CD34阳性成人急性髓系白血病临床及生物学研究

检测了４０例成人初治急性髓系白血病细胞ＣＤ３４抗原的表达，并分析了ＣＤ３４表达与一般临床特征，其它免疫标志，。染色体异常及治疗效果的关系。结果显示，４０例ＡＭＬ中ＣＥ３４阳性表达１７例，占４２．５％，除Ｍ３无ＣＤ３４表达外，ＣＤ３４表达与ＦＡＢ亚型无关。     

急性髓细胞白血病细胞CD34抗原表达的临床及生物学意义

为研究ＣＤ３４在急性髓细胞白血病（ＡＭＬ）中的临床及生物学意义，用流式细胞术检测了１０７例ＡＭＬＣＤ３４表达。结果显示４８．５％的患者ＣＤ３４阳性表达，Ｍ３（１５．８％）低于其他亚型（ｐ〈０．０５）。ＣＤ３４＾＋者肝脾肿大常见。ＣＤ３４表达在ｔ（８；２１）、正常核型及ｔ（１５；１７）组分别为７２．０％、４０?７％及１７．６％。ＣＤ３４＾＋者的完全缓解率（４１．９％）明显低于ＣＤ３４＾－者的５９．６     

Hes1对急性髓系白血病患者骨髓CD34+CD38-细胞的作用及其机制研究

  目的  研究Hes1对急性髓系白血病(AML)患者骨髓CD34+CD38-细胞的作用及其机制。  方法  收集初治AML患者及正常供者骨髓样本后,通过密度梯度离心法获取单个核细胞,流式细胞术检测CD34+CD38-细胞比例及其细胞周期。通过免疫磁珠法分选CD34+CD38-细胞后,体外集落形成实验(CFC)检测其增殖能力,并通过Realtime PCR检测其Hes1的表达量。构建Hes1过表达逆转录病毒载体,感染正常供者骨髓CD34+细胞后,流式细胞术分析其细胞周期的改变,CFC检测其增殖的改变。  结果  AML患者骨髓CD34+CD38-细胞比例明显低于正常对照,流式细胞术结果显示患者来源CD34+CD38-细胞大多数进入静止期,CFC结果显示患者CD34+CD38-细胞体外扩增能力下降。Realtime PCR结果发现患者CD34+CD38-细胞中Hes1表达上调。提高正常供者CD34+细胞中Hes1的表达后,细胞增殖减少,进入静止期。  结论  在AML中CD34+CD38-细胞比例下降,进入静止期,与Hes1的表达上调有关。      

治疗急性髓系白血病的树突状细胞疫苗的研究进展

摘 要 本文综述了免疫治疗急性髓系白血病（acute myeloid leukemia, AML）的树突状细胞（dendritic cells, DC）疫苗的近期研究进展，主要涉及各种类型的白血病肿瘤抗原的确认与制备，包括AML特异融合蛋白及相关多肽、癌症睾丸抗原、酸洗脱多肽、白血病细胞冻融抗原/凋亡细胞、AMLDC融合细胞、白血病细胞来源的DC等；DC的制备与特性以及优化方法；临床试验研究的初步结果等，还对今后的研究方向进行了分析和展望。     

米托蒽醌方案对CD+34高表达急性淋巴细胞白血病的临床疗效观察

目的 探讨米托蒽醌(MTZ)在急性淋巴细胞白血病(ALL)化疗中的作用特点。方法 50例CD34^ 抗原高表达的ALL上,随机选择二种化疗方案(CPMP,CODP)。联合化疗1个疗程后分别比较CR率、骨髓抑制及其他毒副作用;同时将骨髓白血病细胞体外培养72h。进行药物杀伤效应实验。分别比较MTZ、柔红霉素(DNR)在体外对ALL细胞不同分化阶段的抑制作用。结果 CD34抗原高表达的ALL中,以MTZ为主的COMP方案的1个疗程缓解率为92．6％。比CODP方案高27．4％。体外白血病细胞培养加药物抑制试验结果显示,MTZ对分化较早阶段的CD34^ ALL细胞的抑制显著高于DNR。结论 MTZ对ALL具有较强的抗白血病活性,临床骨髓抑制明显。上述特点可能与其主要作用于白血病细胞的分化较早阶段有关。     

慢性粒细胞白血病CD+34细胞的生物学特性

慢性粒细胞白血病(CML)是一种起源于骨髓多能造血干细胞的恶性克隆性疾病,其中90%以上的病例具有ph染色体,其分子基础是bcr/abl基因重排.CD34抗原是一种表达于造血干/祖细胞的膜表面糖蛋白,正常CD+34细胞亚群中90%以上为祖细胞和干细胞,骨髓中约1.5%的单个核细胞表达CD34抗原,未经刺激的外周血中0.1%～0.2%的单个核细胞为CD+34,体内单独或联合应用造血生长因子能够提高外周血中CD+34细胞的数量,并有效地应用于外周血干细胞移植(PBSCT).     

