Bioactivity of Aptamers Targeted to HIV Reverse Transcriptase

描述（由申请人提供）：HIV逆转录酶（RT）是HIV复制所必需的，并且仍然是抗HIV治疗的主要目标。然而，由于艾滋病毒对抗艾滋病毒药物的耐药性迅速发展，有必要制定新的策略来对抗艾滋病毒感染。核酸适体是选择性地与特定分子靶标紧密结合的ssDNA或RNA分子。抗RT适配体已被证明在酶分析显著抑制HIV RT。一些适体被证明抑制多种HIV RTs（普遍型），而其他适体仅抑制特定的RTs（专一型）。虽然一些核酸适体已被证明可以抑制HIV复制，但核酸适体的生物活性尚未被彻底探索，HIV对适体产生抗性的潜在机制尚未确定。本提案的目标是使用单周期HIV复制和复制能力HIV检测来定义DNA和RNA适配体的生物活性。这些实验将比较普遍主义者和专家，以确定促进最有效抑制的适体成分。此外，该项目将探索艾滋病毒对DNA和RNA适配体的耐药性潜力，并确定产生耐药性的范围。特别地，将探讨普遍适体与专业适体产生的抵抗类型。由于普遍适体抑制如此多数量的RTs，希望病毒很难对这些适体产生抗性，抗性突变将降低病毒的适应性。这一建议为未来的研究奠定了坚实的基础，以探索更适用于临床环境的适体递送方法，并在更复杂的实验模型背景下研究适体对HIV复制的影响，如3D细胞培养系统或动物模型，以检查免疫系统在适体治疗中的作用。与公共卫生的相关性：大约10%的新艾滋病毒感染具有耐药性。DNA适体是一种可能的对抗艾滋病毒的武器，它非常稳定，而且很容易用世界上现有的技术廉价高效地合成，但是交付方法必须经过修改才能用于临床。RNA适体也是有吸引力的治疗候选者，在递送方面比DNA适体有几个优势，但尚未得到充分的研究，以确定成功治疗所需的通用成分。该项目将研究这两种适体，努力最终将其优势结合到一个治疗系统中。除了潜在的治疗应用之外，该项目还将增加对艾滋病毒耐药性的了解。

DESCRIPTION (provided by applicant): HIV reverse transcriptase (RT) is required for HIV replication and remains a primary target for anti-HIV therapies. However, due to the rapid development of HIV resistance to anti-HIV drugs, it is necessary to develop new strategies to combat HIV infection. Nucleic acid aptamers are ssDNA or RNA molecules that bind selectively and tightly to specific molecular targets. Anti-RT aptamers have been demonstrated to significantly inhibit HIV RT in enzymatic assays. Some aptamers were shown to inhibit a diverse panel of HIV RTs (universalist) while others inhibited only specific RTs (specialists). Although a few aptamers have been demonstrated to inhibit HIV replication, the bioactivity of nucleic acid aptamers has not been thoroughly explored and the potential mechanisms for the generation of HIV resistance to aptamers have not been determined. The goals of this proposal are to define the bioactivity of DNA and RNA aptamers using both single-cycle HIV replication and replication-competent HIV assays. These experiments will compare universalists and specialists to define aptamer components promoting the most potent inhibition. In addition, the project will explore the potential of HIV resistance to DNA and RNA aptamers and determine the scope of resistance generated. In particular, the type of resistance generated from universalist versus specialist aptamers will be explored. Since universalist aptamers inhibit such a diverse number of RTs, it is hoped that it will be difficult for the virus to develop resistance to these aptamers and that resistance mutations will decrease the fitness of the virus. This proposal sets a firm background for future studies to explore aptamer delivery methods more applicable to a clinical setting and also to study aptamer effects on HIV replication in the context of more complex experimental models, such as a 3D cell culture system or an animal model to examine the role of the immune system in aptamer therapeutics. Relevance to Public Health: Approximately 10% of new HIV infections are drug resistant. DNA aptamers are a possible weapon against HIV and are highly stable and easily synthesized cheaply and efficiently using technology available worldwide, but delivery methods must be modified for clinical use. RNA aptamers are also attractive candidates for therapeutics and have several advantages in delivery over DNA aptamers but have not been studied sufficiently to identify the universal components needed for successful therapy. This project will investigate both aptamers in an effort to eventually couple the advantages into one therapeutic system. In addition to the potential for therapeutic application, the project will increase understanding of HIV resistance.