伴t（8；21）（q22；q22）易位的儿童急性双表型白血病的生物学特征

目的探讨伴t（8;21）（q22;q22）易位的儿童急性双表型白血病（BAL）的生物学特征。方法分析7例伴t（8;21）（q22;q22）易位的儿童BAL,取同期诊断的30例t（8;21）阴性的急性髓细胞性白血病（AML）患儿作为对照组,分析其骨髓细胞形态学、细胞免疫学表型、细胞遗传学、分子生物学（MICM）特征。结果7例伴t（8;21）（q22;q22）易位的儿童BAL占同期连续190例急性髓细胞性白血病（AML）的3．7％。骨髓细胞形态学均显示为AML-M2,分类中原始细胞均显著增多（均P〈0．01）;免疫表型均为髓细胞系伴B淋巴细胞系表达;CD34为高表达阳性;均有t（8;21）（q22;q22）染色体改变,且常伴有染色体复杂易位或缺失等改变;融合基因AMLl／ETO检测均为阳性;7例患儿经治疗后完全缓解（CR）率71．4％,对兼顾AML和急性淋巴细胞白血病（AIJL）的联合治疗方案效果较好。结论BAL是急性白血病的一种亚型,其预后不良可能与染色体异常发生率高、CD34高表达阳性有关,对于BAL采用兼顾AML和ALL的联合治疗方案可提高其疗效。     

GSTT1,GSTM1和NQO1基因多态性与急性髓系白血病发生及其重现染色体异常关系的研究

目的探讨GSTT1,GSTM1和NQO1基因多态性与原发性急性髓系白血病（AML）易感性及AML染色体核型异常的关系。方法228例AML和241名与患者无血缘关系的正常人群对照,用多重聚合酶链反应（PCR）方法检测GSTT1和GSTM1基因型,用PCR-限制性内切酶片段长度多态性（RFLP）方法分析NQO1基因型。结果AML患者GSTM1无效型（null）比例（62．3％）明显高于正常对照组（52．7％）,而GSTT1无效型（null）比例与正常对照组比较,差异无统计学意义。NQO1C609TC／T和T／T基因型比例AML患者（分别为53．1％和25．0％）、t（8;21）（q22;q22）／AML—ETO阳性患者（分别为64．3％和25．0％）、t（15;17）（q22;q11）／PML—RARα阳性患者（分别为57．1％和26．0％）明显高于正常对照组（分别为49．4％和13．7％）。NQO1C609TC／T和T／T基因型患t（8;21）（q22;q22）／AML-ETO阳性AML的相对危险性分别为4．487（95％CI：1．282—15．705）和6．293（95％CI：1．536—25．782）,NQO1C609TC／T和T／T基因型患t（15;17）（q22;q11）／PML—RARα阳性AML的相对危险性分别为2．531（95％CI：1．286—4．981）和4．149（95％CI：1．856—9．275）。结论NQO1C609TC／T和T／T基因型与我国成人原发性AML,特别是伴重现染色体异常t（8;21）（q22;q22）／AML—ETO阳性及t（15;17）（q22;q11）／PML—RARα阳性AML高度相关。     

急性髓细胞白血病M2型AML1-ETO融合转录本的检测

目的：探讨检测AML1－ETO融合转录本在t（8;21）急性髓细胞白血病M2型M2／t（8;21）的临床诊断和预后判断中的意义。方法：用G显带技术对初治M2患者作染色体核型分析,用半筑巢式逆转录聚合酶链反应（RT／PCR）技术检测AML1－ETO融合基因转录本。结果：14例初治M2中共检出9例存在AMLI-ETO融合基因转录本,阳性率为64．3％。细胞遗传学技术证实9例均有t（8;21）易位;5例为阴性,均无t（8;21）易位;4例伴t（8;21）易位患者在处于完全缓解后5～72月,RT／PCR结果仍持续阳性。结论：应用筑巢式RT／PCR技术近似半定量检测AMLI－ETO融合转录本,对M2／t（8;21）的诊断和预后判断具有重要临床价值。     

