三氧化二砷治疗急性早幼粒细胞白血病疗效观察

目的 :观察三氧化二砷 (As2 O3)治疗急性早幼粒细胞白血病的完全缓解率、高白细胞发生率、肝功损害及融合基因转阴率 ,并与维甲酸 (ATRA )的治疗情况进行比较。方法 :随机分为两组 ,As2 O3组 17例 ,ATRA组 2 1例 ,分别选用 0 .1% As2 O310 m L/ d,静脉滴注 ;ATRA2 5 mg· m- 2 · d- 1 ,分 3次服用。分别观察两组的完全缓解率 (CR)、不良反应以及融合基因转阴率。结果 :As2 O3组 15 / 17例 (88.2 % )获 CR,获得 CR时间为 (2 8.1± 4 .6 ) d;ATRA组 19/2 1例 (90 .4 % )获 CR,获得 CR时间为 (39.4± 8.6 ) d,两组之间 CR无明显差别 ,但 As2 O3组获得 CR时间明显缩短。高白细胞发生率 As2 O3组 10 / 15例 (6 6 .7% ) ,ATRA组 18/ 19例 (94 .7% ) ,P<0 .0 5。肝功能异常 ,As2 O3组 11/ 17例(6 4 .7% ) ,ATRA组 13/ 19例 (6 8.4 % ) ,P>0 .0 5。所有患者治疗前 PML- RARα融合基因阳性。 As2 O3组 CR时 ,2 /14例 (14 % )转阴 ,ATRA组 2 / 19例 (10 .5 % )转阴 ,P<0 .0 5。CR后 1a,As2 O3组 5 / 8例 (6 2 .5 % )转阴 ,ATRA组 4 /11例 (36 .3% )转阴 ,P<0 .0 5。As2 O3组 15例获 CR者 ,无 1例复发。ATRA组 19例获 CR者 ,4例 (2 1% )复发。结论 :与 ATRA相比 As2 O3获得 CR时间缩短 ,高白细胞发生率低 ,CR及 CR     

四硫化四砷对急性早幼粒细胞白血病细胞株NB4的凋亡作用

目的 研究四硫化四砷对NB4细胞的促凋亡作用及这一过程中早幼粒细胞白血病-维甲酸受体α(PML-RARα) 融合基因及其表达产物的变化。方法 通过细胞形态学观察, 流式细胞仪检测及DNA电泳等方法观察四硫化四砷对NB4细胞的诱导凋亡作用，用荧光染色体原位杂交技术, 反转录PCR及Western印迹技术测定这一过程中PML-RARα融合基因及其表达产物的改变。结果 四硫化四砷在0.5～3 μmol·L^-1之间能诱导NB4细胞凋亡，在此过程中，PML-RARα融合基因无明显变化，但PML-RARα融合蛋白和野生型RARα蛋白的表达明显减少。结论 四硫化四砷能诱导NB4细胞凋亡，其作用靶点可能在PML RARα融合蛋白和野生型RARα蛋白。     

全反式维甲酸与亚砷酸联合化疗对急性早幼粒细胞白血病高危患者的疗效

目的观察全反式维甲酸(ATRA)与亚砷酸(ATO)联合化疗对急性早幼粒细胞白血病(APL)患者的疗效。方法回顾性分析86例不同危险分级的初治APL患者的临床资料。根据治疗前白细胞和血小板数将APL患者分为低、中、高危三组。采用ATRA+ATO+蒽环类药诱导缓解,蒽环类药+阿糖胞苷巩固治疗,ATRA+ATO+甲氨蝶呤(MTX)[部分加用6-巯基嘌呤(6-MP)]维持治疗。结果治疗后,完全缓解(CR)率高达95.3%(82/86)。中位随访37个月,高危组和中低危组无事件生存率及中枢神经系统累积复发率差异均无统计学意义(P>0.05)。维持治疗单用MTX者或MTX+6-MP者CR率均为100%。结论 APL患者尤其是高危患者可以从ATO+ATRA+化疗中受益;该方案作为初治APL的一线治疗方案优势明显。     

