Pfizer Oligonucleotides Therapeutics Unit, Coley Pharmaceutical GmbH, Düsseldorf, Germany.
Nucleic Acid Ther. 2011 Dec;21(6):423-36. doi: 10.1089/nat.2011.0323.
The toll-like receptors (TLRs) 7, 8, and 9 stimulate innate immune responses upon recognizing pathogen nucleic acids. Certain GU- or AU-rich RNA sequences were described to differentiate between human TLR7- and TLR8-mediated immune effects. Those single-stranded RNA molecules require endosomal delivery for stabilization against ribonucleases. We have discovered RNA sequences that preferentially activate TLR7, form higher ordered structures, and do not require specific cellular delivery. In addition, a dual activation of TLR8 and TLR9 without affecting TLR7 can be achieved by chimeric molecules consisting of GU-rich RNA and Cytosin (C) phosphordiester or phosphorthioat (p) guanine (CpG) motif DNA sequences. Such chimeras stimulate TLR9-mediated type I interferon (IFN) and TLR8-depending proinflammatory cytokine and chemokine production upon primary human cell activation. However, an RNA-dependent TLR7 IFN-α cytokine release is suppressed by the phosphorothioate DNA sequence contained in the chimeric molecule. To convert the immune response of a single-stranded RNA from TLR7/8 to TLR9, a simple chemical modification at the 5' end proves to be sufficient. Such 8-oxo-2'-deoxy-guanosine or 8-bromo-2'-deoxy-guanosine modifications of the first guanosine in GU-rich single-stranded RNAs convert the immune response to include TLR9 activation and demonstrate strong additive effects for type I IFN immune responses in human primary cells.
Toll 样受体（TLRs）7、8 和 9 在识别病原体核酸后会刺激先天免疫反应。某些富含鸟嘌呤（GU）或尿嘧啶（AU）的 RNA 序列被描述为区分人类 TLR7 和 TLR8 介导的免疫效应。这些单链 RNA 分子需要内体递送来抵抗核糖核酸酶。我们已经发现了优先激活 TLR7、形成更高阶结构且不需要特定细胞递送的 RNA 序列。此外，通过由富含 GU 的 RNA 和胞嘧啶（C）磷酸二酯或硫代磷酸（p）鸟嘌呤（CpG）基序 DNA 序列组成的嵌合分子，可以实现 TLR8 和 TLR9 的双重激活，而不影响 TLR7。这种嵌合体在原代人细胞激活时刺激 TLR9 介导的 I 型干扰素（IFN）和 TLR8 依赖的促炎细胞因子和趋化因子的产生。然而，嵌合分子中包含的硫代磷酸 DNA 序列抑制了依赖 RNA 的 TLR7 IFN-α细胞因子释放。为了将单链 RNA 的免疫反应从 TLR7/8 转换为 TLR9，只需在 5'端进行简单的化学修饰即可。富含 GU 的单链 RNA 中第一个鸟嘌呤的 8-氧-2'-脱氧鸟嘌呤或 8-溴-2'-脱氧鸟嘌呤修饰将免疫反应转换为包括 TLR9 激活，并在原代人细胞中显示出对 I 型 IFN 免疫反应的强烈相加作用。