正常核型急性髓系白血病患者融合基因的检测

目的探讨急性髓系白血病（AML）患者染色体畸变所形成的易位相关融合基因的临床和实验的关系。方法采用多重巢式RT—PCR方法和染色体核型分析技术对60例AML患者的融合基因进行分析。结果18例（30％）正常核型的AML患者分别检测出有PML／RARα、AML1／ETO、TLS／ERG、CBFβ／MYH11和MLL／AF9等5种融合基因的存在。结论多重巢式RT—PCR技术可用于白血病患者初诊时染色体畸变的筛选,可在核型正常的AML患者中检出隐匿的染色体易位,对AML的诊断分型具有重要帮助,进一步指导临床个体化治疗。     

1例伴45,X,-X,t(8;21)(q22;q22),del(9)(q22)M2型急性髓系白血病患儿的早期诊断

目的 通过血常规、外周血涂片及骨髓细胞MICM分型对1例伴45,X,-X,t(8;21)(q22;q22),del(9)(q22) M2型急性髓系白血病患儿进行早期诊断。方法血细胞五分类分析仪检测血常规;外周血涂片经瑞-吉染色后镜检分析血细胞形态;骨髓涂片经瑞-吉染色、过氧化物酶（POX）、酯酶双染色（AS-DNCE、α-NBE）进行细胞形态学诊断;流式细胞仪检测免疫表型;未刺激/24 h培养法、G显带分析染色体核型;FISH技术检测白血病融合基因。结果血常规白细胞计数（WBC）10.9×109·L-1、单核细胞比例（Mon%）29.8%、单核细胞绝对值（Mon#）3.3×109·L-1、血红蛋白（Hb）111g/L、血小板计数（PLT）90×109·L-1;血涂片发现血常规增高的单核细胞中部分为幼稚粒细胞,而非真正的单核细胞,且含有Auer小体;骨髓细胞检查诊断为AML-M2;流式结果符合急性髓系白血病AML-M2免疫表型;染色体核型45,X,-X,t(8;21)(q22;q22),del(9)(q22);融合基因AML1／ETO阳性。结论血常规及外周血细胞Auer小体结合骨髓细胞MICM分型有助于急性髓系白血病的早期诊断。     

急性早幼粒细胞自血病PML／RARα融合基因的检测

目的探讨荧光原位杂交技术（FISH）在检测急性早幼粒细胞白血病（APL）患者PML／RARα融合基闪中的价值。方法对初诊的30例及20例疑似APL的患者进行FISH技术检测PML／RARα融合基因,对治疗过程中PML／RARα融合基因阳性细胞比例进行动态观察,进而协助诊断和指导治疗。结果30例APL中PML／RARα融合基因阳性率90％（27／30）,1例AML1／ETO融合基因阳性,确诊为AML-M2;20例未确诊者中AML1／ETO融合基因阳性率15％（3／20）,PML／RARα融合基因阳性率50％（10／20）,其余7例未检测到以上两种融合基因。达到完全缓解（CR）后有7例融合基因阴性,随访至CR后2年,发现7例PML／RARα融合基因阳性比例较高患者分别于1-3个月后复发。结论FISH技术对APL的诊断具有高灵敏度和高准确度,并精确定位复杂核型中融合信号的位置,可用以确定白血病类型,指导临床治疗和进行治疗后的疗效监测。     

成人急性髓细胞性白血病细胞遗传学和分子遗传学的初步研究

目的：了解成人急性髓细胞性白血病（AML）染色体及相关融合基因的变化和意义。方法：95例AML患者采用直接法及24h培养法制备染色体,同时实时定量逆转录聚合酶链反应技术（RQ—PCR）检测t（8;21）易位的AMl。1／ETO融合基因及t（15;17）易位的PML／RAR~融合基因。结果：95例AMI。患者中异常核型48例（50．5％）,各亚型的异常率为：M3 88．5％,M2 44．7％,M4 21．4％,M1 50．0％,M5 16．7％。最常见的数目异常为＋8染色体4例（8．3％）,最常见的结构异常为t（15;17）21例（45．9％）,t（8;21）16例（33．3％）,QT—PCR检测M2、M3相关的融合基因AML1／ETO及PML／RARa均发现了隐匿易位及变异易位。结论：成人AML进行染色体分析及融合基因的检测,有助于AML诊断及亚型之间的鉴别诊断,有助于判断预后及治疗选择。     

