Having a higher blast percentage in circulation than bone marrow: clinical implications in myelodysplastic syndrome and acute lymphoid and myeloid leukemias.

Determining the percentage of peripheral blood (PB) and bone marrow (BM) blasts is important for diagnosing and classifying acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). Although most patients with acute leukemia or MDS have a higher percentage of BM blasts than PB blasts, the relative proportion is reversed in some patients. We explored the clinical relevance of this phenomenon in MDS (n = 446), AML (n = 1314), and acute lymphoblastic leukemia (ALL) (n = 385). Among patients with MDS or ALL, but not AML, having a higher blast percentage in PB than in BM was associated with significantly shorter survival. In multivariate analyses, these associations were independent of other relevant predictors, including cytogenetic status. Our findings suggest that MDS and ALL patients who have a higher percentage of PB blasts than BM blasts have more aggressive disease. These data also suggest that MDS classification schemes should take into account the percentage of blasts in PB differently from the percentage of blasts in BM.     

Telomerase activity in myelodysplastic syndromes

Myelodysplastic syndromes are clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis and peripheral cytopenias. Telomeres are thought to be critical in maintaining normal hematopoiesis. In this study, we assessed telomere dynamics in order to obtain further insight into the pathogenesis of MDS. We studied telomerase activity (TA) in mononuclear cells from peripheral blood (PB) and bone marrow (BM) from patients with myelodysplastic syndrome (MDS; n=24), acute myeloid leukemia (AML; n=14), chronic myeloid leukemia (CML; n=12) and 11 normal controls using a polymerase chain reaction-based telomeric repeat amplification assay. Telomerase activities (mean+/-S.D.) were found as 0.199+/-0.09, 0.414+/-0.55, 0.253+/-0.26 and 0.181+/-0.05 pg/ml in PB mononuclear cells, respectively (P>0.05). Comparison of TA of BM mononuclear cells from 19 MDS patients versus 10 BM samples from normal controls revealed no significant difference (P=0.3). There was no correlation between the levels of TA and clinical and prognostic parameters of the patients with MDS, such as degree of anemia, platelet counts on presentation, gender, presence of organomegaly, bone marrow fibrosis and BM blast percentages. Patients who had higher TA had significantly inferior survival compared with patients who had lower TA (P=0.005). Consistent with previous data, our results suggest that in patients with MDS, telomerase activity might be insufficient to compensate for the telomere shortening. Furthermore, TA might be prognostically important in patients with MDS. Measurements of enzymatic activity in association with telomere length studies may help to understand the prognostic role of telomere dynamics in patients with myelodysplastic syndromes more reliably.     

Exit of pediatric pre-B acute lymphoblastic leukaemia cells from the bone marrow to the peripheral blood is not associated with cell maturation or alterations in gene expression

Background  

Childhood pre-B acute lymphoblastic leukemia (ALL) is a bone marrow (BM) derived disease, which often disseminates out of the BM cavity, where malignant cells to a variable degree can be found circulating in the peripheral blood (PB). Normal pre-B cells are absolutely dependent on BM stroma for survival and differentiation. It is not known whether transformed pre-B ALL cells retain any of this dependence, which possibly could impact on drug sensitivity or MRD measurements.     

Increased expression of AML1-a and acquired chromosomal abnormalities in childhood acute lymphoblastic leukemia

A semi-quantitative expression analysis of both AML1-a and AML1-total was performed by RT-PCR in 19 children with acute lymphoblastic leukemia (ALL) at diagnosis. AML1-a expression was assessed in 16 bone marrow (BM) and 13 peripheral blood (PB) samples whereas AML1-total was assessed in 17 BM and 16 PB samples. These analyses were also carried out in 15 PB samples of healthy controls. In addition, 18/19 patients were karyotyped: 11 had an unmodified constitutional karyotype (CK) and seven exhibited acquired chromosomal abnormalities (ACA). The expression of AML1-a was significantly increased in BM and PB when compared with the controls (p < 0.013 and p < 0.035, respectively). A significant increase was found in the expression of AML1-a in BM of the ACA group compared with the CK group (p < 0.0009). The expression of AML1-a in BM and PB showed a significant increase in the ACA group compared with controls (p < 0.00001 and p < 0.012, respectively); in contrast, the CK group did not differ from the controls. These observations may mean that the increase of AML1-a favours the progression of leukemia.     

