Development and characterization of neutralizing monoclonal antibodies against clinically relevant human adenovirus subgroup B serotypes.

Human adenoviruses (HAdVs), particularly subgroup B serotypes HAdV-3 and HAdV-55, are associated with severe respiratory disease and currently lack targeted therapies. While neutralizing monoclonal antibodies (nMAbs) offer promising therapeutic potential, the specific nMAbs targeting these serotypes remain poorly characterized. Therefore, this study aimed to generate and evaluate the efficacy of serotype-specific nMAbs against HAdV-3 and HAdV-55. Mice were immunized with HAdV-3 virions, HAdV-55 virions, or recombinant fiber knob proteins (HAdV-55/-7) for the generation of serotype-specific nMAbs, and their efficacy was systematically evaluated using in vitro assays and an in vivo tree shrew model. Through comprehensive screening, eleven MAbs were identified with specificity against HAdV-3 (six clones) or HAdV-55/-7 fiber knob (five clones). Four nMAbs exhibited potent neutralizing activity: 13F12 (half-maximal inhibitory concentration, IC50: 3.8 μg/mL), 8D2 (IC50: 15.1 μg/mL), 3A3 (IC50: 14.9 μg/mL) against HAdV-3 virions, and 8F2 with neutralizing efficacy against HAdV-55 virions (IC50: 30.4 μg/mL). Western blot analysis revealed that MAbs 13F12 and 8F2 targeted the fiber protein, whereas 8D2 and 3A3 bound to the hexon protein. Furthermore, in vivo evaluations demonstrated that 13F12 significantly reduced viral loads in nasal turbinates and attenuated lung pathology in HAdV-3-infected tree shrews. Mechanistically, all tested anti-HAdV-3 nMAbs inhibited infection by blocking viral attachment (P < 0.01 vs. controls). In conclusion, this study underscores the therapeutic potential of targeting viral entry and highlights 13F12 as a promising candidate for HAdV prophylaxis.