YL1004 is a SARS-CoV-2 papain-like protease inhibitor with immunomodulatory and antiviral activity in mice

The evolution of SARS-CoV-2, coupled with its immune evasion mechanisms, underscores the urgent need for antiviral strategies beyond vaccination. The papain-like protease (PL^pro) is a dual-functional enzyme essential for viral replication and suppression of host innate immunity. Here, we present the rational design and characterization of YL1004, a tricyclic oral PL^pro inhibitor demonstrating robust, cross-variant SARS-CoV-2 antiviral activity and favorable pharmacokinetic properties. YL1004 potently inhibits the enzymatic function of PL^pro, disrupts deubiquitination and deISGylation processes, and restores antiviral immune signaling cascades. Notably, YL1004 suppresses the replication of SARS-CoV-2 wildtype, Delta and Omicron variants. Additionally, it is also effective against the M^pro E166V recombinant SARS-CoV-2 strain, which confers resistance to nirmatrelvir. In the lethal SARS-CoV-2 infection model using K18-hACE2 mice, YL1004 confers complete protection to animal survival, significantly reduces viral load in nasal turbinate and lung tissues, and alleviates virus-induced pathological tissue damages. Co-crystal structural analysis revealed a distinctive binding mode, enhancing target engagement through expanded hydrophobic interactive interface and additional hydrogen bonding interactions. Collectively, these findings establish YL1004 as a promising therapeutic candidate, harnessing dual antiviral and immunomodulatory mechanisms to combat SARS-CoV-2 and emerging variants.