CTCF peptides - a new therapeutic alternative to target HIV in the brain.

CTCF 肽 - 一种针对大脑中 HIV 的新治疗替代方案。

基本信息

批准号：
8320910
负责人：
Malgorzata Simm
金额：
$ 39.84万
依托单位：
ST. LUKE'S-ROOSEVELT INST FOR HLTH SCIS
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-09-30 至 2014-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8320910
关键词：
AIDS Dementia Complex Acquired Immunodeficiency Syndrome Affinity Amino Acid Sequence Antiviral Agents Antiviral Response Antiviral resistance Applications Grants Astrocytes Binding Binding Sites Biological Biological Assay Biological Response Modifier Therapy Brain CCCTC-binding factor CD4 Positive T Lymphocytes Cell Culture Techniques Cell Line Cell Surface Receptors Cells Complex DNA DNA Binding DNA Probes Data Dose Evaluation Extracellular Space Foundations Future Genetic Transcription Genome Goals HIV HIV-1 Highly Active Antiretroviral Therapy Human In Vitro Incidence Infection Interleukin-1 Kinetics Length Ligation Maintenance Mediating Messenger RNA Methods Mus N-terminal Names Neurotoxins Nitric Oxide Nuclear Nuclear Extract Nucleotides Oligonucleotides Organ Pathway interactions Patients Penetration Peptides Pharmaceutical Preparations Phase Phenotype Process Proteins Protocols documentation Reaction Recombinant Proteins Recombinants Refractory Reporter Genes Resistance Small Interfering RNA TNF gene TNFRSF5 gene Testing Therapeutic Agents Time Toxic effect Transmembrane Transport Virus Virus Replication base brain tissue cytokine deletion library dimer extracellular fetal in vitro activity in vivo macrophage mutant novel novel therapeutics prevent promoter protein complex research study resistance factors response therapeutic target tool uptake

项目摘要

Virus-infected macrophages are the major therapeutic target in the brain tissue, even though the productive virus replication in these cells is necessary but not sufficient to cause HIV Associated Dementia (HAD). HAART is still generally recommended for patients with AIDS and in general this treatment has been effective in decreasing the incidence of HAD; however a significant group of patients remains refractory to HAART even when its CNS penetration is enhanced by supplementing drugs. For this reason alone, the activation of native cellular mechanisms inducing anti-viral resistance in cells migrating to various organs including CNS should be considered a viable alternative to add to the existing antiviral approaches for HAD. We have induced such responses mediated through soluble molecule [previously named HRF (HIV-1 Resistance Factor)] able to prevent virus replication in CD4+ T cells. Recently we identified the HRF as a truncated form of a potent transcription suppressor - CTCF (tCTCF). The biological activity of tCTCF was confirmed by several experimental methods: (1) the siRNA silencing of CTCF mRNA in HRF(+) cells reversed the HIV-1 resistance phenotype and reduced the biological activity of their cell culture supernatants; (2) depletion of tCTCF from HRF(+) cell culture supernatant abolished its antiviral activity; while (3) the affinity purified tCTCF induced antiviral activity in human macrophages; and (4) the affinity purified tCTCF inhibited the transcription of HIV-1 LTR promoted expression of reporter gene. Our preliminary data showed that upon HIV-1 exposure, CTCF from nuclear extracts of HRF(+) but not HRF(-) control cells bound to DNA probe comprising NF-¿B, Sp1 and YB-1 binding sequences, possibly through its interaction with YB-1 protein. We hypothesize that since NF-¿B and YB-1 binding sites are separated by only 6 nucleotides the CTCF/YB1 complex bound to the virus promoter might obstruct the formation of NF- ¿B/DNA binding, thus halting transcription of virus genome. Taken together we discovered a novel form of CTCF protein that mediates innate responses to HIV-1 in human macrophages. The main goal of this application is to prepare foundation for future delivery of tCTCF as a biotherapeutic for treatment of HIV-1 in CNS. In this grant application we propose: (1) To study the mechanism of tCTCF mediated cellular responses to HIV-1; (2) To determine the minimum tCTCF amino-acid sequence required for the induction of antiviral responses and efficacy assessment of tCTCF treatment in primary human macrophages; and (3) To determine the minimal CTCF toxicity and brain entry in vivo.

病毒感染的巨噬细胞是脑组织中的主要治疗靶点，甚至尽管这些细胞中有效的病毒复制是必要的，但还不足以导致 HIV 相关痴呆 (HAD) 的治疗仍普遍建议采用 HAART。艾滋病患者，一般来说，这种治疗可以有效地减少 HAD 的发生率；然而，有相当一部分患者仍难以接受 HAART 即使通过补充药物增强其中枢神经系统渗透力。单独而言，激活诱导细胞抗病毒耐药性的天然细胞机制迁移到包括中枢神经系统在内的各种器官应被视为可行的替代方案添加到现有的 HAD 抗病毒方法中，我们已经诱导了这种反应。通过可溶性分子介导[以前称为 HRF（HIV-1 抵抗因子）] 能够阻止 CD4+ T 细胞中的病毒复制。最近我们将 HRF 确定为一种有效转录抑制因子的截短形式 - CTCF (tCTCF)。 tCTCF的活性通过以下几种实验方法得到证实：（1）siRNA HRF(+)细胞中CTCF mRNA的沉默逆转了HIV-1耐药表型 (2) tCTCF的耗尽 HRF(+)细胞培养上清液中的HRF(+)消除了其抗病毒活性，而(3)亲和力；纯化的 tCTCF 在人巨噬细胞中诱导抗病毒活性；(4) 亲和力；纯化的 tCTCF 抑制 HIV-1 LTR 的转录，促进报告基因的表达我们的初步数据表明，在暴露于 HIV-1 后，CTCF 会从核中释放出来。 HRF(+) 而非 HRF(-) 对照细胞的提取物与包含 NF-¿ 的 DNA 探针结合乙， Sp1 和 YB-1 结合序列，可能通过其与 YB-1 蛋白相互作用。自从 NF-¿ 以来就保留了这一点B 和 YB-1 结合位点仅相隔 6 个核苷酸与病毒启动子结合的 CTCF/YB1 复合物可能会阻碍 NF-的形成 ¿ B/DNA 结合，从而停止病毒基因组的转录。发现了一种新形式的 CTCF 蛋白，可介导对 HIV-1 的先天反应该应用的主要目标是为人类巨噬细胞奠定基础。未来将 tCTCF 作为治疗中枢神经系统 HIV-1 的生物疗法。我们建议的资助申请：（1）研究tCTCF介导的机制 (2) 确定最小tCTCF氨基酸诱导抗病毒反应和功效评估所需的序列原代人巨噬细胞中的 tCTCF 处理；以及 (3) 确定最小的 CTCF 毒性和体内脑进入。