CD_（34）阳性成人急性髓系白血病临床及生物学研究

检测了４０例成人初治急性髓系白血病（ＡＭＬ）细胞ＣＤ＿（３４）抗原的表达，并分析了ＣＤ＿（３４）表达与一般临床特征、其它免疫标志、染色体异常及治疗效果的关系。结果显示，４０例ＡＭＬ中ＣＤ＿（３４）阳性表达１７例，占４２．５％，除Ｍ３无ＣＤ＿（３４）表达外，ＣＤ＿（３４）表达与ＦＡＢ亚型无关。组与组在年龄、性别、初诊时白细胞数、血红蛋白浓度、血小板计数、肝脾肿大及染色体异常均无显著差异。除淋系抗原在组表达明显增高外，其它髓系抗原表达在两组间无差异。组完全缓解率为４１．４８％，明显低于组的７３．９１％。研究提示，ＡＭＬ可能起源于较早期的造血于细胞恶性转化，检测ＣＤ＿（３４）表达对判断ＡＭＬ疗效有一定价值。     

急性髓系白血病患者来源白血病干细胞的分选与鉴定

 目的　分选急性髓系白血病患者白血病干细胞并进行鉴定,为白血病干细胞的进一步研究奠定基础。方法　采用Ficoll密度梯度离心法从患者骨髓中分离单个核细胞。采用流式细胞术从单个核细胞中分选CD+34 CD+123白血病干细胞,通过集落形成能力和鹅卵石形成能力检测分选后白血病干细胞的自我更新与分化能力,同时检测CD+34 CD+123 细胞纯度,观察其细胞形态。结果　分选后的CD+34 CD+123 白血病干细胞比例占总分选细胞的10.7 %,具有集落形成能力和鹅卵石形成能力,CD+34 CD+123 细胞纯度为62.1 %。结论　成功分选与鉴定了白血病干细胞,可用于其后的进一步研究工作。     

以重组人类CD40配体为主的细胞因子体外诱导急性髓系白血病完全缓解患者及健康人外周血单个核细胞来源的树突状细胞形态及表型分析

 【摘要】　目的　以健康人AB血清与以重组人类CD40配体（rhCD40L）为主的细胞因子体外诱导急性髓系白血病完全缓解（AML-CR）患者及健康人外周血的单个核细胞,并分析其形态及表型差异。方法　以健康人AB血清为基础的培养体系中加入GM-CSF、rhIL-4、rhCD40L等细胞因子,诱导单个核细胞分化形成树突状细胞（DC）。瑞特-吉姆萨染色观察形态;流式细胞术鉴定表型;MTT法检测DC对淋巴细胞抗原刺激能力;ELISA法检测DC培养上清IL-12的分泌。结果　培养7 d后的细胞具有典型的MoDC形态,并上调表达MoDC特征性表面分子CD83及共刺激分子CD40、CD80、CD86,AML-CR组与健康人组相比差异无统计学意义（均P＞0.05）,但加rhCD40L组较未加rhCD40L组间差异有统计学意义（均P＜0.05）。培养后的MoDC可较强地刺激同种自体淋巴细胞增殖,AML-CR组与健康人组间刺激淋巴细胞增殖作用的差异无统计学意义（P＞0.05）,但加rhCD40L组较未加rhCD40L组间差异均有统计学意义（均P＜0.05）。培养的MoDC自培养第5天始即有IL-12的分泌,AML-CR组与健康人组相比IL-12分泌水平差异无统计学意义（P＞0.05）,加rhCD40L组与未加rhCD40L组间差异有统计学意义（P＜0.05）。结论　以rhCD40L为主的细胞因子体外可诱导AML-CR患者及健康人外周血DC生成,两者在形态及表型上无明显差异,DC可刺激白血病特异毒性T细胞增殖,尤以由rhCD40L诱导的DC作用明显。     

Independent prognostic impact of CD15 on complete remission achievement in patients with acute myeloid leukemia

