The Respiratory Viruses Core (RVC) within the Molecular Immunoengineering Section (MIS) at the Vaccine Research Center is focused on the development and testing of countermeasures for respiratory diseases caused by RNA viruses. Activities in the RVC range from antigen design and preclinical immunogenicity testing to exploratory studies using samples from human clinical trials. We have a long-standing interest in respiratory syncytial virus (RSV), which can cause serious illness, particularly for infants and older adults, and have evaluated immune responses to the prefusion F subunit vaccine, DS-Cav1, in a phase I clinical trial. Clinical samples were assessed for neutralizing activity against RSV, and B cell and T cell responses to vaccination were also determined. We continue to explore adaptive immunity to RSV infection and vaccination in ongoing research projects.

Our work also includes a variety of resurging and emerging viral pathogens, including paramyxoviruses, Zika, SARS-CoV-2, and enterovirus D68 (EV-D68). This often involves assay and model development for proof-of-concept immunogenicity and protection studies. We perform a variety of serological assays and use tools such as multiparameter flow cytometry to evaluate cellular responses and for monoclonal antibody discovery. RVC efforts are closely integrated with that of the MIS, and often support VRC-wide vaccine development efforts.

The Molecular Immunoengineering Section (MIS) at the Vaccine Research Center aims to conceive novel vaccine concepts that elicit broad and potent protective immune responses against influenza virus and provide a mechanistic principle for designing vaccines for other hypervariable pathogens such as coronaviruses and HIV-1. MIS is dedicated to advancing vaccine immunogen design beyond structure-based protein engineering by combining concepts and principles from multiple disciplines, including, but not limited to, immunobiology, biochemistry, biophysics, nanotechnology, and computational biology. The mission of the MIS is to define the fundamental rules behind vaccine-elicited immunity. Our interests span from basic immunology and virology to translational sciences with the goal of advancing vaccine concepts that would radically improve immune responses to vaccines. Our work involves various biochemical, biophysical, structural, immunological, and computational techniques and tools. MIS efforts include development of “supraseasonal” and “pre-pandemic” influenza vaccine candidates, “mosaic” antigen display technology, high-throughput high-definition virus neutralization assay systems, and animal models that recapitulate key aspects of human responses to influenza, such as immunological imprinting, preexisting immunity, antibody specificity and repertoire, immunodominance, and pathogenesis. MIS is also integrated with the VRC’s Influenza Program by serving as the lead of the Vaccine Concepts team. The Influenza Program involves multiple VRC Sections and Programs with the goal of advancing candidate vaccines from bench to clinic.

We apply systems immunology approaches to investigate human cutaneous leishmaniasis, a disfiguring chronic skin disease caused by the protozoan parasite Leishmania.

Cutaneous leishmaniasis (CL) is endemic and emerging in the southwestern United States and often also affects military personnel who have been deployed to Leishmania-endemic areas. Leishmania parasites are transmitted to humans via the bite of an infected sand fly. As the disease progresses, the immune response to the parasite in the skin can trigger pathologic inflammation that leads to chronic ulceration and permanent scarring. Current therapies for CL are variably effective with increasing reports of drug resistance. A major roadblock to the development of novel therapeutics for CL is our incomplete understanding of human immunity to Leishmania in the skin.

Our group seeks to define the cellular circuits that drive pathologic inflammation and damage to skin tissue in CL. To do this, we employ a variety of experimental and bioinformatic approaches, including single-cell transcriptomics, multiomic spatial profiling, and mechanistic multiscale tissue modeling to map the cell subsets and intercellular signals that drive CL immunopathology. Current efforts are focused on constructing a comprehensive single-cell and spatial atlas of human CL. This work is being performed in collaboration with the NIAID Leishmaniasis Clinic (Principal Investigator: Dr. Elise O’Connell, Laboratory of Parasitic Diseases). Our goal is to develop novel host-directed therapies for CL patients that 1) inhibit the critical signaling circuits driving pathologic inflammation in the skin and 2) enhance immune control of the parasite.