三氧化二砷治疗急性早幼粒细胞白血病的进展

三氧化二砷(As2O3)用于治疗急性早幼粒细胞白血病(APL)已有20余年,通过特异性降解PML-RARα融合蛋白,As2O3在复发难治和初发APL患者中均获得了较理想的疗效,不良反应较轻,已成为复发难治APL的首选药物.通过进一步临床试验,As2O3有望成为初发APL的一线用药,并可减低甚至取代化疗,使部分APL成为无需化疗而获得治愈的血液学肿瘤.     

全反式维甲酸诱导小鼠间充质干细胞C3H10T1/2表达碱性磷酸酶

目的分析全反式维甲酸对小鼠间充质干细胞C3H10T1/2表达碱性磷酸酶的影响。方法全反式维甲酸（10-6M）、维甲酸核受体RAR的激动剂Ch55与MAPK信号通路的阻滞剂处理C3H10T1/2细胞,钙钴染色检测碱性磷酸酶活性,定量RT-PCR方法检测量效关系以及时相分布;采用萤光素酶报告基因检测碱性磷酸酶的启动子是否接受维甲酸的调控。结果维甲酸处理C3H10T1/2细胞24h即能够明显上调ALP的活性,具有明显的剂量依赖特性,与RAR有关,无MAPK信号通路的参与,在其转录起始位点上游-512bp的范围内检测到核受体RAR的调控位点。结论维甲酸能够通过核受体RAR直接上调碱性磷酸酶的转录水平。     

全反式维甲酸和三氧化二砷治疗初治急性早幼粒细胞白血病的疗效观察

目的观察全反式维甲酸(ATRA)联合三氧化二砷(As2O3)治疗初治急性早幼粒细胞白血病(APL)的疗效。方法对应用ATRA和As2O3联合诱导治疗的20例APL患者的完全缓解(CR)率、达CR所需时间和不良反应进行观察,并与单独应用ATRA组30例进行比较。联合用药组治疗方法为ATRA 25mg/(m2.d),0.1%As2O310ml/d直至CR。结果联合用药组与单独应用ATRA组相比,CR率差异无统计学意义(分别为95%、86.7%,均P>0.05);联合用药组获得CR所需的时间短于单独用药组(平均时间分别为24d、45d,均P<0.05),早期死亡率较单独用药组差异无统计学意义(分别为5%、13.3%,均P>0.05);与单独用药组相比,联合用药组的不良反应并未增加。结论联合用药诱导初发APL缓解的疗效优于单用药组,不良反应少,是一种值得推广应用的方案。     

Targeted therapies for the myeloid leukaemias

Although the standard approach to myeloid leukaemias remains chemotherapy, the agents currently available rarely result in cure. Recent advances in understanding the biology of these disorders have lead to the development of targeted treatment strategies. In acute promyelocytic leukaemia (APL), all-trans retinoic acid (ATRA), sodium phenylbutyrate and arsenic trioxide are agents which either induce differentiation or apoptosis and have been used to successfully achieve remission. The tyrosine kinase inhibitor, STI-571, antisense oligonucleotides, and bcr-abl vaccines are strategies which focus on the oncogenic events in chronic myelogenous leukaemia (CML). Two anti-CD33 monoclonal antibody conjugates, Y90-HuM195 and CMA-676, have been used in acute myelogenous leukaemia (AML) and have shown some efficacy. Although the preliminary results with these targeted therapies are promising, further studies are needed to establish them as effective, less toxic alternatives to the current standard of care.     