The prognostic role of CD15 in acute myeloid leukemia (AML) has been tested in different studies with conflicting results. To address this issue, we retrospectively evaluated a cohort of 460 AML patients of all ages with the exclusion of acute promyelocytic leukemia (M/F 243/217, median age 50.6 years [range 0.9‐81.2]) intensively treated at our institute between January 1999 and December 2010. CD15 positivity was found in 171 of 406 evaluable patients (42.1%). Complete remission (CR) was achieved by 334 patients (72.6%), while 82 (17.8%) were resistant and 44 (9.6%) died during induction: the median CR duration was 15.5 months (range 0.6‐176.0), with 2‐year disease‐free survival rate of 45.1% (95% confidence interval 39.6‐50.6). The median overall survival was 14.4 months (range 0.3‐177.0), with 2‐year overall survival rate of 42.2% (95% confidence interval 37.5‐46.9). At univariate analysis for CR achievement, age < 60 years (P < .001), World Health Organization classification (P = .045), low‐risk karyotype (P < .001), no high‐risk karyotype (P = .006), positivity for AML‐ETO (P = .004)/CBFβ‐MYH11 (P = .003)/CD15 (P = .006)/CD11b (P = .013), negativity for FLT3‐ITD (P = .001), Hb > 8 g/dL (P = .020), and white blood cell < 50 × 10⁹/L (P = .034) had a favorable impact. At a multivariate logistic regression model, CD15 positivity (P = .002), age < 60 years (P = .008), white blood cell < 50 × 10⁹/L (P = .017), and low‐risk/no high‐risk karyotype (P = .026/P = .025) retained an independent prognostic role on CR achievement . The baseline assessment of CD15 positivity appears to have a role in the risk evaluation for CR achievement in AML patients undergoing intensive chemotherapy and should be assessed in prospective studies together with other clinical and biologic features already reported.     

Evaluation of residual CD34+Ph+ progenitor cells in chronic myeloid leukemia patients who have complete cytogenetic response during first‐line nilotinib therapy

BACKGROUND:

Compared with imatinib, nilotinib is a potent breakpoint cluster region/v‐abl Abelson murine leukemia viral oncogene (bcr‐abl) kinase inhibitor, and it induces higher rate and rapid complete cytogenetic response (CCyR), yet no clinical data are available regarding its efficacy against chronic myeloid leukemia (CML) stem cells. Earlier studies demonstrated that clusters of differentiation 34–positive, Philadelphia chromosome–positive (CD34⁺Ph⁺) cells are detectable in about 45% of patients with CML, despite being on long‐term imatinib therapy and having achieved sustained CCyR.

METHODS:

CD34⁺ cells from bone marrow of de novo CML patients in the chronic phase (n = 24) treated with nilotinib (median duration of therapy, 22 months) were isolated and scored for BCR‐ABL by fluorescent in situ hybridization (FISH) analysis. Similar analysis was also performed in 5 de novo CML chronic phase patients who achieved CCyR within 3 months of nilotinib therapy.

RESULTS:

FISH evaluation of a median of 100 CD34⁺ nuclei per patient revealed that only 1 of 20 (5%) evaluable patients showed residual Ph⁺ progenitor cells. In this patient, just 1 of 140 (0.7%) CD34⁺ interphase nuclei was found to be positive for BCR‐ABL. Surprisingly, no CD34⁺Ph⁺ cells were found even in those 5 patients evaluated after 3 months of nilotinib treatment.

CONCLUSIONS:

This study assessed for the first time the persistence of CD34⁺Ph⁺ cells during nilotinib first‐line treatment. Preliminary results showed that in patients in CCyR, even after short‐term nilotinib therapy, residual leukemic progenitors are very rarely detected compared with imatinib‐treated CCyR patients. It is yet to be determined if these findings will have an impact in the path to a cure of CML with tyrosine kinase inhibitors. Cancer 2012. © 2012 American Cancer Society.     

CD34阳性及阴性成年人急性T淋巴细胞白血病临床分析

目的 分析CD34阳性及阴性成年急性T淋巴细胞白血病(T-ALL)患者的临床特点及预后,探讨CD34表达对T-ALL预后的价值.方法 回顾性分析郑州大学第一附属医院血液科2012年1月至2015年7月收治的75例初治成年T-ALL患者.根据CD34的表达情况,分为CD34阳性组与阴性组,对两组患者的临床特点及预后进行比较.结果 75例初治成年T-ALL患者中,CD34阳性组24例(32.0％),CD34阴性组51例(68.0％).CD34阳性组与阴性组在性别、年龄、肝脾大、淋巴结肿大、血小板减少、白细胞增高、染色体异常、4周完全缓解(CR)率、中枢神经系统白血病(CNSL)方面,差异均无统计学意义(均P＞ 0.05);初诊时CD34阳性组血红蛋白(Hb) ＜90g/L、伴髓系抗原表达者的比例高于阴性组,差异均有统计学意义(x 2=5.888,P=0.015;x 2=10.758,P=0.001).18例选择造血干细胞移植(HSCT),57例未选择HSCT.在未选择HSCT的患者中,CD34阳性组中位生存期为5个月,阴性组为32个月,两者差异有统计学意义(x2=9.172,P=0.002).结论 成年T-ALL患者CD34表达与Hb＜ 90 g/L及髓系抗原表达有关;未选择HSCT患者的CD34表达可能与预后呈负相关.     

