Immune defense of cCMV at the maternal-fetal interface

ABSTRACT - PROJECT 1 Congenital cytomegalovirus (cCMV) is the most common infectious cause of sensorineural hearing loss and neurological sequelae in the newborn. Efforts to achieve a maternal vaccine have not yet met with success, in part due to an incomplete understanding of the correlates of protection. In Project 1, we will use a nonhuman primate (NHP) model of cCMV in rhesus macaques (RMs) to interrogate systemic and tissue-level immune determinants of placental CMV transmission at the maternal-fetal (M-F) interface. We hypothesize that an effective cCMV vaccine will need to elicit a cellular response at the M-F interface that balances induction of protective virus-specific immunity with prevention of immunopathology. We will investigate cellular immune defenses at the M-F interface in infection and vaccine settings as follows. Aim 1: Characterize virus-host immune interactions at the M-F interface in primary RhCMV infection and their role in cCMV outcome. Placental tissues and cells from immunocompetent RM dams infected with RhCMV will be subjected to viral and immunologic analysis using PCR, RNAScope, multiplex immunohistochemistry, spatial transcriptomics, sc- RNASeq, and cellular assays for analysis of virus and host immune interactions at the M-F interface in the presence or absence of cCMV transmission. Aim 2: Determine the immunogenicity and durability of single- cycle (sc) RhCMV vaccines eliciting MHC-Ia-restricted “conventional” or MHC-II/MHC-E-restricted “unconventional” virus-specific CD8+ T cells in female RMs and characterize their ability to induce vaccine- specific T cell responses at the M-F interface. Breeding age CMV-seronegative dams vaccinated with a prime- boost regimen of sc-RhCMV vaccines will be timed-mated and evaluated for vaccine-induced T lymphocyte responses in tissues of the M-F interface at term gestation. Aim 3: Determine the protective effect of pre- conception RhCMV vaccine-induced conventional and unconventional CD8+ T lymphocyte responses against congenital transmission. Vaccinated, pregnant dams will be challenged with RhCMV in early 2nd trimester gestation and monitored for protection against cCMV transmission. Aim 1 experiment animals will serve as controls for vaccine groups in Projects 1-3. Project 1 will interact with the MODS Core for organoids, spatial transcriptomics, and spatial proteomics; with the Virology Core for RhCMV challenge virus stocks, RhCMV vaccines, quantitative PCR for RhCMV DNA, and viral sequencing; and with the IDS Core for data analysis, including bioinformatics. Virus-specific immunity and T cell trafficking to the M-F interface is relevant to understanding the type of tissue level immunity that a vaccine needs to protect against cCMV without inducing harmful immunopathology and an adverse pregnancy outcome.