Browsing by Subject "Vaccines"

Vaccines have been a revolutionary intervention for infectious diseases over the past many decades. However, over 2 million people continue to die every year of vaccine-preventable causes, especially in low-resource areas. This is mainly due to inefficiencies in the vaccine supply chain which in entirety lead to loss of potency of vaccines worth over $250 million every year due to temperature fluctuations. Additionally, vaccines that actually reach users may lose effectiveness because of more challenges related to their direct delivery to the recipients; for example, contamination and injuries from misuse of needles. We sought to take a step in addressing these challenges by developing thermally stable vaccine tablets for the sublingual delivery of self-assembled peptide nanofibers. Tablets were engineered from a combination of supramolecular peptide nanofibers plus the excipients, dextran and mannitol. The tablet structure was characterized to assess the impact of tablet formation on the nanofiber structure, as well as for suitability of sublingual delivery. In vivo studies were then carried out in a mouse model to determine the capacity to raise antigen-specific immunogenic responses. Sublingual delivery of the tablet in the mouse model was achieved and an immunogenic response was raised in mice. This proof-of-concept study indicates a step towards improving vaccines with regard to addressing challenges of the vaccine supply chain and vaccine delivery, especially in low resource centers.

Human cytomegalovirus (HCMV) is the most common cause of congenital infection worldwide, impacting 1 in 150 live-born infants. Children afflicted by congenital HCMV frequently suffer from lifelong, debilitating neurologic sequelae including microcephaly, sensorineural hearing loss, and cognitive impairment. Natural maternal immunity to HCMV reduces the frequency of congenital infection, but does not prevent the disease altogether. Thus, a vaccine to reduce the incidence and severity of infant infection is a public health priority. Employing a nonhuman primate model of congenital CMV transmission as well as clinical samples from a partially-efficacious HCMV vaccine trial, we sought to examine both the attributes of anti-HCMV immune responses that provide protective immunity as well as the impact of vaccine-elicited immunity on the in vivo HCMV viral population.

First, we used a nonhuman primate model of congenital CMV infection to investigate the ability of preexisting antibodies to protect against placental CMV transmission in the setting of primary maternal infection and subsequent viremia, which is required for placental virus exposure. Pregnant, CD4+ T cell-depleted, rhesus CMV (RhCMV)-seronegative rhesus monkeys were treated with either standardly-produced hyperimmune globulin (HIG) from RhCMV-seropositive macaques or dose-optimized, potently RhCMV-neutralizing HIG prior to intravenous challenge with an RhCMV swarm. HIG passive infusion provided complete protection against fetal loss in both groups. The dose-optimized, RhCMV-neutralizing HIG additionally inhibited placental transmission of RhCMV and reduced viral replication and diversity. Our findings suggest that the presence of durable and potently-neutralizing antibodies at the time of primary infection can prevent transmission of systemically-replicating maternal RhCMV to the developing fetus.

Next, we assessed the properties of antibody responses elicited by glycoprotein B (gB) + MF59 adjuvant subunit vaccination – the most successful HCMV vaccine tested clinicaly to-date, which demonstrated approximately 50% efficacy in preventing HCMV acquisition in multiple phase 2 trials. Plasma from 33 gB/MF59 vaccinees at peak immunogenicity was tested for gB epitope specificity as well as neutralizing and non-neutralizing anti-HCMV effector functions, and compared to 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. Interestingly, vaccinees had high-magnitude IgG responses against AD-3 linear epitopes, demonstrating immunodominance against this non-neutralizing, cytosolic region. Finally, vaccine-elicited IgG robustly bound trimeric, membrane-associated gB on the surface of transfected or HCMV-infected cells and mediated virion phagocytosis, raising the possibility that non-neutralizing antibody effector functions contributed to the partial protection against HCMV acquisition observed in gB/MF59 vaccinees.