AML1/ETO+融合基因阳性的AML患者骨髓细胞形态学分析

目的 探讨AML1/ETO[t(8;21)(q22;q22)]融合基因阳性的急性髓细胞白血病(AML)的骨髓细胞形态学特点.方法 对66例AML(M1,M2a,M2b,M5,MDS-RAEB-Ⅱ) 骨髓片用瑞氏染液染色后,按照国内AML 分型标准进行观察分析;进行荧光原位杂交技术(FISH)检测AML1/ETO 融合基因.结果 M2a 52例, M2b 8例, M5 2例, M1 2例,MDS-RAEB-Ⅱ 2例,AML1/ETO融合基因阳性为34.84%,其中(14/52)26.9%,(8/8)100%,(0/2)0%,(0/2)0%,(1/2)50%.回顾分析AML1/ETO阳性病例骨髓涂片,都检测到多少不一的异形中幼粒细胞.AML1/ETO阳性病例在治疗过程中首次化疗完全缓解(CR)率为78.4%(9例M2a、8例M2b及1例RAEB-Ⅱ),4例M2a 2次化疗后达到CR,1例M2a 5次化疗仍为NR.结论 AML-M2b都有AML1/ETO融合基因改变,AML1/ETO融合基因阳性AML在骨髓涂片中能检测到异形中幼粒细胞.     

慢性和急性髓细胞白血病伴特异性染色体患者的细胞形态学特征

目的 分析174例初发急性和慢性髓细胞白血病，包括急性原始粒细胞白血病部分分化型（M2b），急性早幼粒细胞白血病（M3）和慢性粒细胞性白血病（CML）患者细胞形态学特征。方法 采用细胞形态学（M）结合细胞遗传学（C）和分子生物学（M）检查方法检测患者骨髓标本。结果 细胞形态学特征：25例M2b为异常中幼粒细胞增生，28例M3为异常早幼粒细胞增生，121例CML为粒系细胞增生。染色体及融合基因检测表明：M2b、M3、CML分别与各自特异性染色体及融合基因t（8；21）（q22；q22）／AML1-ETO、t（15；17）（q22；q21）／PML-RARd、t（9；22）（q34；q11）／BCR—ABL密切相关。结论 ①M2b、M3、CML这3种白血病患者在细胞形态学上都有各自一定的特征。②3种白血病分别与特定的染色体及融合基因具有一定的相关性。③特定的染色体和融合基因作为白血病的标记物，对白血病的诊断、预后估计、监测治疗和微小残留白血病等都具有一定价值。     

伴Y染色体缺失的t(8;21)急性髓系白血病2例

在急性髓系白血病（AML）中，t（8；21）（q22；q22）是一种常见的非随机的染色体异常，这种异常主要见于急性髓细胞白血病部分分化型（AML-M2），约占AML-M2b的90．0％。在未分化型（M1）、早幼粒细胞白血病（M3）、粒-单核细胞白血病（M4）等AML其他亚型及慢性髓系白血病（CML）、骨髓增生异常综合征（MDS）和真性红细胞增多症（PV）患者中也偶见。     

实时定量聚合酶链式反应与巢式聚合酶链式反应在 289例白血病融合基因检测中的比较

 目的 比较实时定量聚合酶链式反应（RT-PCR）与巢式聚合酶链式反应（nPCR）在白血病融合基因BCR/ABL、PML/RARa及AML1/ETO检测中结果。方法 分别应用RT-PCR与nPCR对289例急慢性白血病初诊和完全缓解的患者BCR/ABL、PML/RARa及AML1/ETO融合基因进行检测,结合患者骨髓细胞形态学及临床转归予以分析。结果 46例初诊慢性粒细胞白血病（CML）患者中,RT-PCR法与nPCR法检测BCR/ABL融合基因阳性率分别为95.7%和93.5%（P=0.997）;69例完全缓解CML患者中,RT-PCR法与nPCR法检测阳性率分别为78.4%和58.1%（P<0.005）。32例初诊急性早幼粒细胞白血病（APL）患者中,RT-PCR法与nPCR法检测PML/RARa融合基因阳性率分别为93.8%和90.6%（P=0.996）;114例完全缓解APL患者中,RT-PCR法与nPCR法检测阳性率分别为12.3%和7.0%（P<0.025）。12例初诊急性粒细胞白血病M2（ANLL-M2）患者中,RT-PCR法与nPCR法检测AML1/ETO融合基因阳性率分别为50.0%和50.0%（P=1.0）;16例完全缓解ANLL-M2患者中,RT-PCR法与nPCR法检测阳性率分别为50.0%和12.5%（P<0.05）。结论 RT-PCR较nPCR更为敏感而准确,应用RT-PCR可以提高微小残留病的检出率,为临床诊断和治疗提供更为有效的分子生物学依据。     

