Investigation of Quinoxaline-1,2,3-triazole Derivatives for Targeting SARS-CoV-2 via RBD Binding and PLpro Inhibition

Objective: A library of ten derivatives of 3-benzyl-N1-substituted quinoxalin-2-ones was designed and investigated for targeting SARS-CoV-2. Methods: The target compounds were synthesized as N1-substituted quinoxalines and quinoxaline/triazole hybrids via a click reaction. The anti-SARS-CoV-2 activity of these compounds was evaluated via spike protein and papain-like protease (PLpro) inhibition assays. Results and Discussion: The inhibition assays revealed a remarkable dual inhibitory activity for most compounds, ranging from 76.2 to 86.9%. Compounds (IIIb) and (IIIc) were identified as the most potent inhibitors based on enzyme kinetics studies. They exhibited a mixed inhibition type against both the PLpro enzyme and the spike protein. The most effective compound, (IIIc), demonstrated the lowest K_i competitive and K_i noncompetitive values for PLpro (0.23 ± 3 × 10^–4 and 0.57 ± 1 × 10^–3 µM) and spike protein (0.83 ± 1 × 10^–4 and 1.03 ± 2 × 10^–4 µM), respectively. These results indicate that compound (IIIb) acts as a competitive inhibitor (lower K_i value). Interestingly, molecular docking studies of both enzymes’ active sites revealed a significant binding affinity of our compounds, supporting the biological results. In silico prediction studies indicated that most of the candidates comply with Lipinski’s and Veber’s rules, demonstrating acceptable drug-likeness parameters and no predicted CNS side effects. Conclusions: The investigated compounds exhibited favorable inhibitory activity against SARS-CoV-2 and strong binding interactions, suggesting their potential as therapeutic candidates against COVID-19.