widely used for risk‐adapted treatment. This article summarizes current evidence supporting the use of MRD, including clinical significance, current international clinical practice, impact statement, and recommended indications. The proposed MRD recommendations have been endorsed by the MRD Working Group of the Pediatric Oncology Group of Ontario and provide the foundation for a strategy that aims at equitable access to MRD evaluation for children with leukemia.     

儿童急性淋巴细胞白血病微小残留病与复发的相关分析

目的 分析小儿急性淋巴细胞白血病(急淋)缓解时和缓解不同时期微小残留病(MRD)的水平与复发的相关性。方法：用极限稀释定量PCR法和巢式PCR法追踪检测MRD，数据处理用Kaplan Meier方法及COX回归模型等。结果：46例急淋患儿缓解时MRD值与骨髓复发呈正相关(r=0.4396，P<0.01)，骨髓复发组MRD值为7.359×10-3，与未复发组MRD值(3.954×10-4)差异有显著性(P<0.01)；缓解期MRD持续阳性或由阴性转为阳性者，骨髓复发的相对危险度明显增高(P<0.05)。结论：缓解时MRD水平及缓解期MRD定性结果可作为估计急淋预后的一个重要指标，动态追踪检测MRD是指导治疗和预防复发的有效手段。     

WT1基因在急性白血病患儿中的表达及其临床意义

目的探讨WT1基因在急性白血病患儿中的表达及其临床意义。方法采用实时定量RT-PCR方法检测198例急性白血病患儿(男124例,女74例;年龄1~13岁)WT1基因的表达水平。应用SPSS 13.0软件进行统计学分析。结果急性白血病患儿WT1基因的中位数为932.99,对照组儿童为38.50,病例组显著高于对照组;ALL患儿WT1基因的中位数为195.73,AML患儿WT1基因的中位数为6 297.75,ALL患儿与急性髓性白血病(AML)患儿之间存在统计学差异(P<0.01),且在AML患儿FAB分型中M3、M4表达水平相对较高(中位数分别为8 081.83和8 123.54);WT1基因在急性白血病初诊组及复发组患儿中表达水平较高(WT1基因中位数分别为932.99和2 840.54),而完全缓解组表达水平显著下降(WT1基因中位数为144.32);可进行临床疗效评估的97例急性白血病患儿中,第1个疗程后完全缓解组与未缓解组初诊WT1基因表达水平差异有统计学意义,其中位数分别为311.98和3 768.43;对3例复发患儿进行WT1基因表达水平动态监测,发现复发前WT1的表达水平均有上升趋势。结论WT1基...     

Value of Routine Bone Marrow Examination for Detection of Bone Marrow Relapse in Children with Standard Risk Acute Lymphoblastic Leukemia

The value of routine bone marrow examination (RBME) in children during and after treatment for standard risk acute lymphoblastic leukemia (SR-ALL) was Investigated. The clinical symptoms and peripheral blood findings at the time of bone marrow relapse of 28 children were reviewed and compared with those of 28 matched controls in continuous complete remission. Five (45%) children with bone marrow relapse during maintenance therapy and six (35%) after cessation of cytostatic treatment were asymptomatic at the time of relapse. Signs indicative of relapse duriny treatment were lymphoblast cells in the peripheral blood, thromhocytopenia, hepatomegaly, anemia, or leukopenia in decreasing order of frequency. Afer cessation of treatment these signs were lymphoblasts in the peripheral blood, hepatomegab, splenomegaly, thrombocytopenia, or leukocytosis. Except for one case with thrombocytopenia, no signs suspicious for relapse were found in the control groups. When each sign was evaluated separately only the presence of lymphoblasts in peripheral blood and hepatomegaly were significant symptoms for relapse after cessation of treatment. The mean percentage of lymphoblasts in the bone marrow at the time of relapse was significant& lower for patients with an unpredicted relapse (46.8%) than patients with clinical and/or laboratory evidence of relapse (79.5 %). When lymphoblasts were present in the peripheral blood the percentage of lymphoblasts in the bone marrow was always more than 40%, both during and after cessation of treatment. These data suggest a relation between clinical and laboratory symptom and progression of the disease. It is concluded that 467% of relapses are detected by RBME in the absence of clinical or laboratory symptoms. This early detection may have a positive prognostic influence with more effective treatment for relapsed ALL.     