成人和儿童急性白血病常见融合基因表达的比较

目的 比较成人和儿童急性白血病常见融合基因表达的差异.方法 采用多重巢式PCR技术检测159例成人和120例儿童初治急性白血病患者的常见融合基因,比较成人及儿童各融合基因的阳性率.结果 儿童急性淋巴细胞白血病(ALL)患者白血病融合基因总阳性率为51.11%(46/90),高于成人ALL患者的25.53%(24/94)(P<0.05);其中儿童TEL/AML1及TLS/ERG融合基因阳性率高于成人(P>0.05).儿童急性髓系细胞白血病(AML)患者白血病融合基因总阳性率为53.33%(16/30),高于成人AML患者的26.15%(17/65)(P<0.05);其中儿童AML1/ETO融合基因阳性率高于成人(P>0.05).结论 成人及儿童急性白血病融合基因的表达存在一定差异,临床上应有针对性地进行相关融合基因的检测和判读,以提高诊疗质量.     

靶向AML1/ETO的siRNA增强Kasumi-1细胞对组蛋白去乙酰化酶抑制剂的敏感性

目的 探讨AML1/ETO融合基因沉默后kasumi-1细胞株对组蛋白去乙酰化酶抑制剂(HDACI)丙戊酸钠(VPA)的敏感性变化。方法 体外培养人急性髓系白血病细胞株kasumi-1,脂质体介导重组质粒pGCsiRNA-AML1/ETO转染kasumi-1,实时荧光定量PCR(RT-PCR)检测Kasumi-1细胞AML1/ETO和Bcl-2 mRNA的表达变化;MTT法检测VPA对细胞增殖的影响;流式细胞术(FACS)检测细胞周期;Hochest33258染色后荧光显微镜下观察细胞凋亡的形态学变化。结果 靶向AML1/ETO基因的siRNA质粒可有效降低Kasumi-1细胞AML1/ETO基因的表达;与空白对照组相比, 转染组细胞AML1/ETO基因mRNA表达量下降了53.7%(P<0.05), 同时Bcl-2 mRNA的表达量下降了54.5%(P<0.05),而阴性对照组及脂质体组两基因的表达量无明显变化;VPA对Kasumi-1细胞增殖的抑制作用呈浓度、时间依赖性,不同VPA浓度作用下,转染组细胞24、48h的抑制率均高于空白对照组(P<0.05),而阴性对照组及脂质体组与空白对照组相比无明显变化。空白对照组及转染组细胞G0/G1期比例分别为(42.07±5.23)%和(62.6±5.87)%(P<0.01),经含2mmol/L VPA的培养基培养48h后,两组细胞G0/G1期比例分别上升至(69.2±7.02)%和(78.92±6.23)%(P<0.01);转染后细胞出现核固缩、核边集、凋亡小体等改变。结论 特异性AML1/ETO siRNA可显著抑制Kasumi-1细胞AML1/ETO基因的表达,并明显增强Kasumi-1细胞对VPA的敏感性。     

急性髓系白血病H-ras、凋亡及多药耐药的研究

目的研究急性髓系白血病（ＡＭＬ）Ｈｒａｓ／Ｐ２１、抗凋亡基因（ｂｃｌ２）、细胞凋亡与多药耐药基因（ｍｄｒ１／Ｐｇｐ）对疗效的影响,并证实流式细胞术（ＦＣＭ）检测Ｐｇｐ和逆转录多聚酶链反应（ＲＴＰＣＲ）检测ｍｄｒ１ｍＲＮＡ间的相关。方法用ＦＣＭ检测８５例初治ＡＭＬ患者的Ｈｒａｓ／Ｐ２１、ｂｃｌ２、细胞凋亡和ｍｄｒ１产物Ｐｇｐ的表达;用ＲＴＰＣＲ检测其中４７例的ｍｄｒ１／ｍＲＮＡ。结果Ｐ２１＋／Ｐｇｐ＋者完全缓解（ＣＲ）率明显低于Ｐ２１－／Ｐｇｐ－者（分别为５８．１％和１００％,Ｐ＜０．０２５）,高ｂｃｌ２／Ｐｇｐ＋的ＣＲ率明显低于低ｂｃｌ２／Ｐｇｐ－者（分别为６０％和９６．３％,Ｐ＜０．００５）。Ｐｇｐ＋３１例中２９例ｍｄｒ１ｍＲＮＡ阳性,Ｐｇｐ－１６例中仅２例ｍｄｒ１ｍＲＮＡ阳性。结论Ｐｇｐ＋同时Ｐ２１＋、高ｂｃｌ２者ＣＲ低,ＦＣＭ检测Ｐｇｐ和ＲＴＰＣＲ检测ｍｄｒ１ｍＲＮＡ相关性良好     

