Pre-clinical Studies of Therapy for Myelodysplastic Syndrome

骨髓增生异常综合征治疗的临床前研究

• 批准号: 10262194
• 负责人: Peter Aplan
• 金额: $ 62.21万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 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/10262194
• 关键词: Address; Allogenic; Angiogenesis Inhibitors; Apoptosis Inhibitor; Azacitidine; Bone Marrow; Bone Marrow Cells; Bone Marrow Transplantation; Cell Line; Cell Therapy; Cells; Clinical; Collaborations; Cooperative Research and Development Agreement; Cytosine; Cytotoxic Chemotherapy; DNA; DNA Methyltransferase Inhibitor; Data; Decitabine; Disease; Disease remission; Division of Cancer Treatment and Diagnosis; Donor Lymphocyte Infusion; Dose; Dysmyelopoietic Syndromes; Funding; Generations; Genome; Graft-Versus-Tumor Induction; Hematology; Hematopoiesis; Hematopoietic Stem Cell Transplantation; Histocompatibility; Histone Deacetylase Inhibitor; Hypermethylation; Immune; Industry; Institution; Licensing; MCL1 gene; Manuscripts; Mediating; Methylation; Minor; Minor Histocompatibility Loci; Modeling; Mouse Cell Line; Mus; NUP98 gene; Patients; Peripheral; Pharmacologic Substance; Phase I Clinical Trials; Phase III Clinical Trials; Pre-Clinical Model; Preparation; Proteins; Publishing; Relapse; Research Personnel; Role; Saline; Sampling; T-Lymphocyte; T-Lymphocyte Subsets; Time; Transgenes; Transgenic Mice; Transgenic Organisms; Transplantation; Xenograft Model; chemoradiation; cytopenia; cytotoxic; effective therapy; experimental study; graft vs leukemia effect; human disease; inhibitor/antagonist; meetings; mouse model; novel; novel therapeutics; peripheral blood; post-transplant; pre-clinical; preclinical study; predicting response; small molecule

Our initial studies have used the DNA-methyltransferase inhibitor 5-azacytidine; some of these results have recently been published (Genome Res. 4:580-91, 2014).However, since our initial studies used transgenic mice, effective treatment with 5-azacytidine could not replace the MDS bone marrow with completely normal (ie, wildtype or WT) bone marrow, since all of the bone marrow was transgenic. Therefore, in order to distinguish improvement in peripheral blood cytopenia due to differentiation of the MDS clone from elimination of the MDS clone, we have repeated the experiments using chimeric mice, that have both WT and NHD13 bone marrow. These repeat experiments have been performed using Decitabine (DAC), a related DNA methyltransferase inhibitor. Mice treated with DAC showed hematologic improvement and a survival advantage compared with saline-treated control mice; this experiment has now been repeated three times. Sorted BM cells from treated mice show clear differences in global cytosine methylation between the NHD13 and WT samples, and partial reversal of this hypermethylation with DAC treatment (collaboration with Dr. J.P. Issa). A manuscript describing these findings is in preparation. Drs. Difillipantonio, Doroshow, and colleagues from the Division of Cancer Treatment and Diagnosis (DCTD) have developed two novel DNMT1 inhibitors, one of which is now in phase I clinical trials. In collaboration with Dr. Difillipantonio and colleagues, we are now treating chimeric NHD13/WT mice with these compounds to assess efficacy in treatment of MDS. A manuscript describing the generation of chimeric mice with MDS was published in FY2019 (PMID: 30346380). Preliminary results show a survival benefit as well as improvement in hematopoiesis in mice receiving one of these DNMT1 inhibitors, designated T-dCyd. Portions of this data were presented at the 2018 ASH meeting, and a manuscript describing these findings is in preparation. The only curative option for patients with MDS remains allogeneic hematopoietic stem cell transplantation (HSCT). Allogeneic HSCT has two essential components, high-dose cytotoxic chemo-radiotherapy, and an immune-mediated graft versus host (GVH), or graft versus leukemia (GVL) effect. However, the relative contributions of high dose cytotoxic therapy and GVL are not well established in MDS patients. We propose to use transplantation of the NHD13 mice as a means to investigate this question. 1000 CGy induced a remission of 26-38 weeks, defined by normalization of peripheral blood counts and less than 2% circulating host cells. However, despite this period of prolonged remission, and prolonged survival compared to non-transplanted mice, all mice ultimately relapsed, indicating that this myeloablative therapy was not curative. To address the question of a GVL effect, we crossed C57bl6 mice with C3h.SW mice. C3H.SW mice are identical to C57Bl6 mice at the major histocompatibility loci, but have numerous mismatches at minor histocompatibility loci. For this reason, transplantation of C3H.SW donor cells into C57Bl6 host mice has been used to study GVH and GVL. We transplanted BM from C57Bl6/C3HSW mice into C57Bl6 NHD13 recipients. The mice developed little GVH or GVL under these conditions. Subsequent experiments using higher doses of BM or 5 x 10E06 peripheral T cells showed severe GVH. A third trial, using a higher dose of bone marrow cells (10E07)has led to survival 52 weeks post-transplant (PMID: 28953912). These results demonstrate a survival benefit for bone marrow transplant; ongoing experiments are aimed at determining whether donor lymphocyte infusion or co-transplantation of specific T cell subsets will lead to yet greater survival. In addition to the experiments outlined above, we have transferred NHD13 mice to colleagues at many academic institutions, and have licensed NHD13 mice to industry for pre-clinical studies. These colleagues have plans to treat NHD13 mice with a variety of agents, including histone deacetylase inhibitors, apoptosis inhibitors, and angiogenesis inhibitors. One of these compounds (ACE-536 or luspatercept) showed clinical activity in two phase III clinical trials for patients with MDS, and was recently approved for treatment of anemic MDS patients by the FDA. We have recently executed a CRADA with Tolero Pharmaceuticals, who have provided us with funding to study the effects of alvocidib (a Cdk9/Mcl1 inhibitor) and DNMTi on NHD13 mice and cell lines. These studies are now underway.

