biostudies-literatureLizzadro LNational Institute of Allergy and Infectious DiseasesGladstone InstitutesNIAID NIH HHSRoddenberry FoundationP. and E. Taft1530-1551https://www.ebi.ac.uk/biostudies/studies/S-EPMC12834036biostudies-literatureUnknown69(2)The main proteases (M^Pro) of coronaviruses are clinically validated targets for antiviral discovery. Herein, we detail the in vivo optimization of uracil-core M^Pro inhibitors derived from AVI-4516, an in vivo active lead bearing an unactivated propargyl warhead. To expand the anticoronaviral spectrum, we introduced diverse C6 substitution to target the S1' pocket in M^Pro and observed enhanced cellular activity against various nirmatrelvir-resistant mutants. Pharmacokinetic profiling of 12 analogs revealed overall inferior exposure of the C6 aryl analogs. However, PK profiling across three species identified the improved atropisomeric lead (<i>M</i>)-AVI-4773 (5-(5,6-difluoro-1<i>H</i>-benzo[<i>d</i>][1,2,3]triazol-1-yl)-3-((<i>M</i>)-isoquinolin-4-yl)-6-metJournal of medicinal chemistryDiscovery of Coronavirus Main Protease Inhibitors with Enhanced Brain Exposure and Potent Oral Efficacy in SARS-CoV-2 and MERS Infection Models.PMC12834036U19AI171110U19 AI171110G20AI174733Garcia-Sastre AMcCann JTRenslo ARLizzadro LDegotte GAlbrecht RHenderson WRSuazo IDCraik CSHantz ERSettineri NSWhite KMDiallo AMcGovern BLVerba KAKrogan NJLi JZapatero-Belinchon FJMontano MDetomasi TCBenjamin JTaha TYGordon MEShoichet BKDiaz-Tapia RHuang SOtt MMatsui YRosecrans JTorres Pomares DFfalseDiscovery of Coronavirus Main Protease Inhibitors with Enhanced Brain Exposure and Potent Oral Efficacy in SARS-CoV-2 and MERS Infection Models.The main proteases (M^Pro) of coronaviruses are clinically validated targets for antiviral discovery. Herein, we detail the in vivo optimization of uracil-core M^Pro inhibitors derived from AVI-4516, an in vivo active lead bearing an unactivated propargyl warhead. To expand the anticoronaviral spectrum, we introduced diverse C6 substitution to target the S1' pocket in M^Pro and observed enhanced cellular activity against various nirmatrelvir-resistant mutants. Pharmacokinetic profiling of 12 analogs revealed overall inferior exposure of the C6 aryl analogs. However, PK profiling across three species identified the improved atropisomeric lead (<i>M</i>)-AVI-4773 (5-(5,6-difluoro-1<i>H</i>-benzo[<i>d</i>][1,2,3]triazol-1-yl)-3-((<i>M</i>)-isoquinolin-4-yl)-6-met2026-01-01T00:00:00Z2026 Jan2026-06-09T07:21:21.365Z2026-06-09T03:11:28.424ZS-EPMC128340364149020610.1021/acs.jmedchem.5c03015