A Trivalent HSV-2 gC2, gD2, gE2 Nucleoside-Modified mRNA-LNP Vaccine Provides Outstanding Protection in Mice against Genital and Non-Genital HSV-1 Infection, Comparable to the Same Antigens Derived from HSV-1.

HSV-1 disease is a significant public health burden causing orofacial, genital, cornea, and brain infection. We previously reported that a trivalent HSV-2 gC2, gD2, gE2 nucleoside-modified mRNA-lipid nanoparticle (LNP) vaccine provides excellent protection against vaginal HSV-1 infection in mice. Here, we evaluated whether this HSV-2 gC2, gD2, gE2 vaccine is as effective as a similar HSV-1 mRNA LNP vaccine containing gC1, gD1, and gE1 in the murine lip and genital infection models.

Novel Adjuvant S-540956 Targets Lymph Nodes and Reduces Genital Recurrences and Vaginal Shedding of HSV-2 DNA When Administered with HSV-2 Glycoprotein D as a Therapeutic Vaccine in Guinea Pigs.

Herpes simplex virus type 2 (HSV-2) is a leading cause of genital ulcer disease and a major risk factor for acquisition and transmission of HIV. Frequent recurrent genital lesions and concerns about transmitting infection to intimate partners affect the quality of life of infected individuals. Therapeutic vaccines are urgently needed to reduce the frequency of genital lesions and transmission.

Antibodies to Crucial Epitopes on HSV-2 Glycoprotein D as a Guide to Dosing an mRNA Genital Herpes Vaccine.

The toxicity of mRNA-lipid nanoparticle (LNP) vaccines depends on the total mRNA-LNP dose. We established that the maximum tolerated dose of our trivalent mRNA-LNP genital herpes vaccine was 10 μg/immunization in mice. We then evaluated one of the mRNAs, gD2 mRNA-LNP, to determine how much of the 10 μg total dose to assign to this immunogen.

Guinea Pig and Mouse Models for Genital Herpes Infection.

This article describes procedures for two preclinical animal models for genital herpes infection. The guinea pig model shares many features of genital herpes in humans, including a natural route of inoculation, self-limiting primary vulvovaginitis, spontaneous recurrences, symptomatic and subclinical shedding of HSV-2, and latent infection of the associated sensory ganglia (lumbosacral dorsal root ganglia, DRG). Many humoral and cytokine responses to HSV-2 infection in the guinea pig have been characterized; however, due to the limited availability of immunological reagents, assessments of cellular immune responses are lacking.

Trivalent nucleoside-modified mRNA vaccine yields durable memory B cell protection against genital herpes in preclinical models.

Nucleoside-modified mRNA vaccines have gained global attention because of COVID-19. We evaluated a similar vaccine approach for preventing a chronic, latent genital infection rather than an acute respiratory infection. We used animal models to compare an HSV-2 trivalent nucleoside-modified mRNA vaccine with the same antigens prepared as proteins, with an emphasis on antigen-specific memory B cell responses and immune correlates of protection.

Techniques for Characterizing Cytomegalovirus-Encoded miRNAs.

microRNAs (miRNAs) are small noncoding RNAs that regulate gene expression at the posttranscriptional level by binding to sites within the 3' untranslated regions of messenger RNA (mRNA) transcripts. The discovery of this completely new mechanism of gene regulation necessitated the development of a variety of techniques to further characterize miRNAs, their expression, and function. In this chapter, we will discuss techniques currently used in the miRNA field to detect, express and inhibit miRNAs, as well as methods used to identify and validate their targets, specifically with respect to the miRNAs encoded by human cytomegalovirus.

An HSV-2 nucleoside-modified mRNA genital herpes vaccine containing glycoproteins gC, gD, and gE protects mice against HSV-1 genital lesions and latent infection.

HSV-1 causes 50% of first-time genital herpes infections in resource-rich countries and affects 190 million people worldwide. A prophylactic herpes vaccine is needed to protect against genital infections by both HSV-1 and HSV-2. Previously our laboratory developed a trivalent vaccine that targets glycoproteins C, D, and E present on the HSV-2 virion.

Herpes simplex virus type 2 trivalent protein vaccine containing glycoproteins C, D and E protects guinea pigs against HSV-1 genital infection.

A vaccine to prevent genital herpes is an unmet public health need. We previously reported that a trivalent vaccine containing herpes simplex virus type 2 (HSV-2) glycoproteins C, D, and E (gC2, gD2, gE2) produced in baculovirus and administered with CpG/alum as adjuvants blocks immune evasion mediated by gC2 and gE2 and virus entry by gD2. The vaccine protected guinea pigs against HSV-2 vaginal infection.

