Severity-associated cross-reactive anti-sarbecovirus antibody responses in COVID-19 convalescents and isolation of a dual-targeting monoclonal antibody with cross-neutralizing activity.

Background: Sarbecoviruses pose a persistent pandemic threat due to their zoonotic potential and high recombination capacity. This study investigated how disease severity influences cross-reactive antibody responses and the isolation of cross-neutralizing monoclonal antibodies. Methods: Plasma samples from 67 COVID-19 convalescents (severe, n=17; non-severe, n=50) were compared for cross-reactive antibody responses against seven sarbecovirus spike proteins. Memory B cells were sorted using dual S1/S2 fluorescent probes to isolate monoclonal antibodies. Results: Severe convalescents exhibited significantly higher neutralization titers against six animal-derived sarbecoviruses, responses that persisted for up to one year and were further enhanced by vaccination. From dual-targeting B cells, we isolated mAb 1D6, which simultaneously engages the S1 (ACE2-blocking) and S2 (fusion-inhibiting) domains. mAb 1D6 neutralized SARS-CoV-2 (IC50 = 5.708 μg/mL) and pangolin-CoV-GD (IC50 = 14.02 μg/mL). In contrast, antibody 1F2 bound both S1 and S2 across all tested sarbecoviruses but lacked neutralizing activity. Conclusions: Severe COVID-19 was associated with broader and more durable cross-sarbecovirus antibody responses. The dual-targeting antibody 1D6 provides proof-of-concept for a cross-neutralization strategy, although its modest potency requires further optimization for therapeutic application.