Human Cytomegalovirus Acquisition Research Articles

Vaccination with a replication-defective cytomegalovirus vaccine elicits a glycoprotein B-specific monoclonal antibody repertoire distinct from natural infection

Human Cytomegalovirus (HCMV) is the leading infectious congenital infection globally and the most common viral infection in transplant recipients, therefore identifying a vaccine for HCMV is a top priority. Humoral immunity is a correlate of protection for HCMV infection. The most effective vaccine tested to date, which achieved 50% reduction in acquisition of HCMV, was comprised of the glycoprotein B protein given with an oil-in-water emulsion adjuvant MF59. We characterize gB-specific monoclonal antibodies isolated from individuals vaccinated with a disabled infectious single cycle (DISC) CMV vaccine, V160, and compare these to the gB-specific monoclonal antibody repertoire isolated from naturally-infected individuals. We find that vaccination with V160 resulted in gB-specific antibodies that bound homogenously to gB expressed on the surface of a cell in contrast to antibodies isolated from natural infection which variably bound to cell-associated gB. Vaccination resulted in a similar breadth of gB-specific antibodies, with binding profile to gB genotypes 1–5 comparable to that of natural infection. Few gB-specific neutralizing antibodies were isolated from V160 vaccinees and fewer antibodies had identifiable gB antigenic domain specificity compared to that of naturally-infected individuals. We also show that glycosylation of gB residue N73 may shield binding of gB-specific antibodies.

Immune Correlates of Protection Against Human Cytomegalovirus Acquisition, Replication, and Disease.

Human cytomegalovirus (HCMV) is the most common infectious cause of infant birth defects and an etiology of significant morbidity and mortality in solid organ and hematopoietic stem cell transplant recipients. There is tremendous interest in developing a vaccine or immunotherapeutic to reduce the burden of HCMV-associated disease, yet after nearly a half-century of research and development in this field we remain without such an intervention. Defining immune correlates of protection is a process that enables targeted vaccine/immunotherapeutic discovery and informed evaluation of clinical performance. Outcomes in the HCMV field have previously been measured against a variety of clinical end points, including virus acquisition, systemic replication, and progression to disease. Herein we review immune correlates of protection against each of these end points in turn, showing that control of HCMV likely depends on a combination of innate immune factors, antibodies, and T-cell responses. Furthermore, protective immune responses are heterogeneous, with no single immune parameter predicting protection against all clinical outcomes and stages of HCMV infection. A detailed understanding of protective immune responses for a given clinical end point will inform immunogen selection and guide preclinical and clinical evaluation of vaccines or immunotherapeutics to prevent HCMV-mediated congenital and transplant disease.

2651. Protection Against Human Cytomegalovirus Acquisition Is Associated with IgG Binding to Cell-Associated CMV glycoprotein B in Two Historical gB/MF59 Vaccine Cohorts