Vaccines to prevent genital herpes.

Genital herpes increases the risk of acquiring and transmitting Human Immunodeficiency Virus (HIV), is a source of anxiety for many about transmitting infection to intimate partners, and is life-threatening to newborns. A vaccine that prevents genital herpes infection is a high public health priority. An ideal vaccine will prevent both genital lesions and asymptomatic subclinical infection to reduce the risk of inadvertent transmission to partners, will be effective against genital herpes caused by herpes simplex virus types 1 and 2 (HSV-1, HSV-2), and will protect against neonatal herpes.

Nucleoside-modified mRNA encoding HSV-2 glycoproteins C, D, and E prevents clinical and subclinical genital herpes.

The goals of a genital herpes vaccine are to prevent painful genital lesions and reduce or eliminate subclinical infection that risks transmission to partners and newborns. We evaluated a trivalent glycoprotein vaccine containing herpes simplex virus type 2 (HSV-2) entry molecule glycoprotein D (gD2) and two immune evasion molecules: glycoprotein C (gC2), which binds complement C3b, and glycoprotein E (gE2), which blocks immunoglobulin G (IgG) Fc activities. The trivalent vaccine was administered as baculovirus proteins with CpG and alum, or the identical amino acids were expressed using nucleoside-modified mRNA in lipid nanoparticles (LNPs).

Antibody responses to crucial functional epitopes as a novel approach to assess immunogenicity of vaccine adjuvants.

We are interested in developing a vaccine that prevents genital herpes. Adjuvants have a major impact on vaccine immunogenicity. We compared two adjuvants, an experimental Merck Sharp & Dohme lipid nanoparticle (LNP) adjuvant, LNP-2, with CpG oligonucleotide combined with alum for immunogenicity in mice when administered with herpes simplex virus type 2 (HSV-2) glycoproteins C, D and E (gC2, gD2, gE2).

A trivalent gC2/gD2/gE2 vaccine for herpes simplex virus generates antibody responses that block immune evasion domains on gC2 better than natural infection.

Vaccines for prevention and treatment of genital herpes are high public health priorities. Our approach towards vaccine development is to focus on blocking virus entry mediated by herpes simplex virus type 2 (HSV-2) glycoprotein D (gD2) and to prevent the virus from evading complement and antibody attack by blocking the immune evasion domains on HSV-2 glycoproteins C (gC2) and E (gE2), respectively. HSV-2 gC2 and gE2 are expressed on the virion envelope and infected cell surface where they are potential targets of antibodies that bind and block their immune evasion activities.

The Revised 2017 MSPE: Better, But Not "Outstanding".

Aim: The medical student performance evaluation (MSPE) is relied on as an objective summary evaluation by surgical program directors. In 2017, an MSPE task force released recommendations for best practice for their format and content. The purpose of this study was to analyze US medical schools' adherence to these guidelines.

Improving Information Transfer during Transitions of Care Standardized Handoffs.

Multiple studies have shown the detrimental effect of miscommunication during transitions of care. The aim of this study is to determine whether a certain method of "sign-out" can improve information transmission and thereby reduce medical errors. Surgical interns underwent a 90-minute training session before starting residency in five previously verified methods of sign-out.

Vaccine-induced antibodies to herpes simplex virus glycoprotein D epitopes involved in virus entry and cell-to-cell spread correlate with protection against genital disease in guinea pigs.

Herpes simplex virus type 2 (HSV-2) glycoprotein D (gD2) subunit antigen is included in many preclinical candidate vaccines. The rationale for including gD2 is to produce antibodies that block crucial gD2 epitopes involved in virus entry and cell-to-cell spread. HSV-2 gD2 was the only antigen in the Herpevac Trial for Women that protected against HSV-1 genital infection but not HSV-2.

A trivalent subunit antigen glycoprotein vaccine as immunotherapy for genital herpes in the guinea pig genital infection model.

An estimated 417 million people worldwide ages 15 to 49 are infected with herpes simplex virus type 2 (HSV-2), the most common cause of genital ulcer disease. Some individuals experience frequent recurrences of genital lesions, while others only have subclinical infection, yet all risk transmitting infection to their intimate partners. A vaccine was developed that prevents shingles, which is a recurrent infection caused by varicella-zoster virus (VZV), a closely related member of the Herpesviridae family.

Human Cytomegalovirus MicroRNAs miR-US5-1 and miR-UL112-3p Block Proinflammatory Cytokine Production in Response to NF-κB-Activating Factors through Direct Downregulation of IKKα and IKKβ.