Telomerase activity and hTERT mRNA expression in acute leukemia

Telomerase is specifically activated in mostmalignant tumors but is usually inactive in normalsomatic cells, suggesting that telomerase plays animportant role in cellular immortalization andtumorigenesis. Telomerase activity has been the mostgeneral molecular marker for the identification ofhuman cancer and can be detected in 85% of alltumors[1,2]. Although several studies have investigatedtelomerase activity in leukemia, few studies haveinvestigated the diagnostic importance of hTERTexpressi…     

急性白血病患者骨髓及外周血内皮祖细胞的变化及其临床意义

目的 探讨内皮祖细胞(EPC)在急性白血病(AL)患者骨髓及外周血中数量变化及其临床意义.方法 采用流式细胞术(FCM)对43例初治AL患者骨髓及外周血中的EPC进行检测,计算绝对计数,以10例良性血液病患者为对照组.结果 治疗前AL患者骨髓及外周血EPC绝对计数分别为(119.46±72.23)个/μl、(13.69±8.26)个/μl,较对照组的(23.21±12.59)个/μl、(1.86±1.18)个/μl升高(P＜0.01).治疗前AL患者骨髓EPC绝对计数较外周血升高(P＜0.01).完全缓解(CR)组患者治疗后骨髓及外周血EPC绝对计数分别为(26.32±17.44)个/μl、(2.54±2.12)个/μl,较治疗前的(113.18±69.22)个/μl、(14.45±10.76)个/μl降低(P＜0.05),与对照组间差异无统计学意义(P＞0.05);未缓解(NR)组治疗后骨髓及外周血EPC绝对计数[(110.02 ± 67.28)个/μl、(10.04±9.51)个/μl]较对照组增高(P＜0.05),与治疗前差异无统计学意义(P＞0.05).急性髓系白血病(AML)组与急性淋巴细胞白血病(ALL)组间骨髓及外周血中EPC绝对计数差异均无统计学意义(均P＞ 0.05).AL患者外周血EPC绝对计数与βτ微球蛋白、乳酸脱氢酶呈正相关性(均P＜ 0.05).结论 AL患者骨髓及外周血中EPC计数明显增高,其水平可能与AL患者病情发展、疗效及预后有一定关系.     

Transplantation of peripheral blood stem cells as compared with bone marrow from HLA-identical siblings in adult patients with acute myeloid leukemia and acute lymphoblastic leukemia.

PURPOSE: Several studies show that allogeneic peripheral blood stem cells (PBSCs) engraft more rapidly than bone marrow (BM). However, the data are inconsistent with regard to acute and chronic graft-versus-host disease (GVHD), relapse, transplant-related mortality (TRM), and leukemia-free survival (LFS). PATIENTS AND METHODS: Between January 1994 and December 2000, 3,465 adult patients (older than 15 years of age) were reported to the European Group for Blood and Marrow Transplantation Registry from 224 centers. Among acute myeloid leukemia (AML) patients, 1,537 patients received BM and 757 patients received PBSC. In acute lymphoblastic leukemia (ALL) patients, the corresponding figures were 826 versus 345 patients who were analyzed for engraftment, GVHD, TRM, relapse, LFS, and survival. RESULTS: In multivariate analysis, the recovery of neutrophils and platelets was faster with PBSC than with BM (P <.0001). Chronic GVHD was associated with PBSC in patients with AML (relative risk [RR], 2.11; 95% confidence interval, 1.66 to 2.7; P <.0001) and ALL (RR, 1.56; 95% confidence interval, 1.09 to 2.27; P =.02). PBSC versus BM in patients with AML or ALL was not significantly associated with acute GVHD, TRM, relapse, survival, or LFS. In multivariate analysis of patients with AML, factors significantly associated with improved LFS included first remission at transplant (P <.0001), promyelocytic leukemia (M3) versus other French-American-British types (P <.0001), and donor age below median 37 years (P =.02). In patients with ALL, first remission (P <.0001) and methotrexate included in the immunosuppressive regimen (P =.001) were associated with improved LFS. CONCLUSION: Allogeneic PBSC results in faster neutrophil and platelet engraftment and a higher incidence of chronic GVHD than BM. However, acute GVHD, TRM, relapse, survival, and LFS were similar in patients receiving PBSCs versus BM.     