Lastly, we evaluated the impact of gB/MF59-elicited immune responses on the population of viruses acquired by trial participants. In this analysis, we employed quantitative PCR as well as two distinct next-generation sequencing strategies (short amplicon and whole gene) to interrogate genetic differences between the HCMV populations infecting gB/MF59 vaccinees and placebo recipients. For the majority of subject-specific viral populations analyzed, we identified 1 or 2 dominant viral variants, as well as a large number of minor variants present at very low frequency. This finding suggests that the intrahost viral population constitutes a heterogeneous swarm of genetically-distinct virus quasi-species. Additionally, we identified several distinctions between the viral populations of acutely-infected vaccinees and placebo recipients. First, there was reduced magnitude viral shedding in the saliva of gB vaccinees compared to placebo. Furthermore, we noted evidence of genetic compartmentalization at the gB locus in 3 of 4 vaccinees, though only in 1 of 7 placebo recipients. Finally, we observed an enrichment of gB1 genotype HCMV variants among placebo recipients compared to vaccinees, and hypothesize that the gB1 genotype vaccine immunogen might have elicited genotype-specific protection that accounts for the efficacy observed in clinical trial.

Thus, we have made several observations that will inform rational design of the next generation of HCMV vaccines. First, our data suggests that preexisting antibodies can protect against congenital CMV transmission in a rhesus monkey model, and thus that antibodies could be a primary target of vaccines to eliminate congenital HCMV infection. Secondly, our analysis of antibody responses elicited by gB/MF59 vaccination indicates that non-neutralizing antibody functions contributed to the observed 50% vaccine protection and therefore should be a consideration in future vaccine design. Finally, our examination of viral populations in gB/MF59 vaccinees indicates that gB-elicited antibodies had a measurable impact on viral intrahost population dynamics and that gB immunogen strain-specific responses may have defined vaccine protection, suggesting that immunogen strain breadth may be an important factor to consider for future vaccine design.

This dissertation examines the colonial history of medical rights in Latin America through a study of the world’s first vaccine. The Spanish introduced the smallpox vaccine to their empire in 1804, along with royal orders that vaccination be voluntary and medical consent a natural right ceded to parents. Yet, the vaccine first arrived there incubated in the bodies of two enslaved girls. Doctors would continue to rely on enslaved, indigenous, and other dispossessed bodies to conserve the vaccine for those otherwise accorded this ostensibly universal right. Their doing so prompted profound questions about individual liberty, embedding vaccination into struggles over the abolition of slavery, parental rights, and the preservation of colonial rule. By analyzing the politicization of preventative health, the dissertation follows the vaccine through the Spanish Caribbean and Mexico to ask why imperial—and later, national—authorities protected voluntary vaccination, what this choice meant for parents and patients, and what this story can tell us about the meaning and value of consent in an era of both race and rights-making.

To understand how consent operated, I trace the vaccine through the bodies that sustained it, examining the gendered and racialized claims to medical authority that legitimized the vaccine, the state’s patriarchal formulation of consent to it, and the responses and rejections of colonized subjects to both. Medical texts, newspapers, legal codes, orphanage records, plantation guides, and government reports related to the vaccine reveal that recognition of medical rights was inconsistent and often determined by elite assumptions about reason and bodily difference. Racial and sexual politics informed decisions about which bodies were best suited to incubate and test the vaccine, whose knowledge was deemed a threat to public health campaigns, and ultimately, who should be recognized as a parent, worthy of rights and capable of informed consent.

Amidst political and social unrest, I argue that these articulations worked to uphold colonial structures of power, as healthcare became woven into the fractional freedoms accorded to and claimed by subjects and citizens. Medical consent, as it was envisioned and employed in vaccination policies, helped to reinforce these hierarchies even after independence. Mexico retained voluntary vaccination, but the medical rights of women and men, particularly those of indigenous and African descent, remained restricted by assumptions about culture and competence. By tracing the vaccine through the postcolonial era, my project addresses the enduring effects of colonialism across political discourses of liberalism and access to resources and care. Such historicization suggests the limits of consent and prompts a more ethical conceptualization of "informed refusal" that embraces and respects indigenous and other cultural articulations of bodily autonomy.