CD14+单核细胞系白血病细胞来源树突细胞体外刺激特异抗白血病T细胞应答

背景与目的:白血病细胞能在体外分化为树突细胞(dendriticcells,DCs),从而有希望用于白血病的免疫治疗。本研究旨在探讨CD14高表达的单核细胞系白血病(M4、M5)细胞分化而来的DCs体外诱导抗白血病T细胞应答的能力。方法:取5例初诊CD14高表达的M4或M5型白血病患者的骨髓标本,分离单个核细胞(bonemarrowmononuclearcells,BMMNCs),将白血病细胞分为3组:贴壁白血病细胞组、非贴壁白血病细胞组及总白血病细胞组。流式细胞术(flowcytometry,FCM)比较3组细胞的CD14表达。用含GM-CSF、IL-4和TNF-α或不含细胞因子的培养液培养细胞7～10天后,通过细胞形态学观察及FCM检测细胞表型,鉴定单核白血病细胞来源的DCs(monocyticleukemiacell-deriveddendriticcells,Mo-LDCs);采用异基因混合淋巴细胞反应(allogeneicmixedlymphocytereaction,Allo-MLR)以及细胞毒性T淋巴细胞(cytotoxicTlymphocytes,CTL)抗白血病细胞毒分析检测Mo-LDCs的免疫功能,染色体核型分析结合异常表面抗原确定Mo-LDCs的白血病来源。结果:3组中贴壁白血病细胞的CD14含量最高,在细胞因子联合诱导下,可分化为大量CD83 成熟DCs。在同一病例的3组细胞以及不同病例的总单核细胞组间,培养前CD14的表达率与诱导后CD83 DCs的产率成正相关(r=0.967,P=0.007)。Mo-LDCs具有典型的成熟DCs的形态及表型特征,在Allo-MLR中能刺激同种T细胞明显增殖,并能刺激扩增特异性抗白血病CTL。同时,Mo-LDCs持续存在所起源白血病的核型异常和异常表达的髓系抗原。结论:在细胞因子组合诱导下,M4、M5亚型AML的CD14 细胞可分化为具有免疫功能的Mo-LDCs,单核系白血病细胞的CD14表达高低可能预示其DCs分化能力。Mo-LDCs具有经典的DCs的表型及功能,还具有白血病的克隆异常,可用于M4、M5患者的免疫治疗。     

Comorbidity is an independent predictor of complete remission in elderly patients receiving induction chemotherapy for acute myeloid leukemia

BACKGROUND.: Elderly patients with acute myeloid leukemia (AML) have a poor prognosis, which is explained by the disease itself and by host-related factors. The objective of this study was to determine the prognostic role of comorbidities in this population. METHODS.: For this single-center, retrospective study, the authors analyzed the outcome of 133 patients aged >/=70 years who received induction chemotherapy for nonpromyelocytic AML between 1995 and 2004. Comorbidities were evaluated by using an adapted form of the Charlson comorbidity index (CCI). RESULTS.: The median patient age was 73 years. The CCI score was 0 for 83 patients (68%), 1 for 16 patients (13%), and >1 for 23 patients (19%). The complete remission (CR) rate was 56%, and the median overall survival was 9 months. In multivariate analysis, 4 adverse prognostic factors for CR were identified: unfavorable karyotype, leukocytosis >/=30 g/L, CD34 expression on leukemic cells, and CCI >1. A score could be generated to allow the stratification of patients into low-, intermediate-, and high-risk groups with CR rates of 87%, 63%, and 37%, respectively. The risk of early mortality and the probability of survival also were different in the 3 risk groups (P = .02 and P = .01, respectively). CONCLUSIONS.: The results from this study indicated that associated comorbidities are independent factors that may influence achievement of CR in elderly patients with AML. Such a scoring system may be useful in the prognostic staging systems that are used to identify patients with AML who can benefit from induction chemotherapy.     