急性髓系白血病患者造血微环境中EPCs数量、微血管密度与VEGFR-2表达的意义北大核心CSCD

目的探讨急性髓系白血病(AML)患者外周血及骨髓中内皮祖细胞(EPCs)的数量变化以及骨髓活检标本中微血管密度(MVD)和血管内皮生长因子受体-2(VEGFR-2)表达的临床意义。方法采用流式细胞仪检测30例AML患者以及10例健康对照组外周血及骨髓中EPCs的数量,进行相对及绝对计数;应用常规石蜡包埋骨髓病理切片进行免疫组织化学染色检测30例AML患者骨髓活组织中MVD和VEGFR-2表达水平,分析其与临床特征的关系。结果 (1)治疗前AML患者外周血和骨髓中EPCs计数与对照组比较,差异有统计学意义(P<0.01);治疗后AML患者完全缓解(CR)组与未缓解(NR)组外周血中EPCs比较,差异有统计学意义(P<0.01);CR与NR组骨髓中EPCs计数比较差异也有统计学意义(P<0.05)。(2)AML骨髓切片上MVD和VEGFR-2表达水平与健康对照组比较差异有统计学意义(P<0.05)。经Spearman秩相关检验,EPCs绝对计数、MVD、VEGFR-2三者之间呈正相关关系。(3)观察外周血中EPCs绝对计数≥20个/微升以及<20个/微升两组生存曲线,以100周为观察终点,<20个/微升组生存期明显优于前者。(4)经Cox回归分析,发现治疗前外周血EPCs绝对计数、WBC计数、VEGFR-2、β2-微球蛋白为危险因素。骨髓巨核细胞数与ECOG评分为保护因素。结论 EPCs计数结合MVD和VEGFR-2检测对评价AML血管内皮功能可能有一定的临床意义,可作为判断患者疗效及预后的指标之一。     

Asparagine and aspartic acid concentrations in bone marrow versus peripheral blood during Berlin-Frankfurt-Münster-based induction therapy for childhood acute lymphoblastic leukemia

A recent study suggested that mesenchymal cells in bone marrow (BM) may counteract l-asparaginase (L-Asp)-containing acute lymphoblastic leukemia (ALL) therapy by secreting asparagine. Herein, we compared asparagine and aspartic acid concentrations in the BM and peripheral blood (PB), in order to determine whether this in vitro observation could be translated into in vivo differences of amino acid levels between both compartments. Asparagine and aspartic acid concentrations in BM (days 15 and 33) and PB (days 12, 15 and 33) were measured during L-Asp-containing Berlin-Frankfurt-Münster (BFM)-based 5-week multi-agent remission induction therapy in 11 children diagnosed with ALL at the St. Anna Children's Hospital in Vienna, Austria. The level of asparagine depletion did not differ significantly between both compartments at any time point measured, but aspartic acid concentrations were significantly higher in BM than PB at days 15 and 33 (p < 0.05). In the context of the reported mesenchymal asparagine production in BM, an increased asparagine production may indeed take place in BM. However, it may be overcome by continuous action of L-Asp, which is mirrored by increased aspartic acid levels but unchanged low asparagine levels in BM, suggesting a higher BM turnover of asparagine generated by L-Asp during induction therapy.     

替尼泊苷联合方案治疗交叉表达双系相关抗原急性白血病的临床研究

目的 :探讨替尼泊苷联合方案治疗交叉表达淋系和髓系抗原急性白血病的化疗疗效。方法 :应用替尼泊苷联合阿糖胞苷、环磷酰胺、长春地辛及泼尼松组成 TA或 TA+COP方案 ,治疗伴淋系抗原表达的急性髓细胞白血病 (L y+ AML) 2 4例和伴髓系抗原表达的急性淋巴细胞白血病(My+ AL L) 1 7例。结果 :完全缓解 (CR)率为 5 6 .1 % (2 3/ 4 1 ) ,总有效率为 80 .5 % (33/ 4 1 )。与常规方案 (DA、VDL P)诱导疗效相比有显著性差异 (P<0 .0 1 )。其中 L y+ AML 的 CR率为 5 8.3% ,总有效率为 79.2 % ;My+ AL L 的 CR率为 5 2 .9% ,总有效率为 82 .4 %。复治性 L y+ AML / My+ AL L 总有效率为 72 .7% (8/ 1 1 )。两种方案的骨髓抑制明显 ,毒副作用能够耐受。结论 :L y+ AML 和 My+ AL L不宜采用常规诱导缓解方案 ,TA和 TACOP方案宜作为此类患者的首选治疗方案     

Acute myeloblastic leukemia developing in patients with mediastinal lymphoblastic lymphoma.