Targeted therapies for the myeloid leukaemias

Although the standard approach to myeloid leukaemias remains chemotherapy, the agents currently available rarely result in cure. Recent advances in understanding the biology of these disorders have lead to the development of targeted treatment strategies. In acute promyelocytic leukaemia (APL), all-trans retinoic acid (ATRA), sodium phenylbutyrate and arsenic trioxide are agents which either induce differentiation or apoptosis and have been used to successfully achieve remission. The tyrosine kinase inhibitor, STI-571, antisense oligonucleotides, and bcr-abl vaccines are strategies which focus on the oncogenic events in chronic myelogenous leukaemia (CML). Two anti-CD33 monoclonal antibody conjugates, Y90-HuM195 and CMA-676, have been used in acute myelogenous leukaemia (AML) and have shown some efficacy. Although the preliminary results with these targeted therapies are promising, further studies are needed to establish them as effective, less toxic alternatives to the current standard of care.     

细胞遗传学在急性早幼粒细胞白血病的诊断及预后评估中的作用

目的探讨细胞遗传学在急性早幼粒细胞白血病（APL）的诊断及预后评估中的应用价值。方法应用常规细胞遗传学分析APL初诊患者39例,采用R显带技术进行染色体核型分析。随访分析完全缓解率、复发率。结果39例APL患者中,染色体异常38例,其中30例为典型t（15;17）易位,8例为涉及15和17号染色体的复杂核型。形态学误诊为M2和M5各1例,经核型分析确诊。经全反式维甲酸和（或）化疗联合诱导治疗,37例获完全缓解,2例具有复杂核型者未取得完全缓解,6例具有复杂核型者第1次完全缓解后3年内复发,复发率86％（6／7）,其他核型组完全缓解后3年内复发率仅10％（3／31）。结论细胞遗传学对细胞形态不典型APL具有确诊价值,复杂染色体异常者预后不佳。     

One acute promyelocytic leukemia patient underwent complete molecular remission with consistent presence of t (2; 3) (p25; q21) karyotype

Objectives: This study aimed to explore why one acute promyelocytic leukemia (APL) patient underwent complete molecular remission in the persistent presence of the t (2; 3) (p25; q21) karotype.

Methods: One APL patient overexpressed PML/RARα (bcr1) and WT1 genes in the presence of the Fms-like tyrosine kinase-internal tandem duplication mutation, while cytogenetics showed t (2; 3) (p25; q21) and t (15; 17) (q22; q21). Cytogenetics and molecular biology were monitored throughout the treatment.

Results: After 5 weeks of induction chemotherapy, this case gained complete molecular biology remission with the presence of t (2; 3) (p25; q21). This status was still present during the follow-up consolidate and maintenance therapy.

Conclusion: For this patient, t (2; 3) (p25; q21) may be one kind of balanced translocation that leads to miscarriages or causes abnormalities in children, unrelated to leukemia or other malignancies.     

PML-RARα融合基因相关蛋白原核表达载体的构建、表达和纯化

目的 构建PML-RAR o[融合基因相关重组表达质粒,在大肠埃希菌中表达可溶性蛋白并进行纯化.方法 利用聚合酶链反应(Polymerase Chain Reaction,PCR)以PML-RAR α全长质粒为模板分别扩增出长度均为1 200 bp的PML和RAR o[序列,并将它们分别插入PET32a(+)质粒中,构建重组表达质粒,化学法转化大肠埃希菌DH5α进行克隆,菌落PCR筛选阳性转化子,双酶切和测序鉴定重组质粒的正确性.正确重组的表达质粒分别命名为PML-Flag-PET32a(+)和Flag-RARα-PET32a(+).将正确重组的表达质粒转化感受态大肠埃希菌BL21(DE3),经异丙基β-D-半乳糖苷(IPTG)诱导表达,利用表达蛋白的组氨酸“标签”(His-tag)进行Ni2+-树脂柱亲和层析纯化,SDS-PAGE和Western blotting鉴定纯化蛋白质.结果 PCR扩增获得目的基因,重组表达质粒经EcoRI/HindⅢ双酶切和测序鉴定证明构建正确.转化感受态BL21(DE3)并经诱导后得到高效表达,SDS-PAGE和Western blotting显示纯化蛋白为目的蛋白.结论 成功构建重组表达质粒,并经诱导表达后可纯化出目的蛋白,为进一步的抗体制备等实验奠定了基础.     