Comparison of Autologous and Allogeneic Bone Marrow Transplantation in Children with Acute Leukemia

Thirty-one patients with acute non-lymphocytic leukemia (18 patients) or with high-risk refractory acute lymphocytic leukemia (13 patients) underwent bone marrow transplantation between March 1980 and March 1990. The high-dose conditioning regimen employed included cyclophosphamide followed by fractionated total body irradiation (12 GY). Fourteen patients who had an HLA-identical sibling donor received allogeneic bone marrow transplantation (ailo-BMT); the other 17 patients received autologous bone marrow transplantation (auto-BMT) purged with 4-hydroperoxycyclophosphamide (4HC). Four of the 14 allo-graft recipients died of leukemic relapse and 2 others died of graft-versus-host disease. Three of the 17 auto-graft recipients died of relapse and 1 suffered relapse in the testes. The actuarial risk of relapse was 29% for the allo-BMT patients and 24% for the auto-BMT patients (P<0.05). The event-free survival rate at five years was 57% and 74% respectively (P<0.05). Although there was no difference between them, a trend toward a higher survival rate and a lower mortality and morbidity was observed in the auto-BMT group. These results suggest that autologous bone marrow transplantation purged with 4HC is an effective and useful treatment for children with acute non-lymphocytic and lymphocytic leukemia who have no HLA-identical donor.     

儿童急性白血病WT1基因表达的临床意义

目的 构建WT1质粒,建立实时定量聚合酶链反应(RQ-PCR)监测儿童白血病骨髓样本WT1基因的实验方法,初步研究WT1基因在儿童白血病中的表达特点及临床意义.方法 设计合成WT1基囚引物探针,利用分子克隆方法构建含有目的 基因WT1片段的质粒,并以此作为阳性对照检测儿童白血病骨髓样本.通过Taqman荧光探针RQ-PCR方法分别对WT1目的 基因及GAPDH内参基因进行检测,利用双标准曲线法对WT1基因的表达水平进行绝对定量,检测结果用WT1与GAPDH基因拷贝数比值(WT1/104·GAPDH)表示.实验结果应用SPSS 13.0软件进行统计学处理.结果 经酶切电泳和测序分析证实构建的质粒中成功插入了WT1基囚片段,作为RQ-PCR方法中的阳性对照;成功建立了WTl摹因的RQ-PCR检测方法.对儿童白血病标本的检测结果显示:初发AML患儿WT1基因表达水平(9.2331±37.6228)明显高于正常对照组(0.0482±0.0348),其次为BCR-ABL阳性的CML患儿(0.1080±0.0616),初发ALL患儿WT1基因表达水平(0.0605±1.6124)与正常组的差异无统计学意义.缓解中的AML患儿WT1基因表达水平(0.3150±0.7216)较初发时明显下降(P=0.000).结论 实验构建的WT1质粒及建立的RQ-PCR方法可用于儿美童白血病WT1基因检测.WT1基囚的表达水平在一定意义上可作为儿童AML治疗过程中肿瘤负荷的动态监测指标.     