我们的最初研究使用了DNA-甲基转移酶抑制剂5-氮杂丁胺。这些结果中的一些最近已发表（基因组res。4：580-91，2014）。但是，由于我们的初步研究使用了转基因小鼠，因此使用5-氮杂丁胺的有效治疗无法完全正常（即，IE，野生型或wt）骨髓，因为所有骨髓都是转基因。因此，为了区分由于MDS克隆与消除MDS克隆的分化，为了区分外周血细胞质的改善，我们使用嵌合小鼠重复了实验，这些嵌合小鼠具有WT和NHD13骨髓。这些重复实验已使用Decitabine（DAC），一种相关的DNA甲基转移酶抑制剂。与盐水处理的对照小鼠相比，用DAC处理的小鼠表现出血液学改善和生存优势。该实验现已重复三次。来自处理的小鼠的分类BM细胞在NHD13和WT样品之间显示出明显的差异，以及通过DAC治疗（与J.P. ISSA博士的协作）对这种高甲基化的部分逆转。描述这些发现的手稿正在准备。博士。癌症治疗和诊断分区（DCTD）的Difillipantonio，Doroshow和同事已经开发了两个新型的DNMT1抑制剂，其中一个现在正在I期临床试验中。与Difillipantonio及其同事合作，我们现在正在用这些化合物治疗嵌合NHD13/WT小鼠，以评估MDS治疗的功效。描述用MDS的嵌合小鼠产生的手稿在2019财年发表（PMID：30346380）。初步结果表明，接受这些DNMT1抑制剂之一，指定为T-DCYD的小鼠中的造血效果和改善。这些数据的一部分是在2018年ASH会议上介绍的，描述这些发现的手稿正在准备。对于MDS患者而言，唯一的治疗方法仍然是同种异性造血干细胞移植（HSCT）。同种异体HSCT具有两个必不可少的成分，高剂量的细胞毒性化学疗法，以及免疫介导的移植物与宿主（GVH），或移植物与白血病（GVL）效应。但是，高剂量细胞毒性疗法和GVL的相对贡献在MDS患者中尚未很好地确定。我们建议将NHD13小鼠的移植作为研究这个问题的一种手段。 1000 CGY诱导了26-38周的缓解，这是由外周血计数的归一化和循环宿主细胞的标准化所定义的。然而，尽管这段时间长期缓解，并且与未移植小鼠相比，所有小鼠最终都复发了，这表明这种骨髓性治疗尚未治愈。为了解决GVL效应的问题，我们用C3H.SW小鼠越过C57BL6小鼠。 C3H.SW小鼠与主要组织相容性基因座的C57BL6小鼠相同，但在较小的组织相容性基因座处有许多不匹配。因此，将C3H.SW供体细胞移植到C57BL6宿主小鼠中已被用于研究GVH和GVL。我们将BM从C57BL6/C3HSW小鼠移植到C57BL6 NHD13接收者中。在这些条件下，小鼠几乎没有GVH或GVL。随后使用较高剂量的BM或5 x 10E06外周T细胞进行的实验显示出严重的GVH。第三次试验，使用较高剂量的骨髓细胞（10E07）导致移植后52周生存（PMID：28953912）。这些结果表明骨髓移植具有生存益处。正在进行的实验旨在确定供体淋巴细胞输注或特定T细胞亚群的共移植是否会导致更大的生存率。除了上面概述的实验外，我们还将NHD13小鼠转移到许多学术机构的同事，并已将NHD13小鼠许可到行业进行临床前研究。这些同事计划用各种药物治疗NHD13小鼠，包括组蛋白脱乙酰基酶抑制剂，凋亡抑制剂和血管生成抑制剂。这些化合物之一（ACE-536或Luspatercept）在两期MDS患者的III期临床试验中显示出临床活性，最近批准了FDA治疗贫血MDS患者。我们最近与Tolero Pharmaceuticals执行了CRADA，他们为我们提供了研究Alvocidib（CDK9/MCL1抑制剂）和DNMTI对NHD13小鼠和细胞系的影响。这些研究正在进行中。