AML1/ETO阳性急性髓系白血病患者galectin-3的表达及临床意义

目的 探讨AML1/ETO阳性急性髓系白血病（AML1/ETO+ AML）患者半乳凝集素3（galectin-3,gal-3）表达及其临床意义。方法 采用实时定量RT-PCR（RQ-PCR）法检测53例AML1/ETO+ AML患者骨髓单个核细胞中gal-3 mRNA表达水平,ELISA法检测患者外周血gal-3蛋白水平,分析gal-3 mRNA表达与疾病预后的相关性。结果 ①gal-3 mRNA及蛋白在初诊AML1/ETO+ AML患者中呈高表达（ P<0.001）,初诊患者gal-3 mRNA与蛋白表达呈正相关（r=0.732, P<0.001）,疾病完全缓解期外周血gal-3蛋白表达量下降（ P<0.001）;②初诊时gal-3 mRNA表达水平越高,患者白细胞数量越低（ P=0.014）,CD34表达和附加染色体异常的比例越高（ P=0.001, P=0.026）;③gal-3 mRNA高表达组与低表达组在完全缓解（CR）、部分缓解（PR）、诱导期死亡（即早期死亡）率上无明显差异（ P＞0.05）,但两组患者在复发率上差异有统计学意义（ P=0.029）;④gal-3 mRNA高表达组患者中位总生存率（OS）及无病生存率（DFS）均较低表达组缩短（ P=0.007, P=0.015）,累积复发率增高（ P=0.045）,但累积死亡率无明显差异（ P＞0.05）。Cox风险比例模型提示gal-3 mRNA是影响AML1/ETO+ AML患者OS及DFS的独立指标。结论 骨髓gal-3 mRNA有可能成为AML1/ETO+ AML患者评估预后及指导治疗的重要参考指标。     

MiR-130a is aberrantly overexpressed in adult acute myeloid leukemia with t(8;21) and its suppression induces AML cell death

Background: Emerging evidence has revealed that miRNAs can function as oncogenes or tumor suppressor genes in leukemia. The ectopic expression of miR-130a has been reported in chronic leukemia, but our understanding of the biological implications of miR-130a expression remains incomplete.

Methods: We quantified a cohort of de novo acute myeloid leukemia (AML) by bead-based miRNA and real-time quantitative PCR (Rq-PCR). The luciferase reporter gene assay was analyzed after the plasmid constructs which contain 5’-UTR of miR-130a and a Renilla luciferase reporter plasmid were transfected simultaneously into 293T cells. MTT and caspase 3/7 apoptosis assays were used to test cell viability and apoptosis.

Results: We identified miR-130a as significantly overexpressed in t(8;21) AML. Expression of miR-130a decreased significantly once patients with t(8;21) achieved complete remission, but increased sharply at the time of relapse. In patients with t(8;21) AML, KIT mutational status was associated with miR-130a expression—with higher expression associated with KIT activating mutations. Increased miR-130a expression in t(8;21) AML was associated with slightly worse event-free survival; however, no impact on overall survival was observed. Knockdown of AML1/ETO protein in the SKNO-1 cell line resulted in decrease of expression of miR-130a. Direct binding of AML1/ETO fusion protein with the promoter sequence of miR-130a was detected with luciferase reporter gene assay. Following miR-130a knockdown, SKNO-1 demonstrated increased sensitivity to etoposide.

