慢性粒细胞性白血病骨髓移植后的染色体观察

为了观察慢性白血病骨髓移植术后患者的生存情况，笔者对费城染色体进行了检测，结果表明：患者术后９个月，ＸＸ核型细胞占８７％，ＸＹ核型细胞占１３％，费城染色体阴性；术后１年，ＸＸ核型细胞占６４％，ＸＹ核型细胞占３４％，费城染色体阳性细胞占２％．慢性白血病骨髓移植术后观察费城染色体，能及时反映预后情况     

慢性粒细胞白血病急变与bcr/abl，p53，p16基因相关性研究

应用RT-PCR、PCR-SSCP方法对33例慢性粒细胞白血病（Chronicmyeloidleukemia,CML）进行了bcr/abl,p53,p16基因检测.发现33例CML中32例bcr/abl基因阳性（97.0%）,22例慢性期无p53基因突变,而11例急变组中一例外显子5和一例外显子7发生p53基因突变,且均发生于急粒变组,2例出现p16基因纯合缺失,且均发生在急淋变组,33例CML均未见p16基因突变.结果提示bcr/abl基因与CML发生有关,而p53基因突变可能与CML急粒变有关,p16纯合缺失可能与CML急淋变有关.     

Green synthesis and chemical characterization of gold nanoparticle synthesized using Camellia sinensis leaf aqueous extract for the treatment of acute myeloid leukemia in comparison to daunorubicin in a leukemic mouse model

According to chemotherapeutic properties of medicinal plants, pharmacologists have always tried to synthesize and formulate the new chemotherapeutic supplements or drugs of metallic nanoparticles using plants. In this study, Camellia sinensis leaf aqueous extract-based gold nanoparticles (AuNPs) are reported for the first time to exert a dietary therapeutic potential compared to Daunorubicin in an animal model of acute myeloid leukemia. The synthesized AuNPs were characterized using different techniques including UV-Vis., FT-IR spectroscopy, TEM, EDS, FE-SEM, and XRD. DPPH free radical scavenging test was done to evaluate the antioxidant potentials of HAuCl₄, C. sinensis, AuNPs, and daunorubicin. For the analyzing of cytotoxicity effects of HAuCl₄, C. sinensis, AuNPs, and daunorubicin, MTT assay was used on HUVEC, Human HL-60/vcr, 32D-FLT3-ITD, and Murine C1498 cell lines. In vivo design, induction of acute myeloid leukemia was done by 7,12-Dimethylbenz[a]anthracene (DMBA) in 75 mice. Then, the animals were randomly divided into six subgroups, including control, untreated, HAuCl₄, C. sinensis, AuNPs, and daunorubicin. FTIR findings suggested antioxidant compounds in the nanoparticles were the sources of reducing power, reducing gold ions to AuNPs. SEM and TEM images exhibited a uniform spherical morphology and diameters of ~20-30 nm for the nanoparticles. DPPH test revealed similar antioxidant potentials for daunorubicin and AuNPs. These nanoparticles similar to daunorubicin had low cell viability dose-dependently against Human HL-60/vcr, 32D-FLT3-ITD, and Murine C1498 cell lines without any cytotoxicity on HUVEC cell line. AuNPs similar to daunorubicin, significantly (p≤0.05) increased the anti-inflammatory cytokines and the lymphocyte, platelet, and RBC parameters and decreased the weight and volume of liver and spleen, the pro-inflammatory cytokines, and the total WBC, blast, neutrophil, monocyte, eosinophil, and basophil counts, as compared to the untreated mice. According to the above results, it appears that AuNPs can be used as a chemotherapeutic drug for the treatment of acute myeloid leukemia in the clinical trial.     