Human cytomegalovirus (CMV) is the most common cause of congenital infection worldwide, affecting approximately 1 in 150 infants, and is a leading cause of morbidity and mortality among transplant recipients. Congenital CMV (cCMV) infection can lead to permanent hearing loss, brain damage, and neurodevelopmental delay, and cCMV alone is responsible for nearly 25% of all infant hearing loss. Chronic CMV infection has also been associated with a heightened inflammatory state and increased risk of aging-related diseases, including as cardiovascular disease and type 2 diabetes. Over the last fifty years, there have been many efforts to develop vaccines that can prevent CMV disease. However, vaccine development for CMV faces many challenges, including a limited understanding of the immune responses protective against infection.

Like other beta herpesviruses, CMV can establish a lifelong, persistent infection in hosts, marked by periods of latency and reactivation, and preexisting immunity does not protect against reinfection. Moreover, CMV is known to predominantly spread by direct cell-to-cell transmission, complicating efforts to design vaccines that not only prevent viral entry via antibody neutralization but also inhibit the spread of cell-associated virus. To identify targets for vaccine development, we investigated the antibody immune responses associated with protection from CMV in historical vaccine trials and the lineage maturation of neutralizing antibodies elicited in natural infection.

The most efficacious CMV vaccine to-date is the glycoprotein B (gB) subunit vaccine combined with the MF59 adjuvant (gB/MF59), which achieved 50% protection against primary CMV acquisition in multiple Phase 2 clinical trials. CMV gB is a viral envelope protein that mediates fusion with host cell membranes and is required for viral entry into all known cell types and for cell-cell spread. Previous studies had found that in cohorts of CMV-seronegative postpartum women and renal transplant recipients, gB/MF59 vaccination did not elicit broadly neutralizing antibodies but instead generated robust nonneutralizing antibody responses, namely antibody-dependent cellular phagocytosis (ADCP). These studies lacked the statistical power to determine whether ADCP responses contributed to the partial efficacy of the gB/MF59 vaccine. We aimed to define the immune responses correlated with protection from primary CMV acquisition in Phase 2 gB/MF59 clinical trials in cohorts of adolescent girls and postpartum women. We first evaluated the vaccine-elicited sera IgG binding, neutralizing, and nonneutralizing responses against CMV, and we observed distinct immunogenicity profiles in the adolescent and postpartum cohorts, wherein adolescent vaccinees but not postpartum vaccinees developed broadly neutralizing CMV antibodies. We then compared the sera antibody responses between vaccinees who acquired infection and those who remained uninfected during the course of the trial. By multiple logistic regression analysis, we found that protection against primary CMV acquisition was associated with the presence of sera IgG binding to cell-associated gB, but not IgG binding to soluble gB as used in the gB/MF59 vaccine. These results suggested that there may be conformational differences between cell-associated and soluble gB. Supporting this, we identified gB-specific monoclonal antibodies (mAbs) that differentially recognized these gB structures. Our findings indicated the importance of the native, cell-associated gB conformation in future CMV vaccine design.

In our immunogenicity studies of gB/MF59, we observed that the vaccine failed to elicit antibodies against the gB antigenic domain 2 site 1 region (AD-2S1), which is a highly conserved, linear epitope at the far N terminus of gB that is known to be a target for potently neutralizing antibodies in natural infection. The presence of sera antibodies against gB AD-2S1 in naturally infected individuals have been associated with decreased risk for cCMV transmission and less severe CMV disease in transplant recipients. Yet, only about half of naturally infected individuals develop anti-gB AD-2S1 antibodies, and gB-based vaccines to-date have failed to elicit these responses. Thus, it remained unclear how to generate neutralizing antibodies against this poorly immunogenic epitope by vaccination. With the goal to identify a gB AD-2S1 structure that could elicit these potently neutralizing antibodies from the germline, we employed a B cell lineage-targeted vaccine strategy. We mapped the phylogeny of a well-characterized, potently neutralizing anti-gB AD-2S1 mAb from its germline precursor, then by empiric testing, we identified the antibody mutations that conferred neutralizing function. We found that a single heavy chain mutation in the CDR1 region was both necessary and sufficient to confer neutralizing function to the otherwise nonneutralizing IGHV3-30/IGKV3-11 germline ancestor mAb. This study identified a critical, early B cell receptor mutation that can serve a target for mutation-guided CMV vaccine design.