Conclusions: Our data suggest that miR-130a is directly activated by AML1/ETO, and may act as a factor which is associated with leukemia burden, event-free survival, and chemotherapy sensitivity in t(8;21) AML.     

NPM1基因在急性髓细胞白血病中的表达及其临床意义

目的探讨急性髓细胞白血病(AML)患者骨髓单个核细胞(BMMNC)野生型NPM1基因的表达及临床意义。方法采用半定量逆转录聚合酶链反应(RT-PCR)方法检测59例AML患者,分析NPM1基因表达水平与患者性别、年龄、外周血细胞数、骨髓原始幼稚细胞数及临床疗效的关系,并对部分患者NPM1表达水平进行动态观察。结果初治AML患者中NPM1阳性表达率为73.8%,难治复发组NPM1阳性率为100%,完全缓解组NPM1阳性表达率为66.7%,对照组NPM1阳性表达率为60%,但各组之间阳性率差异无统计学意义(P>0.05)。分析表达水平显示,难治复发组NPM1中位表达水平(0.798 8)高于初治组(0.541 0)、完全缓解组(0.483 6)及对照组(0.371 8)(P<0.05)。NPM1高表达组患者有效为63.1%,低于低表达组(88.9%)和不表达组患者(100%)(P<0.05)。NPM1表达强度与外周血白细胞数和骨髓原始细胞数呈正相关(P<0.05)。对12例初治时NPM1基因高表达患者进了动态观察,1个疗程化疗后获完全缓解5例,其表达水平达正常中位水平以下,其余7例为未缓解患者,NPM1表达水平未见明显变化。结论 NPM1基因在AML中均存在不同程度的过度表达现象,NPM1高表达者提示预后差,完全缓解率低,可作为AML预后不良的指标,PCR动态检测AML患者治疗前后NPM1表达水平,可作为判断AML预后和监测微小残留病(MRD)的一项指标。     

All-trans retinoic acid as a single agent induces complete remission in a pat ient with acute leukemia of M2a subtype

Objective To present a special case with the karyotype and molecular marker of acute myelo id leukemia (AML)-M(2) who was induced to complete remission by all-trans reti noic acid (ATRA) alone.Methods A recently hospitalized young female patient with acute leukemia was initially d iagnosed as M(3) subtype based on morphological French-American-British (FAB) classification. Karyotype analysis using standard G and R banding techniques an d RT-PCR were applied to further define the diagnosis. After primarily culture d bone marrow cells from the iliac aspiration were tested for in vitro induced d ifferentiation, the patient was treated with oral all-trans retinoic acid alone , 60 mg per day until complete remission was achieved. Peripheral blood and bo ne marrow changes were monitored over the whole treatment course. Results The characteristic chromosomal aberration for M(3) , the t(15;17) reciprocal tran slocation, was not found while a t(8;21) translocation was verified. Furthermor e, an amplified product of the AML-1/ETO fusion gene instead of the PML/RARα f usion gene was detected by RT-PCR and the diagnosis was corrected from M(3) to M(2) . Primary cultured bone marrow cells can be fully induced to terminal diffe rentiation after 4 days exposure to ATRA. A hematological complete remission wa s achieved after 40 days treatment with ATRA as a single therapeutic agent, sug gesting an alternative pathway mediating ATRA-induced myeloid differentiation. Conclusion A leukemia patient with a subtype other than M(3) , such as M(2) in this case, ma y also be induced to complete remission by the mechanism of ATRA-induced termin al differentiation. This implies that there may be a pathway other than PML/RAR α fusion gene product which mediates ATRA-induced myeloid maturation in leukem ia cells.     

