Enhanced rhLCV production in lymphoblastoid cell lines derived from rhLCV-infected cynomolgus macaque PBMCs

ABSTRACT Epstein-Barr virus (EBV) infects more than 90% of adults worldwide and causes a range of diseases, including multiple malignancies and autoimmune disorders. However, due to a host range restriction, EBV cannot infect commonly used experimental animals, posing a significant obstacle to developing EBV-specific prophylactic and therapeutic agents. Rhesus lymphocryptovirus (rhLCV), an ortholog of EBV, naturally infects rhesus macaques, which is a surrogate model for EBV research. In this study, we demonstrate that cynomolgus macaque (Macaca fascicularis), a primate closely related to rhesus macaque, is susceptible to rhLCV infection. rhLCV can immortalize B cells of cynomolgus macaques to develop cy-LCLs. We developed a high rhLCV-producing cy-LCL cell line, LCL111, and optimized the induction conditions to increase viral production, surpassing the original rhLCV producer LCL8664. Importantly, EBV gHgL-specific monoclonal antibody (mAb) AMMO1 and gB-specific mAb 3A5 can cross-react with rhLCV proteins and block the formation of cy-LCLs. Overall, we established an efficient rhLCV-producing cell line, and rhLCV infection of cynomolgus macaques represents a promising alternative surrogate model for efficiency evaluation of EBV vaccines and mAbs. IMPORTANCE Epstein-Barr virus (EBV) naturally infects only humans, creating a major barrier to evaluating the efficiency of vaccines and therapies in vivo . As an EBV ortholog, rhesus lymphocryptovirus (rhLCV) offers a biologically relevant surrogate system. However, its application has been primarily limited to rhesus macaques. Here, we demonstrate that cynomolgus macaque lymphocytes are also susceptible to rhLCV in vitro, and the newly transformed cy-LCL111 shows superior and sustained rhLCV production ability. rhLCV infection of cynomolgus macaque lymphocytes can be efficiently neutralized by anti-EBV gH/gL nAbs AMMO1 and anti-EBV gB mAbs 3A5, highlighting the potential of cynomolgus macaques as an in vivo model to assess anti-EBV mAb and vaccine efficacy. Our findings support the use of cynomolgus macaques as an additional model for EBV research and offer a useful platform for evaluating EBV-specific prophylactic or therapeutic strategies.