Only a limited number of gB AD-2S1 mAb sequences have been published to-date, with few clonally related members. To identify additional antibodies for lineage analysis and for evaluation as potential passive therapeutics, we developed a pipeline to sequence anti-CMV gB AD-2S1 mAbs and estimate their antigen binding in high-throughput. This study leveraged modern advances in B cell sequencing technologies to expedite the discovery of potently neutralizing CMV antibodies.

In sum, this work contributes to our understanding of protective immunity to CMV. We found that the immune correlate of protection for the gB/MF59 vaccine was sera IgG binding to cell-associated gB, suggesting that the next generation of gB-based vaccines should be designed to adopt a cell-associated or native conformation. Then, we investigated the lineage maturation of potently neutralizing antibodies against CMV gB AD-2S1 and identified a critical mAb mutation from the germline associated with the development of neutralization function, which will may serve as target for immunogen design. Additionally, we developed a pipeline for the high-throughput isolation of anti-gB AD-2S1 mAbs. These findings provide insight into antibody-mediated protection from CMV and illuminate paths forward for rational vaccine design.

There are currently over two million Palestinian refugees residing in Jordan, 370,000 of whom reside in refugee camps. Due to conflict-affiliated disease outbreaks among children in the region, the UN Relief and Works Agency for Palestine Refugees (UNRWA) has identified incomplete vaccination as a critical public health issue and has invested in the development and implementation of a text message reminder service for preventing loss-to-follow-up. Childhood immunization rates in UNRWA catchment regions are generally high, yet little is known about risk factors for missed appointments, which impose a substantial administrative burden due to the need to contact patients for rescheduling. Stronger user characterization is necessary for improved targeting and minimized cost as we develop a more robust SMS system capable of scaling across all health facilities.

This mixed-methods study prospectively recorded 6 months of immunization history among a cohort of children born in June 2014 at Taybeh Health Center in Amman. Demographic information was collected at the time of birth, and caregivers of cohort members were invited to participate in interviews that assessed immunization knowledge, preferences, decision-making, and experience with the SMS reminder system. Patients were more likely to significantly delay appointments during the Ramadan holiday and for doses further from the child date of birth. Future policies that might bridge these gaps include targeting pre-appointment SMS reminders to high-risk patients, implementing holiday shifts in clinic hours, and regularly updating patient contact information.

