Long-term follow-up of allogeneic bone marrow transplantation after reduced-intensity conditioning in patients with chronic myelogenous leukemia in the chronic phase

Although allogeneic transplantation is a curative therapy for chronic myelogenous leukemia (CML), treatment-related mortality is still a major cause of death after transplantation, especially in older patients. We investigated the safety and efficacy of reduced-intensity conditioning consisting of low-dose (600 cGy) total body irradiation and cytosine arabinoside (1 g/m2) together with a continuous infusion of granulocyte colony-stimulating factor and cyclophosphamide (120 mg/kg) in patients with CML in the chronic phase. Fractionated splenic irradiation (5 Gy) was also administered as part of the conditioning treatment. Eight patients older than 40 years underwent allogeneic bone marrow transplantation from an HLA-matched sibling following this conditioning. Regimen-related toxicities (equal to or greater than grade III) were not observed. Rapid restoration of 100% donor chimerism was confirmed by fluorescence in situ hybridization methods in 5 sex-mismatched transplant recipients. One patient died from severe acute graft-versus-host disease and another from Pneumocystis carinii pneumonia early in the course of transplantation. A sustained engraftment was achieved in 5 long-term survivors; in 1 case, the graft was rejected but the Philadelphia chromosome and BCR/ABL-negative autologous hemopoiesis were restored. After a minimum follow-up period of 60 months, 6 patients, including the patient with restored autologous hemopoiesis, were still alive and in remission with 100% donor chimerism. Six years after the transplantation, 1 patient experienced a cytogenetic relapse, which was successfully treated with donor lymphocyte infusions. In summary, this reduced-intensity conditioning resulted in a cure with markedly reduced regimen-related toxicities in this relatively older cohort of patients with CML.     

Long-term leukemia-free survival after allogeneic marrow transplantation in patients with acute myelogenous leukemia

Between February 1982 and April 1995, 62 patients (37 male, 25 female) with acute myelogenous leukemia (AML) with a median age of 32 years (19–51 years) received allogeneic marrow grafts from an HLA-identical sibling (n=60) or an HLA-mismatched family member (n=2). At the time of transplant, 35 patients were in first complete remission (CR), five in second CR, eight were primary refractory, eight were in untreated relapse and six in refractory relapse. The FAB subtypes were as follows: M1 (n=17), M2 (n=13), M3 (n=6), M4 (n=19), M5 (n=6), M6 (n=1). For conditioning most patients were given total body irradiation combined with cyclophosphamide (CY,n=50) or CY and busulfan (n=9). For graft-versus-host disease prophylaxis patients received cyclosporin A (CSA) and methotrexate (MTX) (n=32), MTX alone (n=12), CSA and methylprednisone (n=5), or CSA alone (n=13). As of April 1995, probability of leukemia-free survival projected at 10 years after BMT was 60% for patients transplanted in first CR compared with 10% for patients transplanted beyond first CR. Transplant-related mortality was 11% after BMT in first CR and 39% after BMT beyond first CR. Probability of relapse projected at 10 years after BMT is 32% for patients who received transplants in first CR and 81% for patients who received transplants beyond first CR. Thus, high-dose chemo/radiotherapy followed by allogeneic marrow infusion has a high curative potential for patients with AML who receive transplants in first CR and offers the chance of long-term disease-free survival for some patients with advanced disease.     

Clonal expansion after bone marrow transplantation in a patient with chronic myelogenous leukemia

Summary A patient with chronic myelogenous leukemia (CML) having the standard [t(9; 22), Ph] translocation is presented where the Philadelphia (Ph) chromosome disappeared following bone marrow transplantation (BMT). The Ph chromosome reappeared in host cells after one year of stable hematologic remission. Three additional cell lines, all possessing the Ph chromosome with other abnormalities were consistently present in her marrow cells. Two years after BMT, ninety percent of her dividing bone marrow cells had become leukemic. The patient's clinical status remains unchanged, despite complex cytogenetic findings. The high incidence of multiple aberrant leukemic clones present in this case remains intriguing. Possible mechanisms for this unique transformation after BMT are discussed.     

