IgE and B cell repertoires in human allergy and immunotherapy [electronic resource]

Here you will learn more about IgE and molecular basis of allergic disease. The overarching goal of this work has been to better understand the characteristics of human B cell antibody repertoires, the role they play in the pathology of allergic disease and their contribution in modifying allergic symptoms, and leading to the acquisition of tolerance during allergen specific immunotherapy. In Chapter 2, (previously published as Levin and King, et. al, (2016) in JACI), we examine how IgE repertoire persistence and evolution may hold promise as markers for events accompanying specific immunotherapy of allergic disease. In this chapter, we have combined the use of use phage display technologies to determine IgE specificities, along with high throughput sequencing of blood and tissue B cell repertoires. In this way we were able to identify and track the fates of allergen-specific clones over the course of immunotherapy. We discovered that members of the same allergen-specific B cell clones could be found in both nasal mucosa and the blood, and found evidence of large clonal expansions, persistence, and isotype switching of members of allergen specific IgE containing clonal lineages as immunotherapy progressed. Chapter 3 follows up on this study presented in Chapter 2. In this chapter, we ask what happens in the B cell repertoires of the patients over a longer term of treatment. We look at the fates of allergen specific clones 3 years after the start of immunotherapy, when patients are expected to have reached the optimal improvement of symptoms and are preparing to go off treatment. In our data from this timepoint, we observed a trend toward detection of fewer IgE expressing cells as members of the known allergen specific clones, and more detection of non-IgE expressing clone members, including IgG4, supporting the idea that specific immunotherapy can give rise to IgG4-expressing members of allergen-specific B-cell clones in allergic patients, in a time frame associated with the therapeutic effect of the treatment. In Chapter 4 we discuss more generally the characteristics and differences in B cell repertoires between the blood and nasal biopsy. We found that B cell repertoires in nasal mucosal tissue differed from those observed in the peripheral blood in many ways, which we describe in this chapter. We detected, for instance, distinct antigen experienced IgD and IgM cell populations in tissue, not previously described, and saw a higher proportion of IgG3 usage in nasal tissue, with less IgG2 present. Specific immunotherapy did not appear to elicit global changes in the B cell repertoires in either the nasal tissue or blood. Chapter 5 (previously published as Hoh, et. al, (2016) in JACI), on patients with food allergies and compliments many of our findings from patients with sensitivities to aeroallergens. Investigating allergen-specific B-cells in peanut allergic patients, we found that these cells usually tended to express mutated antibody genes, were typically of switched isotypes, and were often able to bind to both linear and conformational epitopes. Even well-characterized linear epitopes of allergen proteins could be recognized by multiple independent B-cell clones within a single patient. We saw that oral immunotherapy was associated with increased frequencies of allergen-binding B-cells in the blood, and, in one example, progressive somatic mutation of IgG4, but not IgE, members of an allergen-specific clone. Peanut allergen-specific B-cell clones in allergic patient blood typically expressed mutated isotype-switched antibodies, including IgE, and target common epitopes. Oral immunotherapy increased frequencies of specific clones and may also preferentially induce IgG4 somatic mutation. Chapter 6, (previously published as Looney, et. al, (2016) in JACI) discusses methods for generally evaluating somatic mutation patterns in antibody heavy chain genes of B cell clones containing IgE-expressing to search for evidence of direct (from IgM or IgD) versus indirect (from an intermediate switched isotype) isotype switching, a topic that has been a matter of much debate within the field. In this study, we found that both allergic and healthy individuals showed evidence to support the model for indirect switching, particularly from IgG1, as the predominant pathway to IgE expression in humans. Analysis of antibody mutation patterns in allergic and healthy human subjects indicated that most IgE is derived from B cells that previously expressed IgG and had encountered antigen, rather than from naïve IgM+ B cells. Prior to the work presented in this dissertation, studies of B cell repertoires in allergic patients had largely relied on few sequences, or not delved into differences between isotypes. By the use and combination of new technologies and methods, applied to larger cohorts of patients, and across multiple timepoints and/or tissues we have been able to add both depth and breadth to the prior knowledge of human IgE lineages, their origins, and the ways they are being modified over the course of allergen specific immunotherapy. There is still much to learn, but we hope that this work will provide a basis from which to evaluate allergen-specific human antibody repertoires in healthy and diseased individuals, and lay a foundation for future studies aimed at increasing and applying our knowledge for continued improvement in the diagnosis, treatment, and ultimately, the prevention of allergic disease.

Keywords: Biology.
Advisor: Boyd Scott
Fire Andrew Zachary primary advisor. Thesis advisor
Galli Stephen J. advisor. Thesis advisor
Nadeau Kari advisor. Thesis advisor
Stearns Tim advisor. Thesis advisor