Pathogenesis And Treatment Of Aplastic Anemia

再生障碍性贫血的发病机制和治疗

• 批准号: 7321592
• 负责人: NEAL S YOUNG
• 金额: --
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7321592
• 关键词: Pathogenesis; Treatment; Aplastic; Anemia

Aplastic anemia (AA) and other types of bone marrow failure have clinical and laboratory features consistent with an autoimmune pathophysiology, with a diversity of putative inciting antigens, including viruses, chemicals, medical drugs, and tumor antigens. Whatever it?s specific etiology, a majority of patients respond with hematologic improvement after immunosuppressive therapies. One important clinical feature of AA is its association with clonal hematologic diseases, especially paroxysmal nocturnal hemoglobinuria (PNH) and myelodysplastic syndromes (MDS). In the clinic, studies have been directed towards more effective immunosuppression in aplastic anemia and the application of immunosuppressive regimens to related bone marrow failure syndromes. We have completed analysis of rabbit ATG, a more potent formulation antilymphocyte globulin recently licensed for use by the FDA in the US: about 1/3 of patients who have failed a course of horse ATG will respond hematologically to transfusion-independence on treatment with rabbit ATG; the majority of patients who relapse after horse ATG will respond to rabbit ATG. In patients with refractory aplastic anemia, unresponsive to horse ATG, we have also observed hematologic recovery using the monoclonal antibody CAMPATH, directed against CD52, a ubiquitous antigen on human lymphocytes. These clinical results ave led to the institution of a triple arm, randomized protocol for severe aplastic anemia on presentation; approximately 30 patients have been entered into this protocol, which compares horse ATG with a long course of cyclosporin; rabbit ATG with standard cyclosporin; and CAMPATH alone. Hematologic responses have been observed in all arms, including for CAMPATH , but more patients will be required for adequate comparison of the primary endpoints of hematologic response as well as relapse and clonal evolution. In studies in other diseases, we have established that the monoclonal antibody declizumab is effective not only in moderate aplastic anemia but also in pure red cell aplasia, leading to transfusion-independence in single lineage marrow failure in this format about 1/3 of patients, with no or minimal drug toxicity. In myelodysplasia, analysis of outcomes in a large number of patients who were treated with ATG with or without cyclosporin have shown a clear survival advantage, in comparison to historical data obtained in multi-center observations. Also in myelodysplasia, CAMPATH has been effective in several patients in a new protocol as a single agent, again with modest toxicity in comparison to ATG. The Hematology Branch has also participated in a large multi-center trial of Ecluzimab, a monoclonal antibody to the C5A component of complement; the results of this study will be published shortly. In the laboratory, efforts have concentrated on the mechanism of immune suppression, and the pathophysiology of late clonal events, including PNH, MDS, and acute leukemia. Studies of T-cell clonotypes continue to show a varied pattern in patients who respond, relapse, or are refractory to immunosuppressive therapy, although in general dominant clonal types recede with clinical improvement and recur, often accompanied by new clones of