Mechanisms of Transformation by the BCR/ABL Oncogene

The Philadelphia chromosome generates a chimeric oncogene in which the BCR and c-ABL genes are fused. The product of this oncogene, BCR/ABL, has elevated ABL tyrosine kinase activity, relocates to the cytoskeleton, and phosphorylates multiple cellular substrates. BCR/ABL transforms hematopoietic cells and exerts a wide variety of biological effects, including reduction in growth factor dependence, enhanced viability, and altered adhesion of chronic myelocytic leukemia (CML) cells. Elevated tyrosine kinase activity of BCR/ABL is critical for activating downstream signal transduction and for all aspects of transformation. This review will describe mechanisms of transformation by the BCR/ABL oncogene and opportunities for clinical intervention with specific signal transduction inhibitors such as STI-571 in CML.     

Improved survival of patients with chronic myelogenous leukemia undergoing allogeneic bone marrow transplantation

A total of 28 patients with chronic myelogenous leukemia (CML) in chronic phase (CP) received bone marrow allografts from HLA-matched siblings at the University of Florida between August 1984-July 1992. The present study compares the disease-free survival (DFS) for those patients who were transplanted before or after August 1988 using the same conditioning regimen. The analysis shows significant difference in 3-year DFS for those patients transplanted post- vs. pre-August 1988 (69.6% vs. 20%, respectively; P = 0.006). A decrease in pneumonitis due to different etiologies from pre-August 1988 (6/13, 46%) to post-August 1988 (1/15, 7%) was statistically significant (P = 0.029). A decrease, although statistically insignificant, in the overall incidence and severity of acute and chronic graft vs. host disease (GVHD) after August 1988 was also noticed. This study indicates significantly improved outcome for patients with CML in CP who have been treated in the University of Florida after August 1988. Better supportive care and prophylaxis for GVHD most likely contributed to such improvement.     

慢性粒细胞白血病BCR/ABL融合基因及复杂变异的染色体核型分析

目的探讨慢性粒细胞白血病(CML)患者BCR/ABL融合基因及其复杂变异的染色体核型变化及临床意义。方法在常规细胞遗传学(CC)方法检测基础上,运用分子生物学方法——荧光原位杂交(FISH)技术,采用多种位点特异性DNA探针(染色体全染、特殊位点、双色易位融合探针),对2002年9月至2007年2月中国医科大学附属第一临床学院血液科56例门诊及住院CML患者(慢性期51例,加速及急变期5例)进行染色体核型分析。结果56例CML患者有5例为细胞培养失败,均为慢性期患者,该5例细胞培养失败病例经FISH技术证实均出现Ph 染色体,余46例慢性期患者中有42例出现Ph 染色体,其中2例为双Ph 染色体;3例合并有复杂的染色体变化,包括染色体数目及结构的异常,分别为-2,-6,-19,-16,-20,-Y, 8以及t(2;4)(p16;p15)。CML慢性期共有47例患者出现Ph 染色体,总阳性率为92.17%。5例加速及急变期患者均出现Ph 染色体,其中3例患者合并有复杂的染色体核型变化:-Y,del(10),I(9)。结论染色体核型分析对CML的疾病分期、进展、治疗及预后有重要的临床意义。FISH技术在CML染色体核型分析上可作为重要技术补充手段弥补CC检查的不足。     

Allogeneic stem cell transplantation for chronic lymphocytic leukemia: lessons to be learned from minimal residual disease studies

Allogeneic stem cell transplantation (alloSCT) is a potentially curative treatment strategy for poor-risk chronic lymphocytic leukemia (CLL). The crucial anti-leukemic principle of alloSCT in CLL appears to be the graft-versus-leukemia effect (GVL). Evidence for GVL in CLL is particularly provided by studies analysing the kinetics of minimal residual disease (MRD). The purpose of this review is to summarize the methodologies of MRD assessment, its proven benefits and its further perspectives for optimizing the outcome of transplantation. Proven value of quantitative MRD monitoring by RQ-PCR or MRD-flow consists in using it as an indicator of long-term disease control and potential cure. As MRD kinetics correlates with GVL activity, its suitability for guiding GVL-inducing immunomodulation is currently under investigation. In conclusion, quantitative MRD monitoring seems to be mandatory to assure safe and effective immunotherapy in the context of alloSCT for CLL, which should, however, be best performed within clinical studies.     