Emerging evidence indicates that human cytomegalovirus (HCMV) manipulates host cell signaling pathways using both proteins and noncoding RNAs. Several studies have shown that HCMV induces NF-κB signaling early in infection, resulting in the induction of antiviral proinflammatory cytokines with a subsequent reduction of these cytokines late in infection. The mechanism for late cytokine reduction is unknown.

An HSV-2 Trivalent Vaccine Is Immunogenic in Rhesus Macaques and Highly Efficacious in Guinea Pigs.

A genital herpes vaccine is urgently needed to prevent pain and suffering, reduce the incidence of neonatal herpes, and decrease the risk of HIV acquisition and transmission that accompanies genital infection. We evaluated a trivalent HSV-2 subunit antigen vaccine administered with CpG and alum in rhesus macaques and guinea pigs. The vaccine contains glycoproteins C, D and E (gC2, gD2, gE2) to block virus entry by gD2 and immune evasion by gC2 and gE2.

Cytomegalovirus microRNAs.

The discovery that animals, plants and DNA viruses encode microRNAs (miRNAs) has transformed our understanding of the regulation of gene expression. miRNAs are ubiquitous small non-coding RNAs that regulate gene expression post-transcriptionally, generally by binding to sites within the 3' untranslated regions (UTR) of messenger RNA (mRNA) transcripts. To date, over 250 viral miRNAs have been identified primarily in members of the herpesvirus family.

Techniques for characterizing cytomegalovirus-encoded miRNAs.

microRNAs (miRNAs) are small noncoding RNAs that regulate gene expression at the posttranscriptional level, by binding to sites within the 3' untranslated regions of messenger RNA (mRNA) transcripts. The discovery of this completely new mechanism of gene regulation necessitated the development of a variety of techniques to further characterize miRNAs, their expression, and function. In this chapter, we will discuss techniques currently used in the miRNA field to express, detect, and inhibit miRNAs as well as methods used to identify their targets.

Cytomegalovirus miRNAs target secretory pathway genes to facilitate formation of the virion assembly compartment and reduce cytokine secretion.

Herpesviruses, including human cytomegalovirus (HCMV), encode multiple microRNAs (miRNA) whose targets are just being uncovered. Moreover, miRNA function during the virus life cycle is relatively unknown. We find that HCMV miRs UL112-1, US5-1, and US5-2 target multiple components of the host secretory pathway, including VAMP3, RAB5C, RAB11A, SNAP23, and CDC42.

Human cytomegalovirus US7 is regulated synergistically by two virally encoded microRNAs and by two distinct mechanisms.

Human cytomegalovirus (HCMV) encodes at least 14 microRNAs (miRNAs) that act posttranscriptionally to repress gene expression. Although several HCMV miRNA targets of both cellular and viral origin have been identified, our knowledge of their function remains limited. HCMV miRNA targets, as well as phenotypes associated with HCMV miRNA mutants, have been difficult to identify since the downregulation of targets by a single miRNA is often less than 2-fold.

A viral microRNA down-regulates multiple cell cycle genes through mRNA 5'UTRs.

Global gene expression data combined with bioinformatic analysis provides strong evidence that mammalian miRNAs mediate repression of gene expression primarily through binding sites within the 3' untranslated region (UTR). Using RNA induced silencing complex immunoprecipitation (RISC-IP) techniques we have identified multiple cellular targets for a human cytomegalovirus (HCMV) miRNA, miR-US25-1. Strikingly, this miRNA binds target sites primarily within 5'UTRs, mediating significant reduction in gene expression.

Blocking antibody access to neutralizing domains on glycoproteins involved in entry as a novel mechanism of immune evasion by herpes simplex virus type 1 glycoproteins C and E.

Herpes simplex virus type 1 (HSV-1) glycoprotein C (gC) blocks complement activation, and glycoprotein E (gE) interferes with IgG Fc-mediated activities. While evaluating gC- and gE-mediated immune evasion in human immunodeficiency virus (HIV)-HSV-1-coinfected subjects, we noted that antibody alone was more effective at neutralizing a strain with mutations in gC and gE (gC/gE) than a wild-type (WT) virus. This result was unexpected since gC and gE are postulated to interfere with complement-mediated neutralization.

The functions of herpesvirus-encoded microRNAs.

Bioinformatic and direct cloning approaches have led to the identification of over 100 novel miRNAs expressed in DNA viruses, although the function of the majority of these small regulatory RNA molecules is unclear. Recently, a number of reports have now identified potential targets of viral miRNAs, including cellular and viral genes as well as an ortholog of an important immuno-regulatory cellular miRNA. In this review, we will cover the identification and characterization of miRNAs expressed in the herpesvirus family and discuss the potential significance of their role in viral infection.