急性早幼粒细胞白血病18例临床分析

目的　探讨急性早幼粒细胞白血病(APL)的临床特点、最佳治疗方案、维A酸综合征(RAS)的防治。方法　分析18例APL患者的临床资料。结果　18例APL患者中以出血起病16例。维A酸联合化疗治疗8例,疗程>6周期2例,分别于第2 6个月、第72个月复发死亡;2例早期死亡;4例<4周期者2例无病生存5年以上,2例于5年后复发,治疗后再次完全缓解。维A酸、砷剂联合化疗治疗10例无病生存2～37个月。结论　APL是一种特殊类型急性白血病,临床以出血为主要症状,RAS以预防为主。     

急性早幼粒细胞白血病PML-RARα融合基因异构体cDNA实时定量PCR标准品及标准曲线的构建

目的克隆急性早幼粒细胞白血病PML-RARα融合基因异构体L型、S型cDNA作为实时定量聚合酶链反应(FQ-PCR)检测PML-RARα融合基因的标准品,从而构建FQ-PCR检测标准曲线。方法应用反转录-聚合酶链反应(RT-PCR)从急性早幼粒白血病患者骨髓单个核细胞的总RNA中逆转录扩增的PML-RARα融合基因异构体L型、S型cDNA,将纯化的PML-RARα融合基因异构体L型、S型cDNA与pMD18-T载体进行连接,转化宿主菌E.coli DH5a感受态细胞,氨苄青霉素培养基筛选阳性克隆,提取重组质粒cDNA用限制性核酸内切酶HindⅢ、EcoRⅠ进行双酶切鉴定并测序分析,最后进行实时荧光定量PCR检测。通过检测重组质粒260 nm吸光度,确定原液的重组质粒拷贝浓度并以此制备荧光定量PCR标准品。结果酶切鉴定、PCR扩增及测序分析均证实PML-RARα融合基因重组到pMD18-T载体。结论成功克隆急性早幼粒白血病PML-RARα融合基因异构体L型、S型cDNA。     

Arsenic trioxide in hematological malignancies: the new discovery of an ancient drug

Currently, Arsenic Trioxide (ATO) is considered the treatment of choice for patients with relapsed acute promyelocytic leukemia (APL). Recently, a durable remission with minimal toxicity by single agent ATO or ATO + ATRA in newly diagnosed APL was reported by different groups. These regimens have minimal toxicity and can be administered on an outpatient basis after remission induction, thus they could become a real, less toxic and more economic option to ATRA + anthracyclines in particular in low risk APL, or in patients that cannot undergo chemotherapy because of age or comorbid conditions and in patients that refuse chemotherapy. Significantly, these therapies are a successful attempt to cure a tumoral disease without chemotherapy. The results of clinical trials of ATO administration as single agent in multiple myeloma (MM) and myelodisplastic syndromes (MDS) were encouraging and showed clinical effects but they were not close to APL success. On the contrary, results of clinical trials to treat non-APL acute myeloid leukemia (AML) were disappointing. We suggest that a combination therapy with drugs targeting specific pro-survival molecules or capable to enhance pro-apoptotic pathways may lead to an improvement of ATO efficacy against hematological malignancies, in particular AML. Our pre-clinical studies showed that ATO is capable to induce cell death in acute leukemia cells but the apoptotic function is limited since it can induce also a mechanism of cell defense by activating pro-survival molecules such as MEK-ERK, Bcl-xL, Bcl-2. By combining ATO with specific MEK inhibitors, we demonstrated that the block of MEK-ERK phosphorylation, the induction of Bad de-phosphorylation, and activation of p53AIP1 apoptotic pathway interrupt the pro-survival mechanisms of ATO and kill the leukemic cells by apoptotic synergism. Our results provide an experimental basis for combined or sequential treatment with MEK inhibitors and ATO in AML. The renaissance of ATO as a drug in moderne medicine may be considered, together with ATRA success, a victory of empirical analysis, that had (and has) great impact on Chinese culture.