实时定量PCR监测造血干细胞移植后BCR/ABL融合基因的表达

目的监测慢性粒细胞白血病（CML）患者异基因造血干细胞移植（allo-HSCT）后BCR／ABL融合基因表达水平的动态变化,从而判断移植效果,指导临床治疗。方法应用Taqman实时定量RT-PCR方法,以B-actin作为内参照,检测10例初发CML患者及25例接受allo-HSCT治疗的CML患者在移植前、后不同时段外周血中BCR／ABL mRNA表达水平。结果15例接受移植的患者移植前、移植后1、2个月的NBCR／ABL（％）中位数分别为：6．57（0．14～38．83）、0．10（0～1．71）、0（0～0．52）,3个月后全部为0。移植后早期检测BCR／ABL转录本较移植前逐渐下降,移植后1、2个月NBCR／ABL（％）均较移植前下降（x^2均为13．07,P〈O．01）;移植后2个月较1个月下降（x^2=8．10,P〈0．01）。其余10例患者从移植后3～43个月不同时段起连续检测NBCR/ABL（％）也全部为0。结论实时定量RT-PCR方法检测CML患者的BCR／ABL融合基因的表达,操作迅速,灵敏度、特异性好,用于移植后微小残留病的检测对评估预后并指导临床治疗具有重要意义。     

Successful allogeneic bone marrow transplantation after reversion to chronic phase of blast crisis in chronic myeloid leukemia

A female with chronic myeloid leukemia (XX Ph 1 +) in blast crisis (localized to pleura and lymph nodes) was treated by polychemotherapy. After reversion to the chronic phase, an allogeneic bone marrow transplantation (BMT) was performed. Sixteen months after BMT, no sign of the disease was present (XY Ph 1 -).     

Clinical significance of fragmented red cells after allogeneic bone marrow transplantation

To clarify the clinical significance of the presence of fragmented red cells (FRC) after allogeneic bone marrow transplantation (BMT), we measured the incidence and degree of FRC and their relationships to clinical features. The percentages of FRC (%FRC) were measured in 50 patients on weeks -2, 0, 2, 4, 6, 8, 10, and 12. The %FRC in pre-BMT patients (mean, 0.52%; range, 0.04%-1.56%) was higher than in healthy control subjects (mean, 0.08%; range, 0.02%-0.27%). The highest %FRC (> or = 1.3%) were seen in 2 pre-BMT and 17 post-BMT patients. Eight patients who developed thrombotic microangiopathy (TMA) showed %FRC values that were significantly higher than those in patients without TMA. However, the timing of elevated %FRC was delayed until several days after the onset of intravascular hemolysis and/or a drop in platelet count. Of the patients who did not experience TMA, 5 patients with infection and 4 patients with acute graft-versus-host disease (GVHD) also showed significant elevation of %FRC during the clinical course. Furthermore, multivariate analysis results demonstrated that TMA and infection had a statistically significant effect on the high value of %FRC. These findings indicate that the appearance of FRC is a common phenomenon in patients undergoing BMT and is not a predictive factor for the early diagnosis of TMA, although FRC is one of the main laboratory findings in TMA. Furthermore, an increased %FRC is seen in other post-BMT clinical settings, such as infection and acute GVHD.     

Successful unrelated donor bone marrow transplantation for shwachman-diamond syndrome with leukemia

Shwachman-Diamond syndrome (SDS) is a rare congenital disorder featuring exocrine pancreatic insufficiency, growth retardation, and bone marrow dysfunction. Reports suggest that nearly 25% of all cases are complicated with leukemia. Although stem cell transplantation is the sole option for these patients, successful results are rarely obtained. Poor outcomes are often related to graft failure and cardiac and other organ toxicities. We describe in this report successful unrelated donor bone marrow transplantation for a patient with SDS who progressed to acute myelogenous leukemia. The patient received attenuated intensified chemotherapy because of his intolerance to ordinary chemotherapy and went into remission. Sustained unrelated donor bone marrow engraftment was accomplished after treatment with a reduced amount of cyclophosphamide and antithymocyte globulin with 12 Gy of total body irradiation as a conditioning regimen. To the best of our knowledge, this report is the first to describe unrelated donor bone marrow transplantation with complete engraftment for an SDS patient with myelogenous leukemia.     