基于磁珠分离的生物质谱技术用于急性白血病疗效评价

基于磁珠分离的生物质谱技术,建立急性白血病疗效的血清评价方法。通过对质控血清的连续检测分析,日间及日内重复性均达国际标准,确保血清多肽分析体系的稳定性及可靠性。以30例初发急性白血病血清及其治疗缓解血清为研究对象,经金属铜鳌合磁珠分离和基质辅助激光解吸电离飞行时间质谱仪(MAL-DI-TOF-MS)检测后获得血清多肽谱图。经过FlexAnalysis 2.4软件和Biosun_MS软件分析,在m/z1000～10000范围内,得到27个具有统计学意义(p<0.05)的差异峰,其中治疗缓解后表达上调峰10个,表达下调峰17个,利用上述差异峰建立的诊断模型获得了100%敏感性和90%的特异性。基于磁珠分离和MALDI-TOF-MS检测建立的诊断模型具有较高的敏感性和特异性,对差异峰进行测序鉴定将有助于急性白血病疗效评价和发病机理研究。     

Use of MDLC-DIGE and LC-MS/MS to identify serum biomarkers for complete remission in patients with acute myeloid leukemia

The response criteria for complete remission (CR) in acute myeloid leukemia (AML) are currently based on morphology and blood cell counts. However, these criteria are insufficient to establish a diagnosis in cases with poor quality bone marrow (BM) samples demonstrating a loss of cellular morphology. We investigated whether the sera of patients contained biomarkers that indicate disease response status. First, we performed multidimensional liquid chromatography-differential gel electrophoresis (MDLC-DIGE) to generate protein profiles of two pooled, paired serum samples from patients who had achieved CR; one collected at diagnosis (PreCR) and the other collected after chemotherapy (CR). Then, with the biomarker candidates found, ELISA was carried out for individual PreCR and CR samples, and for other verification sets including nonremission (NR) patients and normal samples. We selected two proteins, complement factor H (CFH) and apolipoprotein H (ApoH), with dye (Cy) ratios showing greater than 2.0-fold differences between the pooled samples. ELISA showed that CFH and ApoH are useful for distinguishing between the recovered (CR and normal) and nonrecovered (PreCR, PreNR, and NR) states in AML (p <0.001). We successfully applied a protein profiling technology of MDLC-DIGE and LC-MS/MS to discover two biomarkers for CR which needs further validation for a clinical setting.     

Speciation of arsenic trioxide metabolites in blood cells and plasma of a patient with acute promyelocytic leukemia

Arsenic trioxide (As₂O₃) has been widely accepted as the second-best choice for the treatment of relapsed and refractory acute promyelocytic leukemia (APL) patients. However, a few studies have been conducted on a detailed speciation of As₂O₃ metabolites in blood samples of patients. To clarify the speciation of arsenic, the blood samples were collected at various time points from a patient with APL after remission induction therapy and during consolidation therapy. The total amounts of arsenic in blood cells and plasma, and the plasma concentrations of inorganic arsenic and methylated metabolites were determined by inductively coupled plasma mass spectrometry (ICP-MS) and high-performance liquid chromatography/ICP-MS, respectively. The total amounts of arsenic in the blood cells were 4–10 times higher than those in plasma. Among all arsenic metabolites, the pentavalent arsenate (As^V) in plasma was more readily eliminated. During the drug-withdrawal period, the initial plasma concentrations of trivalent arsenic (As^III) declined more rapidly than those of methylarsonic acid and dimethlyarsinic acid, which are known as the major methylated metabolites of As^III. On the other hand, during the consecutive administration in the consolidation therapy period, the plasma concentrations of total arsenic and arsenic metabolites increased with time. In conclusion, these results may support the idea that methylated metabolites of As₂O₃ contribute to the efficacy of arsenic in APL patients. These results also suggest that detailed studies on the pharmacokinetics as well as the pharmacodynamics of As₂O₃ in the blood cells from APL patients should be carried out to provide an effective treatment protocol. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Presented at the 4th International Conference on Trace Element Speciation in Biomedical, Nutritional and Environmental Sciences, 25–29 May 2008, Munich-Neuherberg, Germany.     

