Bivalent nanoparticle vaccine elicits cross-protective antibody against oncogenic herpesviruses

Background: Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV) are oncogenic gamma-herpesviruses causally linked to multiple malignancies but lack approved vaccines. As virus-associated cancers represent a substantial global tumor burden, preventive vaccination offers a rational approach to cancer prevention. Both viruses depend on the conserved gHgL heterodimer for receptor engagement and entry, suggesting gHgL as a shared vaccine target for broad gamma-herpesvirus protection. Methods: We developed a bivalent nanoparticle vaccine (bi-NP) that co-displays EBV and KSHV gHgL on the self-assembling I53-50 scaffold and compared its immunogenicity and protecting efficacy with single-antigen and soluble counterparts. Structural and antigenic integrity of immunogens were validated by size-exclusion chromatography, negative-stain electron microscopy and binding to specific monoclonal antibodies. Immunogenicity was evaluated in mice, rabbits, and non-human primates (NHPs), while in vivo protection was evaluated using heterologous (murine gamma-herpesvirus 68, MHV-68) and homologous (EBV/KSHV co-infection in humanized mice) challenge models. Results: The bivalent gHgL nanoparticle elicited consistently higher and more balanced antibody responses against both EBV and KSHV across species, effectively blocking viral infection of their respective susceptible cell types. Bi-NP immunization induced cross-reactive antibodies recognizing conserved gHgL epitopes, increasing the frequency of B cells cross-binding both EBV and KSHV gHgL in rabbits, eliciting antibodies cross-neutralizing MHV-68, and conferring in vivo cross-protection against MHV-68 mice challenge. Moreover, passive transfer of IgG purified from bi-NP-immunized macaques protected humanized mice from EBV/KSHV co-infection, markedly reducing viral DNA copies, preventing weight loss, and diminishing EBER + and LANA + pathology. Conclusions: This study establishes gHgL as a rational bivalent vaccine target and demonstrates that nanoparticle co-display of EBV and KSHV antigens elicits cross-reactive, broadly protective antibodies across divergent gamma-herpesviruses. The findings highlight a promising platform for preventive strategies against gamma-herpesvirus-associated cancers and potential zoonotic infections.