格列卫治疗单倍体骨髓移植后复发的难治性急性淋巴细胞白血病缓解 1例报告

目的：探讨格列卫治疗Ph^1阳性儿童急性淋巴细胞性白血病单倍体骨髓移植后复发的疗效。方法：用格列卫，300mg，1次／d，顿服治疗1例单倍体骨髓移植后复发难治性急性淋巴细胞白血病。治疗前后观察血常规、骨髓象、BCR／ABL融合基因。结果：治疗4周后达完全缓解，BCR／ABL基因转阴。结论：格列卫治疗Ph^1阳性儿童急性淋巴细胞性白血病单倍体骨髓移植后复发有效。     

Donor cell leukemia in a patient developing 11 months after an allogeneic bone marrow transplantation for chronic myeloid leukemia

A 38-year-old female with chronic myeloid leukemia underwent an allogeneic bone marrow transplantation from her full-matched brother. Eleven months later, she readmitted with an acute leukemia that was shown to be of donor origin. The patient never achieved a remission even after chemotherapies with cytarabine and mitoxantrone, donor lymphocyte infusion, and second allogeneic peripheral blood stem cell transplantation. Donor cell leukemia (DCL) is sometimes misdiagnosed as relapse by clinicians and the real incidence may be higher than expected. Cytogenetic and molecular techniques may be helpful to clarify the issue of the leukemia. The current case is another case of DCL reported in the literature after an allogeneic transplant for a kind of leukemia.     

Elevated platelet count features the variant type of BCR/ABL junction in chronic myelogenous leukaemia

A variant form of BCR/ABL junction was identified in a patient with chronic myelogenous leukaemia (CML). The BCR/ABL fusion mRNA of this patient showed in-frame junction between BCR exon c3 and ABL exon 2. Although the diagnosis of CML was made, the patient showed clinical features of essential thrombocythaemia (ET) rather than that of typical CML. Treatment with interferon-α showed no cytogenetic response. The c3-a2 type of BCR/ABL junction seems to be associated with elevated platelet count and thus could form a novel clinical entity different from typical CML.     

Human leucocyte antigen alloimmunization after bone marrow transplantation: an association with chronic myelogenous leukaemia

Platelet refractoriness due to human leucocyte antigen (HLA) alloimmunization is a significant risk to allogeneic bone marrow transplant recipients. To identify factors contributing to this risk, we reviewed the records of 317 consecutive, paediatric, allogeneic bone marrow transplant recipients at a single institution. The 6-year estimated cumulative incidence of platelet refractoriness due to HLA alloimmunization was 2.6% +/- 0.9%. The incidence among patients with chronic myelogenous leukaemia (CML) 12.5% +/- 5.3% was significantly greater than that of other patients (1.1% +/- 0.6%, P < 0.001). Graft rejection (P = 0.003) and the number of platelet transfusions during the first 90 d after bone marrow transplantation (BMT) (P = 0.0025) were also significantly associated with alloimmunization. The association with CML and with graft rejection was not seen among patients alloimmunized before transplantation. Eight patients developed alloimmunization, of whom three had mismatched grafts and four had unrelated grafts. Alloantibody specificities, identified in seven patients, were unrelated to host or graft major histocompatibility complex (MHC). Host recognition of alloantigens in transfused blood products, not graft-host recognition, therefore seems predominantly responsible for the alloimmunization. These results show that paediatric CML patients have a significantly increased risk of platelet refractoriness due to HLA alloimmunization after BMT. Identifying the mechanism for the increased alloimmunization risk may assist in the development of therapies to prevent platelet refractoriness.     