BackgroundHuman cytomegalovirus (CMV) is the most common congenital infection worldwide. A CMV glycoprotein B (gB) subunit vaccine with MF59 adjuvant achieved ~50% protection in phase II clinical trials in postpartum and adolescent women. Interestingly, postpartum vaccinees showed poor virus neutralization but robust antibody-dependent cellular phagocytosis (ADCP). In this study, we performed a combined humoral immune correlate of risk analysis in vaccinees to define vaccine-elicited immune responses associated with protection and targets for vaccine candidate immunogenicity.MethodsgB/MF59 vaccinees who became infected and those who remained uninfected were 2:1 matched on race and number of vaccine doses. This study included 42 women from the adolescent (14 infected, 28 uninfected) and 33 from the postpartum cohorts (11 infected, 22 uninfected). IgG binding to whole gB, gB-neutralizing epitopes, FCRs, and whole virions were assessed by standard or multiplex ELISA. IgG binding to gB mRNA-transfected HEK293Ts was measured by flow cytometry. Neutralization of Towne, TB40/E, and AD169-repaired-GFP strains were measured in MRC-5, BJ5Ta, and/or ARPE-19 cells. Phagocytosis was assessed by THP-1 uptake of fluorescently conjugated TB40/E and AD169-repaired-GFP virions. Multiple linear regression controlling for cohort was performed for the combined log-transformed group data (apriori significance cut-off of P < 0.05, Benjamin–Hochberg FDR < 0.2).ResultsVaccine-elicited antibodies in adolescent and postpartum cohorts exhibited similar magnitude IgG binding to soluble HCMV gB protein, gB-neutralizing domains, and gB-transfected cells. Autologous Towne strain neutralization was observed in both cohorts, but heterologous strain neutralization was observed only in adolescent vaccinees (P = 0.001). Both cohorts exhibited robust phagocytosis of HCMV virions. Regression analyses revealed that risk of HCMV acquisition in vaccinees was associated with magnitude IgG binding to gB-transfected cells (P = 0.006, FDR = 0.15), not neutralization or phagocytosis responses.ConclusionProtection against primary HCMV infection was significantly associated with vaccine-elicited IgG binding to gB-transfected cells, suggesting the importance of a native, cell-associated gB conformation in future vaccine candidates.Disclosures All authors: No reported disclosures.

HCMV glycoprotein B subunit vaccine efficacy mediated by nonneutralizing antibody effector functions

Human cytomegalovirus (HCMV) is the most common congenital infection worldwide, frequently causing hearing loss and brain damage in afflicted infants. A vaccine to prevent maternal acquisition of HCMV during pregnancy is necessary to reduce the incidence of infant disease. The glycoprotein B (gB) + MF59 adjuvant subunit vaccine platform is the most successful HCMV vaccine tested to date, demonstrating ∼50% efficacy in preventing HCMV acquisition in multiple phase 2 trials. However, the mechanism of vaccine protection remains unknown. Plasma from 33 postpartum women gB/MF59 vaccinees at peak immunogenicity was tested for gB epitope specificity as well as neutralizing and nonneutralizing anti-HCMV effector functions and compared with an HCMV-seropositive cohort. gB/MF59 vaccination elicited IgG responses with gB-binding magnitude and avidity comparable to natural infection. Additionally, IgG subclass distribution was similar with predominant IgG1 and IgG3 responses induced by gB vaccination and HCMV infection. However, vaccine-elicited antibodies exhibited limited neutralization of the autologous virus, negligible neutralization of multiple heterologous strains, and limited binding responses against gB structural motifs targeted by neutralizing antibodies including AD-1, AD-2, and domain I. Vaccinees had high-magnitude IgG responses against AD-3 linear epitopes, demonstrating immunodominance against this nonneutralizing, cytosolic region. Finally, vaccine-elicited IgG robustly bound membrane-associated gB on the surface of transfected or HCMV-infected cells and mediated virion phagocytosis, although were poor mediators of NK cell activation. Altogether, these data suggest that nonneutralizing antibody functions, including virion phagocytosis, likely played a role in the observed 50% vaccine-mediated protection against HCMV acquisition.

Dynamics of Human Cytomegalovirus Infection in CD34+ Hematopoietic Cells and Derived Langerhans-Type Dendritic Cells.