急性白血病患儿血浆基质细胞衍生因子-1及其受体CXCR4表达的临床意义

目的探讨儿童急性白血病(AL)血浆基质细胞衍生因子-1(SDF-1)的水平和骨髓细胞表面CXCR4的表达及其临床意义。方法收集50例AL患儿、20例健康儿童和10例非恶性血液病患儿(对照组)。采用ELISA法分别检测AL患儿初诊时、缓解6个月、复发时和对照组儿童外周血浆SDF-1水平。用流式细胞仪检测50例AL患儿初诊时和10例非恶性血液病患儿骨髓细胞表面CXCR4的表达。结果1.AL初诊组和缓解组血浆SDF-1水平明显高于对照组(Pa<0.01),且初诊组高于缓解组(P<0.01);急性淋巴细胞白血病(ALL)组血浆SDF-1水平高于急性髓细胞白血病(AML)组(P<0.01);SDF-1水平在ALL和AML各亚型之间差异无统计学意义;复发的4例患儿,其血浆SDF-1水平在初诊时和复发时均高于缓解时(Pa<0.05);髓外浸润组SDF-1水平与非髓外浸润组比较差异无统计学意义(P>0.05)。2.AL患儿骨髓细胞表面CXCR4的相对荧光强度明显高于对照组(P<0.01);ALL组高于AML组(P<0.01);CXCR4在ALL的L1组与L2组的表达,无统计学差异,但T-ALL组高于B-ALL组(P<0.05...     

WT1基因在恶性造血系统疾病儿童中的表达及其意义

目的探讨WT1基因在儿童恶性造血系统疾病中表达的意义。方法采用巢式RT-PCR观察104例恶性造血系统疾病患儿WT1基因表达的相对水平;其中男69例,女35例;年龄1.5～14.0岁;同期检测72例成人恶性造血系统疾病患者作比较,选择12例健康体检者及10例非恶性造血系统疾病患儿骨髓标本或外周血作阴性对照(男9例,女13例;年龄3～12岁)。结果68例急性白血病(AL)患儿中WT1阳性表达49例(72.1%),3例慢性粒细胞白血病(CML)表达阴性,23例恶性淋巴瘤中7例阳性(30.4%),10例骨髓增生异常综合征(MDS)中阳性3例(30.0%),AL与MDS、CML及恶性淋巴瘤WT1阳性表达率比较差异均有显著性意义(Pa<0.01);而在健康体检者及造血系统非恶性造血系统疾病患儿中均表达阴性。各组与成人患者比较无年龄上差异(Pa>0.05),WT1基因表达在ALL均阴性,与急性粒细胞白血病(ALL)比较无显著性差异,在MDS中难治性贫血组显著低于原始细胞增多性难治性贫血及转化型原始细胞增多性难治性贫血(Pa<0.05)。结论WT1基因在恶性造血系统疾病患者中高表达,可作为检测AL微小残留及预测复发的肿瘤标志物;其与MDS疾病进程关系密切,并可作为对MDS疾病发展的高危评估指标之一。     

Immunohistochemical Detection of Post-Therapy Residual Testicular Lymphoblasts in Childhood Acute Lymphoblastic Leukemia (All)

The aim of this study was to evaluate the diagnostic value of immunohistochemistry with monoclonal antibodies (MoAbs) in detecting residual blast cells in testicular biopsies from children with acute lymphoblastic leukemia (ALL). In a prospective study of 26 patients, testicular biopsies were performed after completion of therapy, and the average follow-up after biopsies was 29 months. After immunostaining, seven patients with negative biopsies on routine histology showed scattered, strongly calla-positive cells as well as cells reacting with anti-B (CD22) MoAb. Among these seven patients with residual blast cells, four had relapsed either in testes (n = 1), bone marrow and testes (n = 1), or in the bone marrow (n = 2). In contrast, among the 15 patients without residual blast cells, all but 1 remained in complete remission. In four other cases no definite conclusion was possible after immunohistochemical study. Four testicular biopsies from patients with occult infiltration were used as positive controls. Negative controls consisted of testicular biopsies from children with testicular ectopia and postmortem testicular tissue specimens. Results suggest that the risk of relapse is significantly higher in patients with positive immunohistochemical findings indicating persistent residual blast cells. However, the predictive value of these findings requires confirmation on a larger number of cases to have therapeutic implications.     