Spontaneous complete remission of chronic myeloid leukaemia following haematological relapse after allogeneic bone marrow transplantation

A 31-year-old woman with Philadelphia (Ph) chromosome-positive chronic myeloid leukaemia (CML) underwent allogenic bone marrow transplantation during accelerated phase. Non-T-cell-depleted marrow from a male sibling mismatched at one Class 2 histocompatibility locus was infused after conditioning with total body irradiation and intravenous cyclophosphamide. Cyclosporin and methotrexate were given for prevention of graft-versus-host disease (GVHD). Prompt engraftment occurred with donor karyotype cells, followed by transient moderate acute GVHD. However, by day 60 after BMT, haematological relapse occurred with increasing splenomegaly, leucocytosis, increasing marrow fibrosis, and cytogenetic mosaicism, consisting of 47% donor metaphases with 53% Ph-positive host metaphases, some containing additional structural changes. Thirty days later further cytogenetic progression was evident. A slowly progressive fungal pneumonia concurrently present was treated with intravenous amphotericin and gradual reduction of cyclosporin. Subsequently, without further cytotoxic chemotherapy, pancytopenia and bone marrow hypoplasia developed, and on day 144 only donor karyotype marrow cells were seen. Chromosomes have remained of donor type on subsequent occasions, and the patient has a normal performance status 25 months after BMT. The patient's course illustrates that factors operating after allogeneic BMT contribute to long-term control of CML. The factors potentially responsible for this spontaneous remission, after early relapse, are discussed.     

Galectin-3 (Gal-3) induced by leukemia microenvironment promotes drug resistance and bone marrow lodgment in chronic myelogenous leukemia

Bone marrow (BM) microenvironment (BMME) constitutes the sanctuary for leukemic cells. In this study, we investigated the molecular mechanisms for BMME-mediated drug resistance and BM lodgment in chronic myelogenous leukemia (CML). Gene-expression profile as well as signal pathway and protein analyses revealed that galectin-3 (Gal-3), a member of the β-gal-binding galectin family of proteins, was specifically induced by coculture with HS-5 cells, a BM stroma cell-derived cell line, in all five CML cell lines examined. It was also found that primary CML cells expressed high levels of Gal-3 in BM. Enforced expression of Gal-3 activated Akt and Erk, induced accumulation of Mcl-1, and promoted in vitro cell proliferation, multidrug resistance to tyrosine kinase inhibitors for Bcr-Abl and genotoxic agents as a result of impaired apoptosis induction, and chemotactic cell migration to HS-5-derived soluble factors in CML cell lines independently of Bcr-Abl tyrosine kinase. The conditioned medium from Gal-3-overexpressing CML cells promoted in vitro cell proliferation of CML cells and HS-5 cells more than did the conditioned medium from parental cells. Moreover, the in vivo study in a mice transplantation model showed that Gal-3 overexpression promoted the long-term BM lodgment of CML cells. These results demonstrate that leukemia microenvironment-specific Gal-3 expression supports molecular signaling pathways for disease maintenance in BM and resistance to therapy in CML. They also suggest that Gal-3 may be a candidate therapeutic target to help overcome BMME-mediated therapeutic resistance.     

IL2- and IL4-dependent proliferation of T-cell clones derived early after allogeneic bone marrow transplantation: studies of patients with chronic myelogenous leukaemia.

In an attempt to explore T-cell functions shortly after allogeneic bone marrow transplantation more fully, IL2- and IL4-dependent proliferation was assessed on CD4+ TCR alpha beta+ T-cell clones derived 4-6 weeks after transplantation. Both allogeneic pooled peripheral blood mononuclear cells and Epstein-Barr virus-transformed B-cell lines (BCL) could function as accessory cells (AC) for PHA activation of T-cell clones. Although minimal clonal proliferation was seen when the T-cell activation signal was BCL+PHA+IL4, a majority of the clones could undergo IL4-dependent proliferation after previous activation with AC+PHA+IL2. For certain clones, IL4 also showed an additive effect with IL2. Thus, IL4 was a growth factor for a majority of the investigated posttransplant T-cell clones, and in vivo modulation of IL4-dependent T-cell functions may thus become a future therapeutic possibility to enhance graft-versus-leukaemia effects in bone marrow transplant recipients.