Acquisition of human cytomegalovirus (CMV) usually occurs by contact between contaminated bodily fluids, such as urine and saliva, and host mucosal cells. Langerhans-type dendritic cells (LC) are the only type of immune cells found in the outermost layers of the oral mucosae, where they not only provide a first line of defense against CMV but can easily be targeted by orally administered vaccines, while their bone marrow resident progenitors are important sites of virus latency. In this work, we tracked the progress of infection in CD34(+) progenitor cells, immature LC (iLC), and mature LC (mLC) exposed to the clinical-like strain TB40-BAC4 or to the vaccine strain AD169varATCC, prior to their long-term maintenance under either immature or mature conditions. We show that the genomes of both strains are efficiently maintained in CD34(+) cells during their differentiation into iLC, although this requires the presence of larger amounts of input AD169varATCC DNA. Lipopolysaccharide- and CD40 ligand-induced maturation of iLC derived from latently infected progenitors was not associated with robust viral genome replication and progeny production, while maturation of directly infected iLC increased and prolonged expression of the viral immediate early proteins. While effective replication of viral genomes from both strains occurred only in mLC, both iLC and mLC produced viral progeny, suggesting that both types of LC may contribute to CMV horizontal transmission in vivo. Human CMV is usually acquired via the oral and nasal mucosae. Langerhans-type dendritic cells (LC) are the only type of immune cells found in the outermost layers of these tissues. Understanding how CMV interacts with LC and their hematopoietic progenitors is thus essential to develop innovative means of defense against this virus. Here we show that the genomes of a virulent and an attenuated strain of CMV are maintained in hematopoietic progenitor cells during their differentiation into immature LC and that maturation of these cells by exposure to lipopolysaccharide and CD40 ligand is not sufficient to trigger virus reactivation. While the extents of viral protein expression and genome replication were broadest in directly infected mature LC populations, similar amounts of viral progeny were detected in the supernatants of immature and mature LC, suggesting that these immune cells of the oral mucosa are likely to be important for CMV transmission within the human population.

Breast milk as a source for acquisition of cytomegalovirus (HCMV) in a premature infant with sepsis syndrome: Detection by real-time PCR

Symptomatic disease due to human cytomegalovirus (HCMV) has been increasingly recognized in low birth weight (LBW) premature infants. Breast milk has been identified as a potential source for these infections. At approximately 60 days of life a LBW, premature infant was diagnosed with HCMV sepsis syndrome, prompting further investigation of the source for HCMV acquisition. The infant had received a combination of both fresh and frozen breast milk from his mother throughout his first 2 months of life. We utilized a real-time PCR assay for HCMV DNA to retrospectively examine viral genome copy number in serial samples of stored, frozen breast milk. This analysis indicated an increase in breast milk viral load over the first 45 days of life, heralding the HCMV sepsis syndrome that was observed clinically. Real-time PCR may be a useful tool in the evaluation of HCMV viral load in breast milk.

Cytomegalovirus and the aging population.

Human cytomegalovirus (HCMV) has the largest genome of any virus known to infect man. The virus has evolved many strategies to manipulate the host immune systems and so can remain latent and evade important immune responses. The human host mounts a substantial immune response against the virus, with up to 1% of the virus-specific CD8+ T cells being directed against specific epitopes. Acquisition of HCMV occurs progressively from an early age, and in developed countries the overall seroprevalence is approximately 60%. In contrast, specific communities such as gay men, lower socioeconomic groups and people residing in developing countries have seroprevalence rates that can exceed 90%. It is a widely held belief that successful control of viral infections decreases with increasing age because of a reduction in the capacity of the immune system. Studies in aging populations have shown a specific expansion of the CD8+, CD28- and CD57+ subset of cells in patients who are HCMV-seropositive. Prior infection with HCMV has also been associated with a significantly increased number of CD4+ and CD8+ lymphocytes, as well as cells expressing CD56 and HLA-DR. Thus, HCMV infection can cause substantial perturbations in T cell subsets and these effects persist in the aging population. In the context of solid organ transplantation, older age of both recipients and donors may serve to increase the frequency of donor-positive recipient-positive (D+R+) transplants, which have only a moderate risk of HCMV disease. In the context of HIV infection, age has been a dominant risk factor for progression to AIDS and death. At present, it does not appear that this can be explained by lack of immune control of HCMV in the aging population, although studies have identified prior HCMV infection as a risk factor for AIDS and death independent of age. We await further investigations to determine whether the immune control of HCMV in the elderly patient is as effective as in the younger adult, and whether this is linked to pathological consequences.