T-cells, on relapse. Current studies analyze the pattern of transcription of cytokines and immunomodulatory and other regulatory genes in isolated clones obtained by flow cytometric sorting based on the specific V-beta subfamily of the T-cell receptor. Other investigations have examined on the signal transduction pathways involved in T-cell polarization and the aberrant T-cell response. Results have including the finding of increased T-bet, a transcriptional regulator that is crucial for T-cell polarization to TH1 cells. New results, all consistent with an immune hypothesis for the pathogenesis of acquired aplastic anemia, include decreased PRF1 gene expression, low perforin, and low cytolytic activity in aplastic anemia, similar to that observed in familial hemophagocytic syndrome. Analogously, we have observed a high prevalence of decreased expression of SAP, a small immunomodulatory protein also important in the regulation of gamma interferon gene expression. In other studies relevant to the genetics of acquired aplastic anemia, we have described mutations in genes of the telomere repair complex, including for telomerase, TERT, and TERC in patients with apparently acquired aplastic anemia. In a large Mennonite kindred with a novel TERT mutation, the mutation has tracked with short telomeres and a spectrum of hematologic manifestations, including frank aplastic anemia, mild anemia, macrocytosis, and no hematologic abnormalities; the presence of the mutation can be inferred in six generations of this highly informative family; one patient with TERT mutation in this kindred suffered early liver failure, requiring liver transplantation. Studies of telomere repair in aplastic anemia have also led to the discovery that a likely mechanism of action of male hormones, historically utilized for the treatment of a variety of bone marrow failure conditions, is telomerase gene up-regulation. In primary lymphocytes and CD34 cells, different androgen preparations act through conversion by aromatase to estrodiol and binding to the estrogen receptor, leading to activation of TERT. Sex hormones can also up-regulate telomerase activity in cells from patients mutant in TERT. We are investigating the functional role in the Shwachman-Bodian-Diamond syndrome gene (SBDS), which we found to be mutated in heterozygosity in patients with apparently acquired aplastic anemia and in their mothers with life-long mild macrocytic anemia. The SBDS gene product does not associated physically with the telomere repair complex; heterogeneity of telomere length in DNA hybridization and other data suggests that SBDS may be related to an alternative pathway of telomere repair. In the area of clonal evolution, we have focused on two cytogenetically defined forms of myelodysplasia, trisomy 8 and monosomy 7 MDS. In trisomy 8 MDS, patients are frequently responsive to immunosuppressive therapy and their blood counts may be dependent on low level continued cyclosporin. In previous work, we demonstrated that trisomy MDS is similar to aplastic anemia in a high prevalence of oligoclonal T-cells, which appear to be specifically reactive to the cytogenetically aberrant clone. In more recent work, we have demonstrated a block in the apoptosis pathway: trisomy 8 cells, while annexin-positive, do not undergo DNA degradation and are capable of proliferation in vitro. This block has correlated with increased gene expression of c-myc, cyclin D1, and