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：
8659350
负责人：
Shujun Liu
金额：
$ 33.79万
依托单位：
UNIVERSITY OF MINNESOTA
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 2016-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8659350
关键词：
Aberrant DNA Methylation Acute Myelocytic Leukemia Address Adverse effects Aging Animal Model Animals Apoptosis Area Attenuated Azacitidine BCL2 gene 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）：急性髓系白血病（AML）是最常见的成人白血病之一，对于大多数患者来说仍然是一种致命的疾病。地西他滨或 5-阿扎胞苷抑制异常 DNA 甲基化可恢复细胞增殖、分化和凋亡的正常模式，但临床反应仅限于少数造血系统恶性肿瘤。此外，化疗药物的脱靶摄取和低效递送会导致不良副作用。因此，迫切需要向白血病细胞靶向递送新型治疗剂。我们的长期目标是开发创新的纳米载体，并阐明控制 DNA 甲基化的调控机制，从而控制白血病的发生，这是开发可用于减轻疾病过程的治疗方案的先决条件。具体假设是，当用作联合疗法时，DNA 低甲基化剂（硼替佐米、miR29b 和 Sp1 siRNA）之间存在协同效应，并且当通过基于脂质体或脂多聚复合物的纳米载体递送时，这种协同效应可能会增强。我们的假设基于以下观察结果：1) miR29b 直接或间接破坏 AML 中的所有 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通过 miR29b 上调消除 Sp1/NF:B 依赖性 DNMT1 转录并消除 DNMT3a 和 3b 表达，进而导致 DNA 低甲基化。 3) Sp1/NF:B 复合物是控制 miR29b 和 DNMT 转录的中央调节因子。两种调节剂的抑制剂都表现出令人鼓舞的抗 DNMT 活性。 4）靶向脂质纳米载体递送化学化合物或寡脱氧核糖核苷酸（ODN）显示出更显着的靶标下调，从而在体外和体内具有更显着的抗肿瘤活性。基于这些观察，本提案的实验重点是纳米载体的设计、合成和优化以及硼替佐米与miR29b或Sp1 siRNA联合的协同效应。具体目标旨在对各种制剂的药物递送功效和特异性进行全面评估，并评估硼替佐米、miR29b和sp1 siRNA作为单一疗法或联合疗法的体外和体内药理活性。具体目标是： 1. 设计、合成和优化靶向脂质体 (Lips) 和脂多聚复合物 (LP)，用于将硼替佐米和 Sp1 siRNA 或 miR29b 递送到 AML 细胞系和 AML 患者原代细胞中。我们将通过以下方式开发合适的靶向脂质体制剂：i）使用聚碳酸酯膜挤出合成纳米载体，然后远程加载药物或新开发的微流体（MF）方法，然后与靶向配体缀合，ii）表征靶向纳米载体的尺寸和尺寸分布、药物递送功效、细胞摄取和毒性。 2.确定Lip-bortezomib与LPs-Sp1 siRNA或Lip-bortezomib与LPs-miR29b的组合在AML细胞系和患者原代细胞中的药理活性。我们将使用 i) 蛋白质印迹和定量 PCR (qPCR)、ii) LC/MS/MS 和 iii) MTS 和 PI 证明，通过靶向纳米载体递送的 DNA 低甲基化剂同时对 DNMT 调控网络进行药理学修饰，可协同诱导体外 DNA 低甲基化/AV染色。 3.在小鼠模型中对Lip-bortezomib与LPs-Sp1 siRNA或Lip-bortezomib与LPs-miR29b的组合的药理活性进行临床前体内评价。我们将通过使用蛋白质印迹、qPCR、LC/MS/MS 和临床疗效评估对患有白血病的小鼠进行药代动力学/药效学 (PK/PD) 研究来确定在体内实现协同作用的组合的有效药理学剂量。该项目将由具有转化研究和实验治疗学（Liu、Marcucci 和 Garzon）、PK/PD（Chan）、纳米工程（JLee 和 RLee）专业知识的研究人员通过跨学科方法进行。如果成功，这项研究将增进对纳米科学的理解，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，并建立抑制异常 DNMT 活性的基本概念。

项目成果

期刊论文数量（11）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Inactivation of Receptor Tyrosine Kinases Reverts Aberrant DNA Methylation in Acute Myeloid Leukemia.

DOI：
10.1158/1078-0432.ccr-17-0235
发表时间：
2017-10-15
期刊：
Clinical cancer research : an official journal of the American Association for Cancer Research
影响因子：
0
作者：
Shen N;Yan F;Pang J;Zhao N;Gangat N;Wu L;Bode AM;Al-Kali A;Litzow MR;Liu S
通讯作者：
Liu S

Liposomal bortezomib is active against chronic myeloid leukemia by disrupting the Sp1-BCR/ABL axis.