Targeting aberrant epigenetics by nanomedicine

通过纳米医学靶向异常表观遗传学

基本信息

批准号：
8094456
负责人：
Shujun Liu
金额：
$ 35.28万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-01 至 2011-09-15
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8094456
关键词：
Aberrant DNA Methylation Acute Myelocytic Leukemia Address Adverse effects Aging Animal Model Animals Apoptosis Area Attenuated Azacitidine Biological Assay Biological Markers Blast Cell Blood Bone Marrow Bortezomib Cell Line Cell Proliferation Cell surface Cells Clinic Clinical Clinical Trials Combined Modality Therapy Complex Confocal Microscopy DNA DNA Methylation DNA Methyltransferase DNA Modification Methylases DNMT3a Data Decitabine Deoxyribonucleotides Disease Disease Progression Dose Down-Regulation Doxorubicin Drug Combinations Drug Delivery Systems Drug Formulations Drug Kinetics Dysmyelopoietic Syndromes Enzyme-Linked Immunosorbent Assay Epigenetic Process Evaluation FDA approved Feedback Flow Cytometry Funding Future Gene Silencing General Population Generations Genetic Transcription Goals Growth Health Hematopoietic Neoplasms Histone Deacetylation Human Hypermethylation Impairment In Vitro Incidence Interleukin-15 Intervention Investigation Knock-out Leukemic Cell Ligands Link Lip structure Lipids Liposomes Malignant - descriptor Malignant Neoplasms Measures Mediating Membrane Messenger RNA Methods Methylation Methyltransferase Gene MicroRNAs Microfluidics Minority Modality Modeling Modification Molecular Biology Multiple Myeloma Mus Myelogenous Nanotechnology New Agents Nucleotides Oligonucleotides Outcome Pathway interactions Patients Pattern Pharmaceutical Preparations Pharmacodynamics Plasma Play Plicamycin Positioning Attribute Process Property Propidium Diiodide Proteasome Inhibitor Protein Isoforms Proteins Protocols documentation RNA Reporting Research Research Personnel Role Small Interfering RNA Solid Neoplasm Specificity Staining method Stains Structure System Therapeutic Therapeutic Agents Therapeutic Uses Toxic effect Trans-Activators Transcript Transferrin Receptor Transgenic Mice Translating Translational Research Translations Treatment Efficacy Tumor Suppressor Genes Tumor Tissue United States Up-Regulation Western Blotting Work adult leukemia annexin A5 antileukemic agent base cancer type chemotherapeutic agent clinical efficacy design diphenyl dosage experience in vitro activity in vivo inhibitor/antagonist innovation interdisciplinary approach leukemia leukemogenesis light scattering liquid chromatography mass spectrometry multicatalytic endopeptidase complex nanocarrier nanoengineering nanomedicine nanoscience neoplastic cell new therapeutic target novel novel strategies novel therapeutics overexpression particle polycarbonate pre-clinical programs promoter receptor research study response synergism targeted delivery therapeutic development tumor uptake

项目摘要

DESCRIPTION (provided by applicant): Acute myeloid leukemia (AML) represents one of the most common adult leukemia and remains as a deadly disease for most patients. Inhibition of aberrant DNA methylation by decitabine or 5-azacitidine restores normal patterns of cell proliferation, differentiation and apoptosis, however, the clinical response is restricted to a minority of hematopoietic malignancies. Further, off-target uptake and low efficient delivery of chemotherapeutic agents leads to undesirable adverse effects. Hence, there is an immediate need for targeted delivery of novel therapeutic agents to leukemic cells. Our long-term goals are to develop innovative nanocarriers and to elucidate the regulatory mechanisms controlling DNA methylation thereby leukemogenesis as a prerequisite to the development of therapeutic protocols that can be used to attenuate the disease process. The specific hypothesis is that there exists a synergistic effect among DNA hypomethylating agents (bortezomib, miR29b and Sp1 siRNA) when used as combination therapy and this synergism may be enhanced when delivered by liposome- or lipopolyplexe-based nanocarriers. We base that hypothesis on the observations that 1) miR29b disrupts all DNA methyltransferases (DNMTs) directly or indirectly in AML thereby leading to DNA hypomethylation. 2) proteasome inhibitor bortezomib depletes Sp1/NF:B-dependent DNMT1 transcription and abolishes DNMT3a and 3b expression via miR29b upregulation, in turn causing DNA hypomethylation. 3) Sp1/NF:B complex is a central regulator governing both miR29b and DNMT transcription. Inhibitors for both regulators display encouraging anti-DNMT activity. 4) targeted lipid-based nanocarrier delivery of chemo-compounds or oligo deoxyribonucleotides (ODNs) display more significant target downregulation thereby more pronounced anti-tumor activity in vitro and in vivo. Based on these observations, the experimental focus of this proposal is on the nanocarrier design, synthesis and optimization as well as the synergistic effect of bortezomib combined with miR29b or Sp1 siRNA. The specific aims are designed to provide a comprehensive assessment of drug delivery efficacy and specificity in a variety of formulations and the evaluation of in vitro and in vivo pharmacological activity of bortezomib, miR29b and sp1 siRNA as monotherapy or combination therapy. The specific aims are to: 1. Design, synthesize and optimize targeted liposomes (Lips) and lipopolyplexes (LPs) for delivering bortezomib and Sp1 siRNA or miR29b into AML cell lines and AML patient primary cells. We will develop suitable targeted liposomal formulations by i) synthesis of nanocarriers using polycarbonate membrane extrusion followed by remote-loading of the drug or newly developed microfluidic (MF) methods and then conjugated to targeting ligands, ii) characterization of targeted nanocarriers for the size and size distribution, drug delivery efficacy, cellular uptake and toxicity. 2. Determine the pharmacological activity of the combination of Lip-bortezomib with LPs-Sp1 siRNA or Lip-bortezomib with LPs-miR29b in AML cell lines and patient primary cells. We will demonstrate that simultaneously pharmacological modification of DNMT regulatory network by targeted-nanocarrier delivered DNA hypomethylating agents synergistically induces DNA hypomethylation in vitro using i) Western blot and quantitative PCR (qPCR), ii) LC/MS/MS and iii) MTS and PI/AV staining. 3. Perform preclinical in vivo evaluation of the pharmacological activity of the combination of Lip- bortezomib with LPs-Sp1 siRNA or Lip-bortezomib with LPs-miR29b in murine models. We will define the effective pharmacological dose of the combinations achieving synergisms in vivo by pharmacokinetic/pharmacodynamic (PK/PD) studies in leukemia-bearing mice using Western blot, qPCR, LC/MS/MS and the assessment of clinical efficacy. This project will be carried out through an interdisciplinary approach by investigators with expertise in translational research and experimental therapeutics (Liu, Marcucci and Garzon), PK/PD (Chan), nanoengineering (JLee and RLee). If successful, this investigation will advance the understanding of nanosciences, enhance the understanding of the roles of proteasome system, miR and DNA methylation in leukemogenesis and establish a fundamental concept for the inhibition of aberrant DNMT activities.