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Human Immunodeficiency Virus (HIV) is a vaccine target that has remained elusive for decades. In 2015, 1.1 million people died of HIV-related causes and 2.1 million new infections occurred. Although an effective HIV vaccine has long been a major goal of the World Health Organization (WHO), HIV has been an extraordinarily challenging vaccine target. This challenge is due to several compounding factors including the evolution of the virus within individuals and across geographic regions. This evolution makes it difficult to develop a universal HIV vaccine that will neutralize all strains of the virus. Furthermore, the virus is exceptionally efficient at mutating to evade the immune systems of infected individuals. The outer surface of the virus is densely glycosylated, making it difficult for the immune system to make effective antibodies against such a shielded structure. Consequently, most antibodies generated against the virus are either non-neutralizing or only partially neutralizing and are ineffective at clearing HIV infection. Thus, alternative strategies to direct the immune system towards making neutralizing antibodies are required for an effective HIV vaccine. Attempts at creating an effective HIV vaccine have centered around stimulating high-affinity antibodies that effectively bind genetically diverse strains of the virus. Through protein engineering, variable dosing regimens, and creation of new antigens, HIV researchers have identified the key factors required for the human immune system to raise functional anti-HIV antibodies. Primarily, the immune system must be directed towards the highly conserved and functional antigenic regions of HIV surface proteins to make protective antibodies. To steer the immune system towards specific, neutralizing epitopes from HIV-1, the repertoire of B cells activated by HIV vaccines must be altered from those elicited by natural infection or traditional immunization approaches. Two ways to alter the B cell populations activated upon immunization are explored in this dissertation. Firstly, lowering the activation threshold of activated B cells by arraying antigens on materials allows for a shift of antibody repertoires toward more epitope specificities and potentially broader binding of antibodies to mutated viral strains. Secondly, the immune system can be focused toward specific epitopes using heterologous immunization regimens where antibodies are selected toward a certain specificity, then evolved to bind native antigens, such as those that would be displayed on HIV virions. Though both of these approaches have been explored in other systems, the unique impact of self-assembling peptides in this space has yet to be explored. Peptide biomaterials with fibrillar morphologies such as β-sheet peptides, worm-like micelles, and peptide amphiphiles have been explored towards numerous biomedical applications including scaffolds for tissue repair, immunotherapies for infectious diseases, cancer, or inflammatory conditions, and depots for sustained drug delivery. Although these materials have shown promise in preclinical applications, the immunological effects of their length and ligand valency are poorly understood. Because both of these features can be utilized to tune immune responses, optimizing them in the context of HIV immunization has the potential to improve the magnitude and quality of antibodies elicited by nanofiber immunogens. To examine the impact of antigen valency and nanofiber size in HIV immunization, structural tools to control these features were developed. The ability to control nanofiber length was achieved in this body of work be engineering a set of peptides we have designed and characterized to stabilize self-assembling interfaces. This newfound control over nanofiber length allows size-based targeting of materials which was not previously possible. In addition to studying size-dependent immunogenicity, the role of glycans in immune responses to nanoscale are a newfound area of interest amongst the biomaterials community. Though some glycans can be readily conjugated to peptide and polymer assemblies, many carbohydrates, such as sialic acids are extremely difficult to functionalize chemically. To overcome this, glycomimetic peptides which resemble diverse glycans have been designed, but are implemented in few therapeutic contexts. This system capitalizes upon the synthetic advantages of utilizing glycomimetic peptides in peptide-based immunogens and represents a broadly applicable strategy to impart lectin-binding properties to peptide materials. Although it is appreciated that multivalency and nanomaterial shape and size can influence immunogenicity, these aspects have yet to be fully exploited in the context of a specific disease. To meet these challenges, we have designed a self-assembling peptide system with control over the lengthwise assembly of nanofibers and have studied the effect of antigen valency on immune responses to these materials in the context of HIV. As immunogens, peptide nanofibers have a unique ability to activate low-affinity B cells, such as those which react to autologous targets, will likely be advantageous for HIV vaccination, where low-affinity B cells are precursors to the induction of broadly neutralizing antibody (bnAb) responses. To determine the utility of peptide nanofibers as platforms for HIV vaccination, We first constructed nanofibers that are covalently linked to the HIV envelope antigen gp120 which demonstrated their ability to raise antibodies with broad binding profiles. As an alternative approach for raising immune responses against HIV antigens, we have explored the use of peptide nanofibers displaying short, linear HIV epitopes as priming immunogens. This approach capitalizes on the ability of nanofibers to generate antibodies against short epitopes and tailors the accumulation of nanofibers in lymph nodes to prime epitope-focused antibodies against HIV virions. Taken together, the studies described here utilize supramolecular control over antigen valency and immunogen size to generate antibody responses to HIV with high affinity and high binding breadth. The supramolecular tools described here provide morphological controls for spontaneously assembling materials which have not yet been utilized for these types of platforms. This tight control over morphology allows us to ask questions with levels of immunological precision that are not common in biomaterials literature.