surviving. Most recently, the antigen in trisomy 8 has been shown to Wilms tumor-1 (WT-1), previously implicated in acute myelogenous leukemia and the subject of vaccine development in studies of malignant hematology. T-cells in a large proportion of patients with trisomy 8 are specific for WT1, as demonstrated as in tetramer methodology. In monosomy 7, the most frequent cytogenetic abnormality to evolve form aplastic anemia, clinical data have suggested an association with either the exogenous administration of granulocyte-colony stimulating factor (G-CSF) or persistent neutropenia and elevated endogenous G-CSF levels. We have shown that monosomy 7 does not arise de novo from normal bone marrow cells, but under conditions of high G-CSF stimulation, small clones of pre-existing monosomy 7 cells expand, as detected by fluorescent insitu hybridization (FISH), due to increased expression of a normal isoform of the G-CSF receptor which lacks the cytocellic signaling domain for differentiation. As a result, under conditions of high G-CSF in vivo, these cells likely are selected to undergo proliferation and not differentiation.

再生障碍性贫血（AA）和其他类型的骨髓衰竭具有与自身免疫病理生理一致的临床和实验室特征，具有多种推定的刺激性抗原，包括病毒、化学物质、药物和肿瘤抗原。不管它吗?S特异性病因学，大多数患者在免疫抑制治疗后血液学改善。AA的一个重要临床特征是其与克隆性血液病，特别是阵发性夜间血红蛋白尿（PNH）和骨髓增生异常综合征（MDS）的相关性。在临床上，研究的方向是在再生障碍性贫血中更有效的免疫抑制，以及将免疫抑制方案应用于相关的骨髓衰竭综合征。我们已经完成了兔ATG的分析，这是一种更有效的抗淋巴细胞球蛋白制剂，最近被FDA批准在美国使用：大约1/3的马ATG治疗失败的患者在接受兔ATG治疗后，血液学上对不依赖输血有反应；大多数马ATG后复发的患者对兔ATG有反应。在对马ATG无反应的难治性再生障碍性贫血患者中，我们也观察到使用单克隆抗体CAMPATH，针对CD52（一种普遍存在于人淋巴细胞上的抗原）的血液学恢复。这些临床结果导致了一种三臂随机方案的建立，用于治疗出现的严重再生障碍性贫血；大约有30名患者参与了该方案，该方案将马ATG与环孢素的长期疗程进行比较；兔ATG加标准环孢素；和CAMPATH。包括CAMPATH在内的所有组均观察到血液学反应，但需要更多的患者来充分比较血液学反应的主要终点以及复发和克隆进化。在其他疾病的研究中，我们已经确定单克隆抗体declizumab不仅对中度再生障碍性贫血有效，而且对纯红细胞发育不全也有效，导致约1/3的单系骨髓衰竭患者输血不依赖，没有或只有很小的药物毒性。在骨髓发育不良中，与多中心观察获得的历史数据相比，对大量接受ATG联合或不联合环孢素治疗的患者的结果分析显示出明显的生存优势。同样，在骨髓增生异常中，CAMPATH作为单一药物在一些患者中有效，与ATG相比毒性较小。血液学分支还参与了一项大型多中心试验Ecluzimab，一种针对补体C5A成分的单克隆抗体；这项研究的结果将很快发表。在实验室，努力集中在免疫抑制的机制和晚期克隆事件的病理生理学，包括PNH， MDS和急性白血病。对t细胞克隆型的研究继续显示出对免疫抑制治疗有反应、复发或难治性的患者的不同模式，尽管一般情况下，优势克隆型随着临床改善而消退，复发时通常伴有新的t细胞克隆。目前的研究基于t细胞受体特异性v - β亚家族，通过流式细胞术分选获得分离克隆，分析细胞因子和免疫调节及其他调控基因的转录模式。其他研究已经检查了信号转导途径参与t细胞极化和异常t细胞反应。结果包括发现T-bet增加，这是一种转录调节因子，对t细胞向TH1细胞极化至关重要。新的结果与获得性再生障碍性贫血发病机制的免疫假说一致，包括再生障碍性贫血中PRF1基因表达降低、穿孔素水平降低和细胞溶解活性降低，与家族性噬血细胞综合征相似。类似地，我们观察到SAP表达降低的高患病率，SAP是一种小的免疫调节蛋白，在γ干扰素基因表达的调节中也很重要。在其他与获得性再生障碍性贫血遗传学相关的研究中，我们描述了端粒修复复合体基因的突变，包括端粒酶、TERT和TERC在明显获得性再生障碍性贫血患者中的突变。在一个具有新型TERT突变的大型门诺派亲属中，该突变被追踪到端粒短和一系列血液学表现，包括明显的再生障碍性贫血、轻度贫血、巨噬细胞增多和无血液学异常；突变的存在可以在这个信息丰富的家族的六代中推断出来；该家族中有1例TERT突变患者出现早期肝功能衰竭，需要肝移植。对再生障碍性贫血中端粒修复的研究也导致发现，端粒酶基因上调可能是男性激素的作用机制，这种激素历来用于治疗各种骨髓衰竭情况。在原发性淋巴细胞和CD34细胞中，不同的雄激素制剂通过芳香化酶转化为雌激素二醇并与雌激素受体结合，导致TERT的激活。性激素也可以上调TERT突变患者细胞的端粒酶活性。我们正在研究Shwachman-Bodian-Diamond综合征基因（SBDS）的功能作用，我们发现该基因在明显获得性再生障碍性贫血患者及其终身轻度大细胞性贫血的母亲中发生杂合性突变。SBDS基因产物与端粒修复复合体没有物理关联；DNA杂交中端粒长度的异质性和其他数据表明，SBDS可能与端粒修复的另一种途径有关。在克隆进化领域，我们专注于两种细胞遗传学定义的骨髓发育不良形式，8三体和7单体MDS。在8三体MDS中，患者通常对免疫抑制治疗有反应，他们的血细胞计数可能依赖于低水平的持续环孢素。在之前的工作中，我们证明了MDS三体与再生障碍性贫血相似，在低克隆t细胞的高患病率中，它们似乎对细胞遗传学异常克隆具有特异性反应。在最近的工作中，我们已经证明了凋亡途径的阻断：8三体细胞，虽然膜联蛋白阳性，但不经历DNA降解，并且能够在体外增殖。该阻滞与c-myc、细胞周期蛋白D1基因表达增加和存活相关。最近，8号三体中的抗原已被证明与Wilms肿瘤-1 （WT-1）有关，该抗原先前与急性髓性白血病有关，并在恶性血液学研究中成为疫苗开发的主题。正如四聚体方法所证明的那样，很大一部分8三体患者的t细胞是WT1特异性的。在单体7中，最常见的细胞遗传学异常演变为再生障碍性贫血，临床数据表明其与外源性给予粒细胞集落刺激因子（G-CSF）或持续性中性粒细胞减少和内源性G-CSF水平升高有关。我们已经证明，单体7不会从正常骨髓细胞中重新产生，但在高G-CSF刺激的条件下，已有单体7细胞的小克隆扩增，荧光原位杂交（FISH）检测到，这是由于G-CSF受体的正常亚型表达增加，缺乏细胞细胞信号传导域用于分化。因此，在体内高G-CSF的条件下，这些细胞可能被选择进行增殖而不是分化。