Blockade of PD-1 and LAG-3 Immune Checkpoints Combined with Vaccination Restores the Function of Antiviral Tissue-Resident CD8⁺T_RMCells and Reduces Ocular Herpes Simplex Infection and Disease in HLA Transgenic Rabbits

Abstract

Chronic viruses such as herpes simplex virus 1 (HSV-1) evade the hosts’ immune system by inducing the exhaustion of antiviral T cells. In the present study, we found that exhausted HSV-specific CD8+ T cells, with elevated expression of programmed death ligand-1 (PD-1) and lymphocyte activation gene-3 (LAG-3) receptors were frequent in symptomatic patients, with a history of numerous episodes of recurrent corneal herpetic disease, compared to asymptomatic patients who never had corneal herpetic disease. Subsequently, using a rabbit model of recurrent ocular herpes, we found that the combined blockade of PD-1 and LAG-3 pathways with antagonist antibodies significantly restored the function of tissue-resident antiviral CD8+ TRM cells in both the cornea and the trigeminal ganglia (TG). An increased number of functional tissue-resident HSV-specific CD8+ TRM cells in latently infected rabbits was associated with protection against recurrent herpes infection and disease. Compared to the PD-1 or LAG-3 blockade alone, the combined blockade of PD-1 and LAG-3 appeared to have a synergistic effect in generating frequent polyfunctional Ki-67+, IFN-γ+, CD107+, and CD8+ T cells. Moreover, using the human leukocyte antigen (HLA) transgenic rabbit model, we found that dual blockade of PD-1 and LAG-3 reinforced the effect of a multiepitope vaccine in boosting the frequency of HSV-1-specific CD8+ TRM cells and reducing disease severity. Thus, both the PD-1 and the LAG-3 exhaustion pathways play a fundamental role in ocular herpes T cell immunopathology and provide important immune checkpoint targets to combat ocular herpes. IMPORTANCE HSV-specific tissue-resident memory CD8+ TRM cells play a critical role in preventing virus reactivation from latently infected TG and subsequent virus shedding in tears that trigger the recurrent corneal herpetic disease. In this report, we determined how the dual blockade of PD-1 and LAG-3 immune checkpoints, combined with vaccination, improved the function of CD8+ TRM cells associated with a significant reduction in recurrent ocular herpes in HLA transgenic (Tg) rabbit model. The combined blockade of PD-1 and LAG-3 appeared to have a synergistic effect in generating frequent polyfunctional CD8+ TRM cells that infiltrated both the cornea and the TG. The preclinical findings using the established HLA Tg rabbit model of recurrent herpes highlight that blocking immune checkpoints combined with a T cell-based vaccine would provide an important strategy to combat recurrent ocular herpes in the clinic.

Unique molecular signatures of antiviral memory CD8+ T cells associated with asymptomatic recurrent ocular herpes

“…Exhaustion of HSV-specific CD8 + T cells may occur during multiple productive replication attempts and/or repetitive virus reactivations, events that might be underestimated with actual molecular detections 73 . Moreover, we found that LAG-3 and PD-1 immune checkpoints blockade in HSV-1 latently infected HLA Tg rabbits restored T cell functions associated with less recurrent ocular herpes infection and disease, confirming our previous studies 18,28,31,35 . The findings in this report prompt to question whether HSV-1 infections can be categorized as a latent or chronic infection.…”

“…flow cytometry assays. Trigeminal ganglia were analyzed by flow cytometry, as we previously reported 86 .…”

“…In the present study, we performed bulk and scRNASeq transcriptomic, phenotypic and functional analyses of blood and tissue-resident CD8 + T cells, specific to multiple and same HLA-A*0201-restricted HSV-1 epitopes from HSV-1 infected symptomatic and asymptomatic patients and from our HLA-A*0201 transgenic rabbit (HLA Tg rabbit) model of recurrent ocular herpes 18,28,31,35 . The identified molecular, phenotypic, and functional signatures of antiviral CD8 + T cells, sharing the same HSV-1 epitope-specificities, were confirmed in HSV-1-infected symptomatic and asymptomatic HLA Tg rabbits.…”

Immune Checkpoints in Viral Infections

“…Related studies show that both CD4 + T-cells and CD8 + T-cells in draining lymph nodes have increased expression levels of PD-1, TIM-3, and LAG-3 after HSV infection [86,87]. This upregulated PD-1 and LAG3 expression led to the exhaustion of tissue-resident CD8 + T-cells and the loss of antiviral response to symptomatic reactivation, whereas a blockade of LAG-3 and PD-1 improved the function of HSV-specific CD8 + T-cells and significantly reduced the reactivation of latent HSV virus in a rabbit model [88,89].…”

Human Epitopes Identified from Herpes Simplex Virus Tegument Protein VP11/12 (UL46) Recall Multifunctional Effector Memory CD4+TEMCells in Asymptomatic Individuals and Protect from Ocular Herpes Infection and Disease in “Humanized” HLA-DR Transgenic Mice

“…There is substantial literature currently on the role of CD8 ϩ T cells in the protection against herpes, whereas the role of CD4 ϩ T cells in such protection is relatively less known (10)(11)(12)(13)(14). Activated CD4 ϩ T cells are retained in latently infected trigeminal ganglia (TG) of mice and humans.…”