Another major focus of our group is investigating human immunity to vector bites and vector-borne pathogens via controlled human vector challenge studies. The immune response to vector bites profoundly affects the susceptibility of the host to vector-borne pathogens. In collaboration with Dr. Matthew Memoli (Laboratory of Infectious Diseases), we performed a human challenge study using mosquitos and sand flies, which revealed a conserved transcriptomic skin response against highly divergent vector species. We are now using single-cell and spatial profiling to map the cellular circuitry of the human skin response to vector bites. Our goal is to develop novel clinical therapies that co-opt these responses to enhance protection against vector-borne pathogens.

Dr. Carroll’s primary research is concerned with understanding how neuroinflammation and glial cell activation (astrocytes and microglia) influence prion pathogenesis and neurodegeneration. Prions are infectious misfolded conformers of the cellular GPI-anchored protein PrP and can spread from cell to cell within the brain by seeded-polymerization. This group of proteinopathies can affect humans, cattle, sheep, and cervids and are resistant to many standard decontamination methods. Initially, it was assumed that prion diseases lack an immunological component due to the absence of a prominent antibody or interferon response. However, Dr. Carroll’s research has shown that prion infection elicits a substantial inflammatory response in the CNS and that many inflammatory effectors increase in expression in response to prion infection.

To address the potential impact of microglia in prion disease, Dr. Carroll performed several studies using the potent CSF-R1 inhibitor PLX5622 to reduce microglia in the CNS. These studies indicated that microglia were indispensable to host defense against prion disease. Moreover, his research implicated astrocytes as potentially affecting pathology during disease, where when microglia were absent, the astrocytes were more highly active, expressing numerous disease-related components. This has led to further investigations to assess astrogliosis during prion infection.

A Uniform Manifold Approximation and Projection (UMAP) of single nuclei RNA (snRNA) sequencing analysis of uninfected and prion-infected mouse thalamus depicting 43 transcriptional clusters from >69,000 nuclei examined.

Credit: NIAID

Using high-throughput deep sequencing of RNA transcripts in longitudinal studies, Dr. Carroll has identified numerous differentially expressed genes in the CNS during prion infection. These investigations have yielded compelling results and suggest that microglia in the prion-infected brain assume an alternative phenotype that is distinct from those seen in other brain disorders. From these RNA-seq studies, it was determined that reactive astrocytes assume an expression signature that is not reliant on the canonical signals described in other neuroinflammatory models. Furthermore, this prion-specific reactive astrocyte expression signature is exacerbated when microglia are reduced in the CNS.

Dr. Carroll has begun to analyze the individual cellular changes in the brain using single nuclei RNA (snRNA) sequencing to better understand the relevant changes in the cell populations in the complex milieu of the CNS during infection. Thus far, the research has focused on gene expression changes in the thalamus at pre- and early-clinical times. The thalamus is affected early during prion infection in rodent models, and thalamic pathology is a key feature in human forms of prion disease.

A new aspect of Dr. Carroll’s research is a collaboration with Dr. Cathryn Haigh (Chief, Prion Cell Biology Unit, NIAID) to study Lyme Neuroborreliosis (LNB). Lyme disease, a global public health concern, is the most common tick-borne disease in North America and Eurasia, with an estimated 14% of the world’s population having become infected. Reported cases of Lyme disease in the U.S. have been on the rise for many years, with over 62,000 confirmed cases reported in 2022, making it the leading reportable arthropod-borne infectious disease. The Centers for Disease Control estimates that the disease is underreported, and the true incidence of Lyme disease in the U.S. is approaching 500,000 cases annually. Lyme disease, caused by bacterial spirochetes of the genus Borrelia, is a multi-systemic disorder affecting the skin, heart, central nervous system, and joints.

To address the need for additional models of LNB and to better understand the responses of the human CNS when exposed to Borrelia, this collaboration has developed two in vitro model systems. The first uses human cerebral organoids differentiated from human induced pluripotent stem cells (iPSCs) as an in vitro tissue model. The second uses iPSCs for differentiation into specific neuronal subtypes, astrocytes, and microglia-like cells for study. Exploiting these human-derived model systems, we are assessing responses to Borrelia infection that are stimulated in isolated cells from specific responses that only occur in these cells when they are part of an integrated organoid network. This project incorporates several cutting-edge technologies, including organoid development, bulk and single-cell RNA sequencing, metabolomics, and lipidomics.

Experimental design and strategy to address potential responses of human cerebral organoids, astrocytes, and neurons after exposure to infectious Borrelia species that cause Lyme Neuroborreliosis.

Credit: NIAID