Determinates of anti-HIV nucleic acid aptamer potency and resistance

抗HIV核酸适体效力和耐药性的测定

• 批准号: 7801719
• 负责人: Donald H Burke
• 金额: $ 3.93万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7801719
• 关键词: AIDS/HIV problem; Acquired Immunodeficiency Syndrome; Amino Acid Sequence; Animal Welfare; Anti-HIV Agents; Bibliography; Cells; Chemicals; Coenzyme A; Country; Data; Elements; Ensure; Environment; Environmental Impact; Equipment; Evolution; Face; Freezing; Future; Gene Therapy Agent; Genetic; Goals; HIV-1; IACUC; In Vitro; Individual; Intention; International; Libraries; Maps; Modification; Molecular; Monitor; Nucleic Acids; Nucleotides; Pharmaceutical Preparations; Principal Investigator; Published Comment; RNA; RNA-Directed DNA Polymerase; Research; Research Ethics Committees; Resistance; Resources; Retrovir; Structure; Surface; Toxic effect; Variant; Vertebrates; Viral; Virus; Work; abstracting; aptamer; base; crosslink; design; drug resistant virus; expiration; high throughput screening; human subject; improved; interest; novel therapeutics; pressure; programs; small molecule

New therapeutic strategies are needed to circumvent the rapid selection of drug resistant virus and the toxicity of current anti-HIV drugs. RNA aptamers targeted to the reverse transcriptase (RT) inhibit viral replication when expressed inside cells, and they offer great potential in future therapies against HIV/AIDS. There is little known about how the virus might evolve in the face of continual selection pressure, or about how the aptamers can be improved to circumvent potential resistance. The long-term goal of this project is to develop RNA aptamers as gene therapy agents that provide long-term antiviral protection against a rapidly-evolving virus. Our emphasis is on defining the influence of RT amino acid sequence variations on inhibition by aptamers, and on identifying new aptamer sequences and structures with improved inhibition, both in vitro and in cells. Aim 1 will determine the biophysical and structural basis for aptamer recognition and for differential inhibition by aptamers across phylogenetically diverse lentiviral RT's. Aim 2 will identify new aptamers with broad recognition and improved potency, and delineate the secondary structures associated with cross-clade enzymatic inhibition. Aim 3 examines inhibition of pseudotyped and replication-competent HIV-1 by intracellularly expressed aptamers, moving from optimization of the design elements that control expression, localization and inhibition to high throughput screens of aptamer and RT libraries. Aim 4 evaluates the de novo evolution of aptamer resistance with emphasis on establishing genetic threshold, resistance loci, and the molecular basis of resistance.

需要新的治疗策略来规避耐药病毒的快速选择和当前抗艾滋病毒药物的毒性。针对逆转录酶（RT）的RNA适体在细胞内表达时会抑制病毒复制，并且它们在未来针对HIV/AIDS的治疗中具有巨大潜力。人们对病毒在面对持续的选择压力时如何进化，或者如何改进适配体以规避潜在的耐药性知之甚少。该项目的长期目标是开发RNA适体作为基因治疗剂，为快速进化的病毒提供长期抗病毒保护。我们的重点是确定 RT 氨基酸序列变异对适体抑制的影响，并鉴定在体外和细胞内具有改善抑制作用的新适体序列和结构。目标 1 将确定适体识别和适体在系统发育上不同的慢病毒 RT 中的差异抑制的生物物理和结构基础。目标 2 将鉴定具有广泛识别性和增强效力的新适体，并描绘与跨进化枝酶抑制相关的二级结构。目标 3 检查细胞内表达的适体对假型和复制能力的 HIV-1 的抑制，从控制表达、定位和抑制的设计元素的优化转向适体和 RT 文库的高通量筛选。目标 4 评估适体耐药性的从头进化，重点是建立遗传阈值、耐药位点和耐药性的分子基础。