Of three patients with mediastinal malignant lymphoma, lymphoblastic type, at the time of diagnosis one also had acute myeloblastic leukemia (AML), and the other two had blood and bone marrow findings indicative of acute lymphoblastic leukemia (ALL). The latter two patients developed the hematologic picture of AML less than eight months later. In all cases, AML was confirmed by cytochemical studies of peripheral blood and bone marrow cells. Autopsy of two of the patients revealed only AML. The myeloid nature of the proliferative cells was demonstrated with the naphthol-ASD-chloroacetate stain (NCA) on postmortem tissue sections. This study further supports the hypothesis of a common origin of neoplastic lymphoid and myeloid cells from pluripotent bone marrow stem cells.     

Detection of minimal residual disease (MRD) after bone marrow transplantation (BMT) by multi-parameter flow cytometry (MPFC)

Multi-parameter flow cytometry (MPFC) was used to detect minimal residual disease (MRD) following bone marrow transplantation (BMT) in 21 patients. Bone marrow (BM)was analyzed pre-transplant and 3-4 months post-BMT while the patients were in clinical and morphological remission. MRD was detected by identifying cells with aberrant antigen expression and/or leukemia-associated phenotype (LAP) using MPFC. Prior to BMT, 8 out of 21 patients exhibited normal antigen expression based on normal BM samples while 13 BM aspirates had abnormal MPFC. Pre-BMT MPFC was abnormal in all 10 patients who were not in complete remission (CR) (>5% blasts in BM) as well as 3 patients acute lymphoblastic leukemia (ALL) who were in CR. In BM from ALL patients, an abnormal uniform B cell population was observed however antigen expression patterns varied greatly between patients. BM from acute myeloblastic leukemia (AML) patients showed an abnormal distribution of CD34+ cells. In addition, a correlation was observed between pre-BMT cytogenetics and MPFC. Only 2 out of 8 (25%) patients with normal MPFC pre-autologous bone marrow transplantation (ABMT) relapsed (AML), while 6 out of 13 (46%) patients with abnormal pre-BMT MPFC relapsed including 2 out of 3 patients who were transplanted in clinical CR. Pre-BMT MPFC may thus be an effective tool for detection of MRD by detection of a pre-transplant MPFC abnormality.     

Role of interleukin-2 in human hematological malignancies

Clinical studies with Interleukin-2 (IL-2) in human hematologic malignancies were initiated in the late 1980s. Based on clinical studies on various solid tumors, and laboratory research on hematopoietic cells, IL-2 was shown to be effective in 150 acute myeloid leukemia (AML) patients mainly for maintenance therapy in first complete remission, or with residual blast cells in the marrow. IL-2 has also been shown to be effective in remission induction in 10 patients with chronic myeloid leukemia (CML). The role of IL-2 in lymphoma patients remains to be established. IL-2 alone or in combination with Interferon-α, may intensify remission and prolong disease-free survival when given post autologous bone marrow transplantation (BMT) to patients with lymphoma and myeloid leukemia, and to a lesser degree, to patients with acute lymphatic leukemia (ALL). IL-2 in combination with HLA-matched or mismatched peripheral blood lymphocytes was also used post autologous BMT in preliminary studies. IL-2 was administered with or without peripheral blood lymphocytes, for prevention of relapse post T-cell-depleted allogeneic BMT in CML, ALL and AML, with encouraging results. The same strategy was shown to be effective in the reinduction of remission in patients with CML, who relapsed post BMT.     

Elevated frequencies of CD4⁺ CD25⁺ CD127lo regulatory T cells is associated to poor prognosis in patients with acute myeloid leukemia

Regulatory T cells (Treg) mediate amelioration of disease and immune homeostasis by inhibiting immune activation and maintaining peripheral immune tolerance. The suppressive mechanisms and clinical significance of Treg have not been completely elucidated in patients with acute myeloid leukemia (AML). Here, we demonstrated that CD127 in combination with CD4 and CD25 can identify FoxP3(+) Treg in peripheral blood (PB) and bone marrow (BM) using multicolor flow cytometry. We showed that the CD4(+) CD25(+) CD127(lo) Treg frequencies were significantly increased and their phenotypes were different in PB from newly diagnosed AML patients compared to those from healthy volunteers (HVs). Moreover, the Treg frequencies were significantly higher in BM than those from PB in the same patients. The Treg frequencies were reduced when patients achieved complete remission (CR) and were increased when patients relapsed. The Treg frequencies at diagnosis in PB and BM of patients who had achieved CR were lower than those of patients who had persistent leukemia or died, respectively. CD4(+) CD25(+) Treg were isolated by magnetic-activated cell sorting and tested for suppressive functions in coculture with allogeneic carboxyfluorescein diacetate succinimidylester-labeled CD4(+) CD25(-) responder cells. Suppression mediated by Treg was higher in AML patients compared to HVs. No significant differences were observed in the cytokines production of Treg, including interferon-gamma (IFN-γ), interleukin (IL)-4,IL-2 and IL-10, between patients with AML and HVs. Our study suggests that Treg may play a role in the pathogenesis of AML, and sequential measurements of Treg frequency may have clinical value in the evaluation of therapeutic effects and clinical outcome.     