描述（由申请人提供）：急性髓样白血病（AML）代表最常见的成年白血病之一，对于大多数患者而言，仍然是致命疾病。抑制法替滨或5-氮杂丁氨酸对异常DNA甲基化的抑制恢复了细胞增殖，分化和凋亡的正常模式，但是，临床反应仅限于少数造血恶性肿瘤。此外，化学治疗剂的脱靶吸收和低效递送导致不良的不良反应。因此，即时需要将新型治疗剂靶向靶向白血病细胞。我们的长期目标是开发创新的纳米载体，并阐明控制DNA甲基化的调节机制，从而将白血病发生为开发治疗方案的先决条件，可用于减轻疾病过程。具体的假设是，当用作联合疗法时，DNA低甲基化剂（Bortezomib，MiR29b和Sp1 siRNA）之间存在协同作用，而当基于脂质体或脂肪型的基于脂质体或脂肪型的纳米携带者时，可能会增强这种协同作用。我们基于观察结果，即1）MiR29b直接或间接地破坏所有DNA甲基转移酶（DNMT），从而导致DNA低甲基化。 2）蛋白酶体抑制剂硼替佐米的耗尽SP1/NF：B依赖性的DNMT1转录并通过miR29b上调废除DNMT3A和3B表达，进而导致DNA甲基化。 3）SP1/NF：B复合物是管理MiR29b和DNMT转录的中央调节器。两种调节剂的抑制剂都表现出令人鼓舞的抗DNMT活性。 4）基于脂质的基于脂质的纳米载体递送化学混合物或寡核糖核苷酸（ODNS）的纳米载体递送显示出更重要的靶标下调，从而在体外和体内更明显。基于这些观察结果，该建议的实验重点是纳米载体设计，合成和优化以及硼替佐米与miR29b或sp1 siRNA的协同作用。该特定目的旨在全面评估各种配方中药物输送功效和特异性，并评估硼替佐米，miR29b和SP1 siRNA作为单一治疗或联合疗法的体外和体内药理活性。具体目的是：1。设计，合成和优化靶向的脂质体（LIPS）和脂质型（LPS），用于将硼替佐米和SP1 siRNA或MIR29B输送到AML细胞系以及AML患者原始细胞中。我们将通过i）使用聚碳酸酯膜挤出来开发合适的靶向脂质体配方，然后通过远程负载进行药物或新开发的微流体（MF）方法，然后将靶向配体的靶向配体（II）结合到靶向Nanocarriers的尺寸和尺寸分布的质量分布，促进性的质量，促进性效率，促进，并赋予药物分布，促进效率，并赋予药物分布，并赋予药物，并具有较大的效率。 2。确定脂质 - 宝唑与LPS-SP1 siRNA或LIP-BORTEZOMIB与LPS-MIR29B在AML细胞系和患者原代细胞中的药理活性。我们将证明，通过靶向纳米载体对DNMT调节网络的同时进行药理修饰，使用i）使用i）Western印迹和定量PCR（QPCR）在体外协同诱导DNA降压剂，从而在体外诱导DNA低甲基化。 3。对腹膜硼替佐米与LPS-SP1 siRNA或Lip-Bortezomib的药理活性进行临床前评估，并在鼠模型中使用LPS-MIR29B。我们将使用Western Blot，QPCR，LC/MS/MS和临床疗效评估来定义通过药代动力学/药物动力学（PK/PD）研究在体内实现协同作用的有效药理剂量。该项目将通过具有转化研究和实验疗法专业知识的研究人员（LIU，Marcucci和Garzon），PK/PD（Chan），纳米工程（JLEE和RLEE）进行。如果成功的话，这项研究将提高人们对纳米科学的理解，增强对蛋白酶体系统，miR和DNA甲基化在白血病发生中的作用的理解，并为抑制异常DNMT活性建立一个基本概念。