Discovery and Mechanistic Characterization of Tetrahydroisoquinoline Derivatives as Inhibitors of SARS‐CoV‐2 Main Protease

SARS‐CoV‐2 main protease (Mpro) is an enzyme for viral replication and an target for antiviral drugs due to its evolutionary conservation across coronaviruses and the absence of closely related human homologs. Tetrahydroisoquinoline (THIQ) derivatives represent a structurally diverse class of nitrogen‐containing heterocycles with proven versatility in medicinal chemistry. Thirty newly synthesized compounds, including THIQ‐based molecules and structurally related amine analogs, were evaluated for their inhibitory activity against Mpro. Six compounds exhibited significant inhibitory potency (IC ₅₀  < 50 μM). Compound 3ia, containing nitro and para ‐bromophenyl groups, was the most active candidate with an IC ₅₀ of 6.61 μM. Fluorescence quenching and thermodynamic analyses revealed that the interactions between active compounds and Mpro occurred predominantly via static quenching. Results demonstrated that compound binding induced conformational changes in Mpro. Differential scanning fluorimetry (DSF) showed that all active compounds increased the thermal stability of Mpro, and microscale thermophoresis (MST) confirmed high‐affinity binding ( K _d  = 25.92 μM for 3ia ). Molecular docking further revealed that the THIQ scaffold of 3ia inserts into the Mpro substrate‐binding pocket, forming π–sulfur interactions with the catalytic Cys145. This work confirms THIQ as a promising lead scaffold for Mpro inhibitors and supports the structure‐based design of improved anticoronavirus candidates.