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Vaccines save millions of lives every year primarily through the induction of antigen-specific antibodies; however, there is a critical lack of understanding of the mechanistic underpinings of vaccine-elicited, antibody-mediated protection. Vaccinologists have long since recognized neutralization as an important antibody function while comparatively neglecting the role of Fc functions of antibodies. Therefore, the goal of my dissertation was to dissect out the functional roles of different subpopulations of antibodies in preventing infection using a controlled human infection model (CHIM) of a pathogen with a mucosal and systemic phase of infection, Salmonella Typhi. The first objective was to determine which isotypes/subclasses within the polyclonal humoral immune response are protective against S. Typhi infection and through what functional mechanisms. The second objective focused on vaccine-derived monoclonal antibodies to identify biophysical properties that affect the ability to recognize S. Typhi Vi antigen and influence downstream antibody Fc-mediated function. Most immune correlates of vaccine-mediated protection focus on either total IgG responses or bulk polyclonal serum with little-to-no defined understanding of how the quality (avidity) and characteristics (epitope-specificity, functional potency) of specific subpopulations impact protection. In this dissertation, I interrogated the humoral response to vaccination with two Vi polysaccharide vaccine constructs: one plain polysaccharide vaccine (Vi-PS) and one Vi-protein conjugate vaccine (Vi-TT). In chapter 2 of this dissertation, I evaluated the binding and avidity of subclass-specific IgA and IgG antibodies to the Vi polysaccharide. Firstly, I demonstrated that Vi-specific IgA magnitude correlated with protection in the typhoid fever CHIM. In addition, Vi IgG1 and IgA2 avidity were higher amongst protected individuals of both vaccine groups. These findings suggest that a combined IgA, perhaps via IgA2, and IgG1 response may be critical in preventing infection. Although IgA magnitude was associated with protection from S. Typhi infection, I did not identify a threshold concentration of Vi-specific IgA that prevented infection. Therefore, I hypothesized that Fc functions, rather than neutralization, may be critical in the mechanism of IgA-mediated protection. Following the findings of the univariate analysis, a multivariate analysis was conducted as part of a wider scientific collaboration in which binding and avidity data, as well as functional data (including FcR binding and Fc-mediated functions), were included to identify the smallest set of immune measurements that could robustly predict protection. In a composite analysis with both vaccine regimens combined, we identified Vi specific IgA magnitude, IgG2 magnitude, and IgA2 avidity as the top features that were enriched among protected individuals. These responses were also linked to antibody-dependent neutrophil phagocytosis and oxidative burst. However, we found enriched antibody-dependent NK cell activation and complement deposition amongst diagnosed vaccinees. These data suggest that a highly specialized mechanism driven by IgA and IgG mediated neutrophil activation via FcαR and FcγRs impart protection whereas broad innate immune activation can be detrimental. In chapter 3 of this dissertation, I interrogated the humoral response to Vi immunization using recombinantly produced monoclonal antibodies derived from vaccinees. I identified a convergent B cell response amongst vaccinees with most antibodies targeting the immunodominant C3 O-acetyl group of the Vi polysaccharide. However, I also identified four unique subdominant epitopes that varied in their accessibility for antibody binding by performing a de-O-acetylation of the polysaccharide backbone and conducting epitope binning competitive assays. Furthermore, I demonstrated that specific subdominant epitopes on the Vi polysaccharide antigen can be effectively targeted by antibody for phagocytosis and complement deposition. I utilized a kinetics rates-based assay, BioLayer Interferometry, to assess the association rate, dissociation rate, and overall avidity of each monoclonal antibody to the Vi antigen. By conducting correlative analyses of kinetic parameters and functional outcomes, I demonstrated that association rate has no substantial association with Fc function, while dissociation rate is highly associated with Fc function. Taken together, these findings highlight a likely mechanism of protection from infection with S. Typhi through cellular phagocytosis mediated cooperatively by IgA and IgG antibodies. This study also highlights the key epitopes that antibodies elicited by intramuscular Vi vaccination target for functional Fc outcomes which is dependent on epitope exposure and on antibody-antigen binding stability (off-rate). Identifying these specific mechanisms of protective immunity against typhoid fever will facilitate evaluation and licensure of the many Vi conjugate vaccines in development that would otherwise require large scale trials, and also help guide any future development of Vi vaccines. More broadly, the analysis here highlights the importance of evaluating specific subpopulations of antibodies, and their quality, as potential mediators of protection for other vaccine constructs rather than focus only on magnitude of bulk polyclonal responses. This type of analysis deeply enhances our understanding of protective humoral immunity while also informing both the development and the evaluation of new vaccine candidates.