@article {Si2021.11.19.469183, author = {Longlong Si and Haiqing Bai and Crystal Yuri Oh and Tian Zhang and Fan Hong and Amanda Jiang and Yongxin Ye and Tristan X. Jordan and James Logue and Marisa McGrath and Chaitra Belgur and Atiq Nurani and Wuji Cao and Rachelle Prantil-Baun and Steven P Gygi and Rani K. Powers and Matthew Frieman and Benjamin R. tenOever and Donald E. Ingber}, title = {Self-assembling short immunostimulatory duplex RNAs with broad spectrum antiviral activity}, elocation-id = {2021.11.19.469183}, year = {2021}, doi = {10.1101/2021.11.19.469183}, publisher = {Cold Spring Harbor Laboratory}, abstract = {The current COVID-19 pandemic highlights the need for broad-spectrum antiviral therapeutics. Here we describe a new class of self-assembling immunostimulatory short duplex RNAs that potently induce production of type I and type III interferon (IFN-I and IFN-III), in a wide range of human cell types. These RNAs require a minimum of 20 base pairs, lack any sequence or structural characteristics of known immunostimulatory RNAs, and instead require a unique conserved sequence motif (sense strand: 5{\textquoteright}-C, antisense strand: 3{\textquoteright}-GGG) that mediates end-to-end dimer self-assembly of these RNAs by Hoogsteen G-G base-pairing. The presence of terminal hydroxyl or monophosphate groups, blunt or overhanging ends, or terminal RNA or DNA bases did not affect their ability to induce IFN. Unlike previously described immunostimulatory siRNAs, their activity is independent of TLR7/8, but requires the RIG-I/IRF3 pathway that induces a more restricted antiviral response with a lower proinflammatory signature compared with poly(I:C). Immune stimulation mediated by these duplex RNAs results in broad spectrum inhibition of infections by many respiratory viruses with pandemic potential, including SARS-CoV-2, SARS-CoV, MERS-CoV, and influenza A, as well as the common cold virus HCoV-NL63 in both cell lines and human Lung Chips that mimic organ-level lung pathophysiology. These short dsRNAs can be manufactured easily, and thus potentially could be harnessed to produce broad-spectrum antiviral therapeutics at low cost.Competing Interest StatementD.E.I. is a founder, board member, SAB chair, and equity holder in Emulate Inc. D.E.I., L. S., H. B., C.O., and R.P. are inventors on relevant patent applications held by Harvard University.}, URL = {https://www.biorxiv.org/content/early/2021/11/22/2021.11.19.469183}, eprint = {https://www.biorxiv.org/content/early/2021/11/22/2021.11.19.469183.full.pdf}, journal = {bioRxiv} }