Infrastructure and Opportunity Fund (IOF) Final Progress Reports
2020
Awardee: Stylianos Bournazos and David Knorr
Title: Modulating the antibody response to vaccination through targeting the CD40 axis
Summary: Our team has been working on developing new ways to enhance the response to vaccinations using therapeutic antibodies targeting the immune system. We're studying the natural antibody response in humans with the goal of finding ways to improve annual vaccination strategies like influenza or COVID. We're specifically focusing on the structure of a part of the antibody called the Fc fragment, which plays a role in how the body fights off pathogens. We're also studying a type of immune cell called T follicular helper cells, which help B cells create long-lived immunity to pathogens. We've tested different immunotherapies, including blocking a protein called PD-1, to see if they can improve the response to vaccinations. More recently, our results in mice and humans suggest that activating a different protein called CD40 may be a more effective way to enhance the response to vaccinations. These studies were aimed at testing the impact of antibodies targeting the CD40 pathway (either enhancing or blocking) on human responses to vaccination.
Awardee: Stephanie Boisson-Dupuis
Title: Inherited human PD-1 deficiency
Summary: The pathogenesis of adverse reactions to PD-1 blockade immunotherapy, including tuberculosis (TB) and autoimmunity remains poorly characterized. We identified and studied a patient with inherited PD-1 deficiency who suffered from TB and died of pulmonary autoimmunity. The patient’s leukocytes lacked expression of PD-1 and did not respond to PD-1-mediated suppression. A variety of inborn errors of interferon (IFN)-γ immunity underlie TB. Intriguingly, the patient’s lymphocytes produced only trace amounts of IFN-γ in response to mycobacterial stimuli, owing to the depletion and dysfunction of multiple IFN-γ-producing T and NK lymphocyte subsets. Another set of inborn errors triggering an expansion of CD4-CD8- double-negative (DN) αβ T cells underlie lymphoproliferative autoimmunity (e.g. Fas deficiency and STAT3 gain-of-function (GOF)). Remarkably, the patient displayed hepatosplenomegaly and expansion of total, activated, and RORγT+ DN αβ T cells, as observed in PD-1-deficient mice, cancer patients with PD-1 blockade, and, patients with STAT3 GOF mutations. RNA sequencing of sorted DN αβ T cells revealed unique transcriptional signatures common to the PD-1-deficient and STAT3 GOF patients, but not seen in FAS-deficient patients, suggesting that PD-1 deficiency triggers lymphoproliferative autoimmunity that apparently phenocopies that seen in patients with STAT3 GOF. Mechanistically, the patient’s myeloid cells produced excessive amounts of STAT3-activating cytokines IL-6 and IL-23 upon stimulation with lipopolysaccharide, culminating in STAT3-dependent RORγT upregulation in T cells. Overall, inherited human PD-1 deficiency underlies TB through undermining IFN-γ production by T and NK lymphocytes and lymphoproliferative autoimmunity through over-activating STAT3. The manuscript outlining this work has been published in Nature Medicine (Ogishi et al. PMC8446316).
2021
Awardee: Sean Brady
Title: Human-microbial-lectins regulate the mucosal immune system
Summary:
Awardee: Junyue Cao
Title: Investigation of the pathogenesis of Psoriasis through a novel single-cell genomic technique
Summary: Our main goal is to develop state-of-the-art genomic approaches for comprehensively profiling the cell population heterogeneity and the identifying associated molecular features in human psoriasis samples.
During the funding period, we focused on developing two high-throughput single-cell profiling methods to profile gene expression and chromatin accessibility at single-cell resolution. First, we developed a new approach EasySci-RNA that is more cost-effective than alternative scRNA-seq technologies (less than 1/500 the cost of commercial methods). In addition, the new approach can profile single-cell gene expression with full gene body coverage, thus recovering cell-type-specific gene expression at isoform resolution.
Moreover, to increase the throughput of single-nucleus chromatin accessibility profiling, we and others recently developed single-cell ATAC-seq by combinatorial indexing (sci-ATAC-seq) - a platform that uses split-pool barcoding to uniquely label open chromatin of a large number of single nuclei. While the sci-ATAC-seq platform has been validated in the analysis of human tissues, there are several limiting factors, including low detection efficiency and high cell loss rate, that limit its further scaleup to many samples. To overcome these limitations, we optimized this platform to further improve its efficiency by over 10-fold and reduced the cost to less than $1000 per million cells.
Through collaboration with Dr. James Krueger’s lab, during the funding period, we tested our methods in human skin tissues from psoriasis patient biopsy at Rockefeller University Hospital. While our optimized methods work with some human tissues, the profiling of skin samples remains challenging. We then tested these psoriasis samples using a commercial 10X single-cell transcriptome profiling platform and recovered a diverse cell population from the skin samples. A manuscript reporting the cell population heterogeneity of psoriasis skin samples is currently under preparation.
2022
Awardee: Gabriel Victora
Title: Reactivation and secondary hypermutation of memory B in humans following hepatitis B immunization
Summary: The proposed research study aims to investigate the ability of B cells to elicit a potent and protective antibody response upon re-exposure to an antigen. Maturation of antibodies takes place in germinal centers (GCs), clusters of proliferating B cells that form within secondary lymphoid organs upon antigenic challenge. Within these structures, iterative cycles of hypermutation of immunoglobulin genes followed by selection lead to the expansion of higher-affinity B-cell clones. Throughout this process, positively selected B cells are exported as memory B cells that can be utilized upon secondary exposure to differentiate into plasma cells, generating high titers of serum antibodies over a short period of time. This property, known as the "booster effect," is of critical importance for the effectiveness of vaccination and is thought to underlie the need for repeated doses or "booster" shots. A common assumption in the vaccinology field has been that, in addition to forming plasma cells, memory B cells will also generate secondary GCs upon boosting with high efficiency, allowing them to re-evolve their antibody genes to adapt to variant strains of a pathogen. However, the extent to which this sequential affinity maturation of memory B cell occurs is still debated.
This study aims to provide insight into these underlying mechanisms of vaccination by investigating the ability of memory B cells to generate secondary GCs and re-evolve their immunoglobulin genes to adapt to variant strains of a pathogen. To achieve this, we proposed to sample and analyze the primary and secondary immune response by lymph node fine-needle aspiration of hepatitis B vaccine-draining lymph nodes of healthy human volunteers. Secondary vaccination will be administered in the deltoid area contralateral to that of the primary doses to achieve an anatomical separation of the two responses. This will allow us to characterize the cellular and clonal composition of the two responses. A longitudinal sampling of the memory B cell compartment in the peripheral blood will be performed to obtain a complete picture of the clonal dynamics induced by this vaccine.
The study was granted initial approval by the Rockefeller Institution Review Board on January 24, 2022, and after continuing review and protocol amendment finalization, screening of potential participants began in September 2022. As of now, 7 volunteers have been screened and consented to participate. 5 participants were excluded due to evidence of past hepatitis B vaccination. Of the remaining 2 participants, one still has to complete the screening process and one is enrolled in the active phase of the study. The enrolled participant received two primary vaccination doses, and the blood samples were collected pre- and post-vaccination accordingly. The process of prescreening healthy volunteers is ongoing in order to achieve the targeted enrollment of up to 12 evaluable participants. This sample size will allow us to reach the planned goal of 5 participants who received all 3 immunizations and have 2 successful lymph node fine-needle aspiration procedures, which will be necessary to achieve the desired level of understanding of the clonal dynamics induced by the vaccine.
2023
TBD
