Sequence motifs in adenoviral DNA block immune activation by stimulatory CpG motifs

Abstract

Unmethylated CpG dinucleotides in particular base contexts (CpG-S motifs) are relatively common in bacterial DNA but are rare in vertebrate DNA. B cells and monocytes have the ability to detect such CpG-S motifs that trigger innate immune defenses with production of Th1-like cytokines. Despite comparable levels of unmethylated CpG dinucleotides, DNA from serotype 12 adenovirus is immunestimulatory, but serotype 2 is nonstimulatory and can even inhibit activation by bacterial DNA. In type 12 genomes, the distribution of CpG-f lanking bases is similar to that predicted by chance. However, in type 2 adenoviral DNA the immune stimulatory CpG-S motifs are outnumbered by a 15-to 30-fold excess of CpG dinucleotides in clusters of direct repeats or with a C on the 5 side or a G on the 3 side. Synthetic oligodeoxynucleotides containing these putative neutralizing (CpG-N) motifs block immune activation by CpG-S motifs in vitro and in vivo. Eliminating 52 of the 134 CpG-N motifs present in a DNA vaccine markedly enhanced its Th1-like function in vivo, which was increased further by the addition of CpG-S motifs. Thus, depending on the CpG motif, prokaryotic DNA can be either immune-stimulatory or neutralizing. These results have important implications for understanding microbial pathogenesis and molecular evolution and for the clinical development of DNA vaccines and gene therapy vectors.

Suppressive Oligodeoxynucleotides Protect Mice from Lethal Endotoxic Shock

“…1A). In contrast, animals treated with the other class of suppressive ODN (known to selectively block CpG-induced immune activation, typified by ODN 2010) (21) were not protected (Fig. 1B).…”

Influence of stimulatory and suppressive DNA motifs on host susceptibility to inflammatory arthritis

“…TCR ϭ T cell receptor. activation and may thus limit the pathologic changes caused by CpG-driven immune responses (10,13,25).…”

“…This immune activation can be inhibited by "suppressive" ODNs, which block the production of Th1 and proinflammatory cytokines (10)(11)(12)(13). Suppressive motifs are rich in poly G or GC sequences, tend to be methylated, and are present in the DNA of mammals and certain viruses (10,11,14).…”

A gene therapy for cancer based on the angiogenesis inhibitor, vasostatin

“…[13][14][15] The expression plasmid pSecTag2B-vaso and the empty vector pSecTag2B were purified by using two rounds of passage over EndoFree columns (Qiagen, Chatsworth, CA), as reported previously by us 36 and others. 37 Western blot analysis Western blot analysis was performed as described. 18 Briefly, COS cells were plated in six-well plates at 2 × 10 5 cells/well, and transfected with 2 g of plasmid and 4 g lipofectin (Life Technologies, Grand Island, NY, USA) in serum-free DMEM following the manufacturer's instructions.…”