Immunohistochemical Detection of Post-Therapy Residual Testicular Lymphoblasts in Childhood Acute Lymphoblastic Leukemia (All)

The aim of this study was to evaluate the diagnostic value of immunohistochemistry with monoclonal antibodies (MoAbs) in detecting residual blast cells in testicular biopsies from children with acute lymphoblastic leukemia (ALL). In a prospective study of 26 patients, testicular biopsies were performed after completion of therapy, and the average follow-up after biopsies was 29 months. After immunostaining, seven patients with negative biopsies on routine histology showed scattered, strongly calla-positive cells as well as cells reacting with anti-B (CD22) MoAb. Among these seven patients with residual blast cells, four had relapsed either in testes (n = 1), bone marrow and testes (n = 1), or in the bone marrow (n = 2). In contrast, among the 15 patients without residual blast cells, all but 1 remained in complete remission. In four other cases no definite conclusion was possible after immunohistochemical study. Four testicular biopsies from patients with occult infiltration were used as positive controls. Negative controls consisted of testicular biopsies from children with testicular ectopia and postmortem testicular tissue specimens. Results suggest that the risk of relapse is significantly higher in patients with positive immunohistochemical findings indicating persistent residual blast cells. However, the predictive value of these findings requires confirmation on a larger number of cases to have therapeutic implications.     

儿童急性髓细胞白血病WT1基因表达及其临床意义

目的研究急性髓细胞白血病(AML)患儿WT1基因表达情况及其与临床预后的关系。方法采用实时荧光定量PCR方法检测45例非M3儿童AML的WT1表达水平,并回顾性分析其与AML预后的关系。结果骨髓幼稚细胞比例60%的AML患儿WT1表达水平高于幼稚细胞≤60%的患儿(P0.05)。M2病例组初诊时WT1表达量低于非M2病例组(P0.05)。完全缓解组患儿的WT1表达量显著低于初诊组和复发组(P0.01)。诱导化疗结束时WT1高表达组的2年无病生存率(DFS)低于WT1低表达组(P0.05)。诱导化疗结束时WT1下降程度≥1个数量级病例组的2年总生存率(OS)和DFS高于WT1下降程度1个数量级病例组(P0.05)。AML患儿骨髓复发2~3个月前WT1表达呈上升趋势。结论 WT1表达水平与儿童AML预后密切相关,动态监测WT1表达在指导儿童AML个体化治疗、预后评估和复发预测方面具有重要临床应用价值。     

Phagocytic Macrophages in the Bone Marrow Biopsies of Children with Hodgkin's Disease

Phagocytic macrophages in bone marrow aspirates have been described as normal and are frequently observed in autoimmune disorders. They are rarely seen in bone marrow biopsies. We observed phagocytosis of leukocytes and nuclear debris by macrophages in the bone marrow biopsies in 20 of 26 children with Hodgkin's disease before therapy.

In contrast, phagocytic activity was present in only 1 of 16 children with solid tumors and 4 of 17 children receiving chemotherapy for neoplasia other than Hodgkin 's disease. In all groups the marrow was not directly involved by tumor. The presence of macrophage activity did not correlate with clinical stage or histological type of Hodgkin's disease or with the peripheral blood count. Its increased frequency in patients with Hodgkin's disease may reflect abnormal macrophage function in those patients.     

早期微量喂养对新生儿肺透明膜病临床康复的影响

目的 观察早期微量喂养在新生儿肺透明膜病(HMD)治疗中的效果.方法 收集2009年1月-2010年12月在本院NICU住院的HMD新生儿的临床喂养情况,分别统计实行早期微量喂养(早期微量喂养组)和常规喂养(常规喂养组)新生儿的一般资料,观察2组患儿达全量胃肠道营养时间、恢复出生体质量时间、胃肠功能紊乱发生率、血胆红素峰值、并发胆汁淤积比例等.采用t检验和x2检验进行统计学分析.结果 常规喂养组56例,早期微量喂养组48例,2组一般资料比较均无统计学差异.在达全量胃肠道营养时间、恢复出生体质量时间、血胆红素峰值、并发胆汁淤积等方面早期微量喂养组明显优于常规喂养组,差异均有统计学意义;2组患儿在住院天数及胃肠功能紊乱、吸人性肺炎、坏死性小肠结肠炎发生率方面差异均无统计学意义.结论 新生儿HMD实行早期微量喂养有利于患儿更早地过渡到全量胃肠道喂养,更好地恢复、增长体质量,减少胃肠外营养相关胆汁淤积的发生,无明显不良反应.     