CXCR4 antagonists mobilize childhood acute lymphoblastic leukemia cells into the peripheral blood and inhibit engraftment.

The role of CXCL12 in the bone marrow (BM) homing and growth of B-cell progenitor acute lymphoblastic leukemia (ALL) has been established. However, the effect of modulating CXCL12/CXCR4 interactions on the retention of ALL cells within the supportive BM microenvironment and the expansion and dissemination of ALL cells in vivo has not been examined. We used mouse models of human childhood and murine leukemia and specific peptide and small molecule CXCR4 antagonists to examine the importance of CXCL12/CXCR4 in the development of leukemia in vivo. CXCR4 antagonists mobilized ALL cells into the peripheral blood (PB). Extended administration of CXCR4 antagonists to mice with leukemia resulted in a reduction in the number of leukemic cells in the PB and spleens of animals compared to control treated animals in three of the five cases tested. There was also a marked reduction in the dissemination of ALL cells to extramedullary sites including liver and kidney in all cases where this occurred. Considering the inhibitory effect of stromal layers on the activity of chemotherapeutic agents and the interactive effect of CXCL12 antagonists with chemotherapeutic agents in vitro, this raises the possibility of using these agents to potentiate the effects of current chemotherapy regimens.     

线粒体DNA在急性髓系白血病中的检测意义

目的探讨线粒体DNA(mitochondrial DNA,mtDNA)拷贝数改变在急性髓系白血病(acute myeloid leukemia,AML)中的临床意义。方法采用实时荧光定量PCR检测60例AML患者骨髓及外周血的mtDNA;以20例健康人外周血为对照,评估mtDNA(ND1单拷贝基因)拷贝数和核基因组人球蛋白(human globulin,hGB)的相对值在AML中的检测意义。结果初发或复发/难治AML患者外周血和骨髓液中mtDNA相对拷贝数高于治疗后缓解组(P<0.01);而同一患者外周血与骨髓液中mtDNA相对含量无明显差异(初发或复发/难治组,P=0.112;治疗后缓解组,P=0.667)。治疗后缓解AML患者外周血mtDNA含量与健康人差异无统计学意义(P=0.13),而初发或复发/难治AML患者外周血mtDNA含量高于健康人(P<0.01)。结论初发或复发/难治的AML患者骨髓液及外周血mtDNA含量均高于治疗后缓解AML患者,且同一患者外周血与骨髓液mtDNA含量无差异。检测AML患者外周血mtDNA相对拷贝数在一定程度上可以反映疾病状态。     

DNA ligases in human leukemia

Following partial purification on sucrose gradient and/or phosphocellulose chromatography, DNA ligase was tested in peripheral white blood and bone marrow cells of nearly 100 patients with various kinds of leukemias, mainly acute leukemias. Terminal deoxynucleotidyl transferase (TdT) was tested in parallel. DNA ligase of acute myeloblastic leukemia (AML) was extracted with the same sedimentation coefficient (5.5S) on sucrose gradient, and eluted with the same KCl molarity (0.3 M) than the one extracted from normal lymphocytes. Acute lymphoblastic leukemias (ALL) were characterized by no detectable DNA ligase activity--in most T or non T-non B-ALL, or a low activity in pre-B and B (Burkitt type) ALL, with levels similar to the one observed in chronic lymphocytic leukemia (CLL). An inverse relationship was observed between DNA-ligase and TdT in ALL, ligase being undetectable in cells positive for TdT and being present in some T or non T-non B, and in all pre-B and B-ALL negative for TdT. AML and chronic myelocytic leukemia (CML) were characterized by a markedly higher DNA-ligase activity. This activity was higher in the most differentiated subtypes--M2, M3 and M4 subtypes of FAB classification--and in CML. Moreover a high degree of correlation was observed in AML between the DNA ligase activity and the S phase fraction measured by 3 H-thymidine autoradiography or flow cytophotometry on the total cell sampling. Besides their clinical interest, these results are discussed in relation with the role of DNA-ligase in DNA replication and repair.