Multiple chemical series emerged from this exercise (Table , Figure , and Supplementary material). Thirteen of the high ranked compounds as well as compound derivatives that scored lower but exhibited high binding affinity by SPR were further assayed for inhibition of double-stranded RNA unwinding activity in a FRET-based assay. Two compounds were found to potently inhibit Nsp13 helicase activity (CACHE2-HO_1431_6: K D 770 nM ± 180, unwinding IC 50 8.6 µM ± 1.7; CACHE2-HO_1454_15: K D 31 µM ± 0.7, unwinding IC 50 57 µM ± 2). Inhibition of dsRNA unwinding by CACHE2-HO_1431_6 was also confirmed in a gel-based unwinding assay with CACHE2-HO_1454_15 partially inhibitory, consistent with CACHE2-HO_1431_6 having a more potent unwinding activity and stronger binding affinity and lower IC 50 value. (Figure ). Note that many compounds in this series have an ester group linker that is likely to be hydrolyzed in cells and represents a serious medicinal chemistry liability, which penalized the final score of this chemical series. However, modifying the ester linker to a more stable group could easily address this liability while conserving potency. But this medicinal chemistry work is beyond the scope of the CACHE study.