Minimal residual disease (MRD) measurement as a tool to compare the efficacy of chemotherapeutic drug regimens using Escherichia Coli-asparaginase or Erwinia-asparaginase in childhood acute lymphoblastic leukemia (ALL)

BACKGROUND: L-asparaginase is a crucial drug in childhood acute lymphoblastic leukemia (ALL) induction therapy, but much debate remains regarding the optimal formulation and dosage. As minimal residual disease (MRD) can accurately measure extremely low levels of lymphoblasts, it is a sensitive reflection of leukemia cell kill. We utilized MRD to compare the efficacy of Erwinia-asparaginase (Erwinia-asp) and E. coli-asparaginase (E. coli-asp) during induction therapy for childhood ALL. PROCEDURE: Of 116 precursor-B ALL patients, 22 were treated with Erwinia-asp, 90 with E. coli-asp, and 4 were switched from E. coli-asp to Erwinia-asp. MRD levels at the end of induction were analyzed for 90 patients (Erwinia-asp = 16; E. coli-asp = 74). Patients were stratified into MRD > or =10(-2), between 10(-2)-10(-4) and < or =10(-4). Toxicity information during induction was available for 110 patients. RESULTS: MRD was the only significant prognosticator compared to conventional criteria. Patients treated with Erwinia-asp were 6.7 times more likely to have MRD levels > or =10(-2) (P = 0.031), reflecting slower lymphoblast clearance. While non-asparaginase related toxicities were similar in both groups, more E. coli-asp patients experienced severe asparaginase-related toxicity. CONCLUSION: E. coli-asp is superior to Erwinia-asp in childhood ALL induction. Although E. coli-asp is more toxic, this is balanced by better response to therapy. Early response to treatment as measured by MRD is a direct reflection of leukemic cell kill and is a significant prognosticator of eventual outcome, making it a good surrogate marker to evaluate the efficacy of induction drugs in childhood ALL.     

急性淋巴细胞白血病外周血WT1基因表达的意义

目的　建立realtimeRT PCR检测WT1基因表达方法 ,了解急性淋巴细胞白血病 (ALL)患儿外周血WT1基因表达水平。方法　建立realtimeRT PCR方法 ,采用PEABI 770 0PCR仪检测ALL 37例患儿、非白血病 1 3例和 1 8例正常儿童外周血WT1基因表达水平。结果　ALL初发 1 3例WT1基因的表达量为 (1 0 5～ 1 0 6 )拷贝 / μgRNA ,部分缓解 1 2例ALL患者的表达水平为 (1 0 2 ～ 1 0 4 )拷贝 / μgRNA ,ALL完全缓解 1 2例表达水平为 (0～ 1 0 2 )拷贝 / μgRNA。 结论　WT1基因在ALL外周血中有高水平表达 ,可作为疗效考核及监测微残留病指标。     

Use of the Nd:Yag Laser in the Surgical Treatment of Lung Metastases in Children

Renal dysfunction may occur in survivors of bone marrow transplantation (BMT). The renal function of children who have survived 5 to 10 years after BMT has not been reported. Bone marrow transplantation was performed in 55 children with acute lymphoblastic leukemia less than 18 years of age at the University of Florida between September 1983 and October 1992. All received a uniform conditioning regimen of high-dose cystosine arabinoside and fractionated total body irradiation. Twenty-three are currently surviving. The survival average period following transplantation is 79 ± 6.6 (SD) months. The longest survival is 129 months after BMT. We retrospectively examined data evaluating kidney function prior to transplantation, within 150 days after transplantation, and at each child's most recent clinic visit (1.7 to 10 years after transplantation). We were able to collect follow-up data regarding renal function for 17 survivors. Two children (11 %) have renal dysfunction in the form of hypertension, glucosuria, and hematuria. One of them had acute renal insufficiency during the first 100 days following BMT. An unexpected finding was the presence of hyperfiltration in 10 patients. In conclusion, in this homogeneous group of children, allogeneic BMT did not lead to significant long-term renal dysfunction.