Non-CpG-Containing Antisense 2-Methoxyethyl Oligonucleotides Activate a Proinflammatory Response Independent of Toll-Like Receptor 9 or Myeloid Differentiation Factor 88

Oligonucleotides with a “CpG ” motif trigger a proinflammatory response through activation of Toll-like receptor 9 (TLR9) and are being studied to exploit these properties for use as adju-vants and cancer therapies. However, oligonucleotides in-tended for antisense applications (ASOs) are designed to min-imize proinflammatory responses by avoiding CpG motifs and by using chemical modifications [i.e., 2-methoxyethyl (MOE) sugars and 5-methyl cytosine residues]. Nonetheless, modified ASOs are capable of eliciting a proinflammatory response at high doses, albeit mild compared with CpG oligos. To deter-mine whether this phenomena is TLR-mediated, wild-type, TLR9 knockout, and myeloid differentiation factor 88 (MyD88) knockout mice were treated with a phosphorothioate-modified oligodeoxyribonucleotide CpG optimal oligo (ISIS 12449), and