ANTIVIRAL EFFICACY OF OLIGONUCLEOTIDES TO HIV-1 POLYPURINE-RICH SEQUENCES

旨在通过以下方式影响HIV致病的合成寡核苷酸 与病毒核酸的序列特异性相互作用显示出希望 作为抗病毒治疗的候选对象。我们一直在探索 寡核苷酸治疗的疗效比较(针对不同的 病毒复制周期的不同阶段)。 我们使用的寡聚蛋白家族的目标是同样富含多嘌呤的 序列，但带有不同的修饰，以便它们可以发挥作用 作为三螺旋(TH)、反义(AS)或非特异性抑制剂。 设计成三螺旋(TH)的ONS治疗感染细胞 随着靶HIV序列p24抗原水平的显著降低 以及减少病毒DNA和RNA的数量 未经处理的细胞。AS ONS对感染细胞的处理也受到抑制 艾滋病毒的表达，但以更有限和较少的序列特异性的方式。 寡聚体迅速被细胞摄取，进入细胞核内 10分钟，并保持可检测到的非降级水平长达48 几个小时。这些寡核苷酸与它们的靶序列显示出一定的相互作用 在体外试验中，但这种联系并不稳定。我们有 继续这些研究，使用携带不同修饰的TH寡核苷酸 设计用于1)增加TH结构的稳定性，2)提供 对细胞核酸酶有更强的抵抗力，以及3)增强抗病毒作用 目标交互的活动。我们也已经开始研究 探讨抑制病毒的主要作用方式 复制。这些研究是在组织培养细胞中进行的。 细胞系(H9细胞)和原代正常人外周血 淋巴细胞(PBL)，使用实验室和初级分离株。

Synthetic oligonucleotides designed to affect HIV pathogenesis by sequence-specific interactions with viral nucleic acid have shown promise as candidates for antiviral therapy. We have been exploring the comparative efficacy of oligonucleotide treatment (aimed at different stages in the viral replicative cycle) of HIV-1 infected cells in vitro. The family of oligos we have used target the same polypurine-rich sequences, but carry different modifications such that they may function as triple helix (TH), antisense (AS), or nonspecific inhibitors. Treatment of infected cells with ONs designed to form a Triple Helix (TH) with the target HIV sequence significantly decreased p24 antigen levels as well as decreasing the amounts of viral DNA and RNA relative to untreated cells. Treatment of infected cells with AS ONs also inhibited HIV expression, but in a more limited and less sequence specific manner. The oligos were rapidly taken up by the cells, entering the nuclei within 10 minutes and maintaining detectable, non-degraded levels for up to 48 hours. These oligos showed some interaction with their target sequences in in vitro assays, but the association was not stable. We have continued these studies with TH oligos carrying different modifications designed to 1) increase the stability of the TH structure, 2) provide more resistance to cellular nucleases, and 3) enhance the antiviral activity of the ON - target interaction. We have also begun studies to investigate the primary mode of action in ON inhibition of viral replication. These studies have been conducted in tissue culture cell lines (H9 cells) and with primary normal human peripheral blood lymphocytes (PBLs), using laboratory and primary isolates.