Synthesis of Dual Inhibitors Targeting SARS-CoV-2 Mpro and hCatL with Selectivity for hCatS

• Grant number: 24/10359-0
• Support Opportunities: Regular Research Grants
• Start date: August 01, 2025
• End date: July 31, 2028
• Field of knowledge: Physical Sciences and Mathematics - Chemistry
• Principal Investigator: Carlos Alberto Montanari
• Grantee: Carlos Alberto Montanari
• Host Institution: Instituto de Química de São Carlos (IQSC). Universidade de São Paulo (USP). São Carlos , SP, Brazil

Abstract

Our extensive efforts to develop inhibitors for human cathepsin L and SARS-CoV-2 main protease (Mpro) have led to the identification of a set of antiviral agents exhibiting superior efficacy compared to nirmatrelvir (synthesized in our laboratory and dubbed Neq1183). Preliminary studies on cytochrome P450 enzymes revealed that Neq1184 shows no activity, indicating that it is a superior candidate to nirmatrelvir concerning CYP3A4 substrate specificity. Neq1184 does not inhibit hCatL and is a weak Mpro inhibitor with a pKi of 6.7, but surpasses Neq1183 as an anti-SARS-CoV-2 agent. Neq1119, with a new in-house engineered scaffold, is sufficiently selective towards CYPs to allow a positive assessment of its degree of metabolism. Neq1119 is an hCatL inhibitor at low-nanomolar concentrations (pKi = 8.8) but is inactive against Mpro and 100 % effective against SARS-CoV-2. Neq1167, independently developed in our Lab, is a dual inhibitor of hCatL and Mpro, and it also has 100 % efficacy against SARS-CoV-2. The 100 % effective antiviral agent, Neq1153, is also a dual inhibitor of hCatL at low-nanomolar concentration (pKi = 8.7) and Mpro with a pKi of 7.4. hCatL is involved in processing viral proteins necessary for viral replication and infectivity. hCatS plays a role in immune responses and other cellular processes, and it is not directly implicated in viral replication. Achieving selectivity for hCatL inhibitors over hCatS is a crucial aspect of drug discovery and development and aligns with the principles of precision medicine, where treatments are tailored to target specific molecules or pathways involved in disease. For instance, Neq1119 is selective for hCatL over hCatS by a difference of 1.7 log units. While Neq1153 is a Mpro low-nanomolar inhibitor and equipotent against hCatL and hCatS, Neq1167 is a Mpro inhibitor equipotent to hCatS, but with a lower affinity towards hCatL. Because hCatS is primarily associated with antigen processing and presentation via the MHC class II pathway and its dysregulation is implicated in autoimmune diseases and chronic inflammatory conditions, we aim to improve the dual affinity of hCatL and SARS-CoV-2 Mpro inhibitors while producing weak or inactive inhibitors for hCatS. (AU)