常见白血病融合基因筛查在白血病诊断与分型中的意义

目的:分析病人白血病细胞染色体畸变涉及的86种融合基因和临床白血病类型的相关性,探讨常见融合基因筛查法在临床诊断和分型中的应用价值.方法:收集161例初发或者复发的白血病患者及8例骨髓增生异常综合征(MDS)患者的骨髓细胞,提取RNA,用32条特异性引物逆转录为cDNA,利用白血病29种染色体畸变形成的融合基因的86种mRNA剪接变异体引物,分8管进行多重RT-PCR,筛查白血病融合基因.结合临床状态和形态学观察了解融合基因与白血病类型的关系.结果:白血病中115例(71%)分别检测出10种白血病常见融合基因,包括AMLl/ETO、PML/RARα、PLZF/RARα、dupMLL、MLL/AF6、MLL/AF10、CBFβ/MYHll、BCR/ABL、Hoxll、Evil.其中52例慢性粒细胞白血病(CML)100%检出BCR/ABL;25例急性早幼粒细胞白血病(APL)中88%检出融合基因,其中21例APL检测出PML/RARα,1例APL检测出PLZF/RARα;AMLl/ETO阳性的17例急性白血病(AL)16例为FAB-M2亚型,1例为混合型白血病;CBFβ/MYH11阳性的4例AL 3例为FAB分型的M4,1例为M5,属于向粒单细胞系统分化的白血病.16例AL检测出MLL基因异常,其中MLL/AF6白血病均为FAB分型的M5,具有典型的原始单核细胞白血病的特征.17例急性淋巴细胞白血病(ALL)5例检测出BCR/ABL.8例MDS病人中2例检测出融合基因,其中AMLl/ETO阳性的MDS-RAEB很快发展为AML.结论:这种以多重RT-PCR为基础的白血病常见融合基因筛查法可以准确、快速而且可靠地确定白血病的分子类型,提供白血病诊断和治疗的依据.     

Chromosomal changes detected by fluorescence in situ hybridization in patients with acute lymphoblastic leukemia

Objectives To investigate patients with acute lymphoblastic leukemia (ALL) for TEL/AML1 fusion,BCR/ABL fusion, MLL gene rearrangements, and numerical changes of chromosomes 4, 10, 17 and 21 by fluorescence in situ hybridization (FISH) and to determine the relationship and the significance of those findings.Methods Fifty-one American patients (34 men and 17 women) were included in this study. Of them there were 41 patients with pro-B cell type ALL, 9 with B cell type ALL and 1 with T cell type ALL.Chromosome metaphases of each sample were prepared according to standard protocols.Fluorescence in situ hybridization was performed using commercially available DNA probes, including whole chromosome painting probes, locus specific probes, specific chromosome centromere probes and dual color/multiple color translocation fusion probes. The digital image analysis was carried out using Cytovision and Quips FISH programs.Results An overall incidence of chromosomal anomalies, including t (9; 22 ), MLL gene rearrangements, t (12;21), and numerical chromosomal anomalies of chromosomes 4, 10, 17 and 21 was found in 33 patients (65%). Thirty-one of them were pediatric patients and two adults. The t(12;21) was the commonest chromosomal anomaly detected in this population; 14 out of the 45pediatric patients (31%) were positive for TEL/AML1 fusion, among which three had an additionalderivative 21[t (12;21) ], four had a deletion of 12p and two had an extra copy of chromosome 21.All 14 patients with positive TEL/AML1 fusion had ALL pre-B cell or B-cell lineage according to standard immunotyping. The percentage of cells with fusion signals ranged from 20% to 80%. All fourteen patients positive for TEL/AML1 gene fusion were mosaic. Three out of the 14 patients positive for the TEL/AML1 gene fusion were originally reported to be culture failures and none of the remaining eleven samples had been found to have chromosome 12 abnormalities by conventional cytogenetic techniques. All pediatric patients with pre-T or T cell lineage and the six adults were negative for TEL/AML1 fusion. One patient had double Philadelphia chromosomes, three had a rearranaement or a deletion of the MLL aene. one had t (4;11)and two had a deletion of the MLL One of the patients with an MLL deletion also had a large ring of chromosome 21, and r (21) was caused by AML1 gene tandemly duplicated at least five times. The second case with the MLL deletion was also unique, the patient had a t (12;21) as well. A total of 20 patients had numerical changes( gain or loss) of chromosomes 4, 10, 17 and 21. Eight patients were found to have trisomies of three or four different chromosomes. Interestingly, seven of these patients did not have TEL/AML1, BCR/ABL or the MLL aene rearranaement, one did have the TEL/AML1 aene fusion. Eleven patients with pro-B cell or B cell type ALL (9 children with ALL, 2 adults with ALL) had numerical changes of chromosome 21 (gain 1 or 2 chromosome 21 ), among them, 10 patients had no structural alteration of chromosome 21, and one was combined by t (12;21 ). Four patients had a monosomy of chromosome 17 and three out of these patients with monosomy 17 also had a fusion signal of TEL/AML1.