急性髓细胞性白血病细胞培养上清对树突细胞分化、成熟、凋亡及功能的影响

背景与目的:树突细胞(dendritic cell,DC)存机体抗瘤免疫中起重要作用.研究已表明肿瘤患者和荷瘤动物体内DC功能受损并伴有DC数量的降低.最近的研究提示这种DC功能的受损可能是肿瘤细胞分泌的可溶性因子介导的免疫抑制所致.本研究拟观察急性髓系白血病(acute myeloid leukemia,AML)细胞分泌的可溶性因子对单核细胞来源DC的分化、成熟、凋亡及其功能的影响.方法:原代AML细胞培养24 h后收集上清.在含有或不含有AML细胞培养上清的培养液中,利用IL-4和GM-CSF刺激单核细胞分化成不成熟DC(immature DC,iDC),IL-1β、IL-6、TNF-α和PGE-2促进DC成熟.流式细胞仪检测DC表型和凋亡率的变化.计算前体细胞频率(precursorfrequency,PF),观察DC对异基因CD4 T细胞和CD8 T细胞的刺激功能.结果:AML细胞培养上清可显著抑制DC表面协同刺激分子CD80和CD86的表达,并可降低促成熟细胞因子对DC的促成熟作用.同时,与对照组相比,AML细胞培养上清可显著诱导DC的凋亡,凋亡率分别为(15.1±4.2)%和(29.4±9.5)%(P＜0.01).相对于正常成熟DC,AML细胞培养上清诱生的DC对异基因CD4 T细胞和CD8 T细胞的刺激功能显著降低(P＜0.01),其中CD4 T细胞的前体细胞频率分别为(5.2±1.6)%和(1.8±0.5)%,CD8 T细胞的前体细胞频率分别为(6.5±2.0)%和(2.1±0.6)%.结论:AML细胞分泌的可溶性因子可抑制DC的发育和功能.     

成人急性白血病中CD117与CD34共表达的临床意义

背景与目的:c-kit受体(c-kitreceptor,c-kitR,CD117)是干细胞因子受体。CD117在急性非淋巴细胞白血病(acutenon-lymphoblasticleukemia,ANLL)中高表达,可作为髓系免疫学标记物,对诊断ANLL有一定参考价值。但是,CD117也可在部分急性淋巴细胞白血病(acutelymphoblasticleukemia,ALL)中表达。CD34为造血干(祖)细胞抗原标记物,在ANLL和ALL中均有高表达。本研究旨在探讨CD117和CD34在急性白血病中共表达的临床意义。方法:采用流式细胞术(flowcytometery,FCM)分别检测92例ALL和81例ANLL初诊患者骨髓单个核细胞(BMMNC)CD117的阳性率和阳性细胞水平;比较ALL和ANLL患者CD117/CD34共表达率的差异,并比较ALL患者中CD117和CD117/CD34共表达率的差异。设立20例健康成人为对照组。结果:在ALL和ANLL患者中CD117阳性率分别为15.2%和71.6%,CD117/CD34共表达率分别为5.4%和55.5%,差异有显著性(P<0.001)。ALL患者中CD117表达率和CD117/CD34共表达率分别为15.2%和5.4%,差异有显著性(P=0.029)。结论:CD117可作为急性白血病的MIC分型诊断之髓系免疫学标志,用以协助ANLL的临床诊断;较之CD117表达,CD117/CD34在ALL中的共表达率更低,可籍此协助排除ALL。     

Polymeric nanoparticle-mediated silencing of CD44 receptor in CD34 acute myeloid leukemia cells

The adhesion receptor CD44 plays an important role in the survival and retention of leukemic stem/progenitor cells (LSPC) within the bone marrow (BM) niche, as well as in the high relapse rates of acute myeloid leukemia (AML). Down-regulating CD44 could be clinically relevant not only for suppression of the deregulated function of LSPC but also in LSPC response to chemotherapeutic agents. Small interfering RNA (siRNA) delivery is a promising approach for AML treatment, and we recently reported effective siRNA delivery into difficult-to-transfect AML cell lines using lipid-substituted polyethylenimine/siRNA complexes (polymeric nanoparticles). In this study, we investigated polymeric nanoparticle-mediated silencing of CD44 in CD34⁺ LSPC cell models (leukemic KG-1 and KG-1a cell lines) as well as primary AML cells. Polymeric nanoparticle-mediated silencing decreased surface CD44 levels in KG-1, KG-1a and primary AML cells by up to 27%, 30% and 20% at day 3, respectively. Moreover, CD44 silencing resulted in induction of apoptosis in KG-1 cells, reduced adhesion of KG-1 and KG-1a cells to hyaluronic acid-coated cell culture plates and BM-MSC, and decreased adhesion of primary AML cells to BM-MSC. Our results suggest that polymeric nanoparticle-mediated silencing of CD44 might be a useful technique for inhibiting LSPC interactions with their microenvironment, thereby prohibiting leukemia progression or sensitizing LSPC to chemotherapy.