利用原子力显微镜探测人正常与慢性淋巴白血病外周血B淋巴细胞

采用Ficoll密度梯度离心法得到人外周血单个核细胞(PBMC),并结合磁珠分选的方法进一步纯化得到正常B淋巴细胞,探索了正常和肿瘤B淋巴细胞之间的差异。通过应用具有高分辨率的原子力显微镜(AFM)对正常人和慢性淋巴白血病人外周血B淋巴细胞进行成像,并对这两种B淋巴细胞的高度、直径、体积及膜表面的颗粒平均高度、平均粗糙度和颗粒分布进行测量,对比观察两组细胞膜表面宏观和纳米结构的变化。结果表明,慢性淋巴白血病B淋巴细胞比正常的B淋巴细胞高大,细胞膜表面颗粒更大且细胞膜粗糙。此外,对这两组淋巴细胞进行了机械性质方面的测量和统计,结果发现慢性淋巴白血病B淋巴细胞粘附力(524.1±160.0)pN比正常B淋巴细胞粘附力(1091±260)pN约小1倍,且癌变的B淋巴细胞硬度明显比正常的小。当正常细胞癌变时,细胞的形貌、超微结构及骨架会发生一定的改变。实验证明应用AFM可在形态学和机械性质上明显区别正常和慢性淋巴白血病B淋巴细胞,为临床诊断慢性淋巴白血病提供新的技术手段。     

Formal Synthesis of (−)-Englerin A and Cytotoxicity Studies of Truncated Englerins

An efficient formal synthesis of (−)-englerin A ( 1 ) is reported. The target molecule is a recently isolated guaiane sesquiterpene that possesses highly potent and selective activity against renal cancer cell-lines. Our enantioselective strategy involved the construction of the BC ring system of compound 1 through a Rh^II-catalyzed [4+3] cycloaddition reaction followed by subsequent attachment of the A ring through an intramolecular aldol condensation reaction. As such, this strategy allows the synthesis of truncated englerins. Evaluation of these analogues with the A498 renal cancer cell-line suggested that the A ring of englerin is crucial to its antiproliferative activity. Moreover, evaluation of these analogues led to the identification of potent growth-inhibitors of CEM cells with GI₅₀ values in the range 1–3 μM .     

The cellular labeling and pH-sensitive responsive-drug release of celastrol in cancer cells based on Cys-CdTe QDs

As one of the active compounds derived from Traditional Chinese Medicine,Celastrol(CSL)had cytotoxicity for human leukemia cancer cells K562 and its multidrug-resistant cell line K562/A02.Here,we introduced cysteamine-modified CdTe QDs as the labeling and drug carrier into CSL research and found that the self-assembly and conjugation of anticancer molecular CSL with the Cys-CdTe QDs could significantly increase the drug’s cytotoxicity for K562 cells.More important,these CSL-Cys-CdTe nanocomposites could overcome the multidrug resistance of K562/A02 cells and efficiently inhibit the cancer cell proliferation by realizing the pH-sensitive responsive release of CSL to cancer cells.The enhanced cytotoxicity was caused by the increase of the G2/M phase arrest for K562/A02 cells as well as for K562 cells.Cys-CdTe QDs can readily bind on the cell plasma membranes and be internalized into cancer cells to trace and detect human leukemia cancer cells in real time.In addition,these Cys-CdTe QDs can facilitate the inhibition of the multidrug resistance of K562/A02 cells and readily induce apoptosis.As a good photosensitizer for the therapy,labeling,and tracing of cancer cells,the combination of CSL with Cys-CdTe QDs can optimize the use of and a new potential therapy method for CSL and yield new tools to explore the mechanisms of active compounds from Traditional Chinese Medicine.     

Capillary electrophoresis method for plasmatic determination of imatinib mesylate in chronic myeloid leukemia patients

Imatinib (IMAT) is a tyrosine kinase inhibitor that has been used for the treatment of chronic myeloid leukemia (CML). Despite the efficacy of IMAT therapy, some cases of treatment resistance have been described in CML. Developing a plasma method is important since there are several studies that provided a higher correlation between IMAT plasma concentration and response to treatment. Therefore, in this investigation we validated a method by CE as an alternative, new, simple and fast electrophoretic method for IMAT determination in human plasma. The analysis was performed using a fused silica capillary (50 μm id×46.5 cm total length, 38.0 cm effective length); 50 mmol/L sodium phosphate buffer, pH 2.5, as BGE; hydrodynamic injection time of 20 s (50 mbar); voltage of 30 kV; capillary temperature of 35°C and detection at 200 nm. Plasma samples pre-treatment involved liquid-liquid extraction with methyl-tert-butyl ether as the extracting solvent. The method was linear from 0.125 to 5.00 μg/mL. The LOQ was 0.125 μg/mL. Mean absolute recovery of IMAT was 67%. The method showed to be precise and accurate with RSD and relative error values lower than 15%. Furthermore, the application of the method was performed in the analysis of plasma samples from CML patients undergoing treatment with IMAT.