Therapeutic applications of interface-mimicking peptides for targeting the SARS-CoV-2 NSP12-NSP8 RdRp complex

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication depends on the NSP12-NSP8-NSP7 complex, which plays a critical role in enhancing RNA-dependent RNA polymerase (RdRp) activity. NSP8 is particularly essential, stabilizing the RdRp complex and supporting viral replication across diverse variants. To disrupt this crucial interaction, we designed four NSP8-derived peptides-N8-Pepα, N8-Pepα_cyc, N8-Pepβ, and N8-PepβD-targeting a key hotspot region within the NSP12-NSP8 interface that governs complex stability and processivity. In vitro assays demonstrated that these peptides effectively inhibit RdRp activity by disrupting the NSP12-NSP8 interaction, leading to significant reductions in SARS-CoV-2 replication in Vero E6 cells. Notably, intranasal administration of N8-Pepα or N8-Pepα_cyc (25 mg/kg) in Balb/c mice provided robust antiviral protection, alleviating weight loss and reducing mortality following challenge with a mouse-adapted SARS-CoV-2 strain. Both prophylactic and therapeutic treatments significantly lowered viral titers and minimized pathological damage in the nasal turbinates and lungs. These results highlight the NSP12-NSP8 interface as a novel and highly conserved target for antiviral therapy and establish NSP8-derived peptides, particularly N8-Pepα and N8-Pepα_cyc, as promising candidates for inhibiting RdRp complex formation and controlling SARS-CoV-2 replication.