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Jun 29, 2021 / Immunology

Probing deeper: Single cell tools refine our understanding of the role of innate immune cells in successful allergen immunotherapy

Jeanene Swanson

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Allergen immunotherapy is a way to treat allergies by exposing people to allergens in increasingly larger doses. If successful, this desensitization process changes the immune system’s response to the allergen, and it produces fewer antibodies and more immune cells, including T cells, B cells, and cells that produce the regulatory cytokine interleukin-10 (IL-10). In a recent clinical study (1), scientists at the University of Amsterdam and Imperial College London made use of single cell tools to study the role that IL-10-producing type 2 innate lymphoid cells play in grass pollen allergy.

Allergen immunotherapy is a type of treatment for seasonal allergic rhinitis (more commonly known as hay fever), a widespread disease of the upper airway that’s caused by allergens like grass pollen. When allergen immunotherapy is given to a person with an allergy, typically subcutaneously or sublingually, it causes a decrease in the body’s immune response, ultimately leading to immune tolerance of the allergen. The therapy works by affecting the immune system in different ways, including suppressing or inducing different types of T cells or by inducing the production of regulatory B cells that secrete IL-10 (2, 3).

Innate lymphoid cells (ILCs) are innate immune cells that live in tissue and regulate the body’s immunity by secreting signaling molecules in response to pathogens. When these cells’ behavior goes awry, this can lead to allergies, asthma, and autoimmune disorders. Recently, several groups discovered a subset of ILCs, type 2 ILCs (ILC2s), that are also able to secrete IL-10 (4, 5). When it comes to allergen immunotherapy, understanding what role these IL-10-producing ILCs play in the clinical response to treatment is key to improving the therapy’s efficacy. In a recent study, Korneliusz Golebski, PhD, at the University of Amsterdam, led a team that used single cell tools to characterize this cell type and describe how it affects the response to grass pollen allergen immunotherapy in a clinical patient cohort.

Deep dive: Toward characterizing IL-10-producing type 2 innate lymphoid cells

In the first part of the study, the team performed a series of in vitro experiments aimed at characterizing ILC precursors, as well as defining the effect of IL-10-producing ILC2s on T-cell immune response and nasal epithelial integrity. Taking sorted ILCs from peripheral blood mononuclear cells of patients enrolled in a prospective, double-blind, placebo-controlled clinical trial comparing allergen immunotherapy to placebo (6), they were able to generate and study which factors contribute to the activation of IL-10-producing ILC2s. Cell culture experiments showed that only ILC2 precursors and mature ILC2s that express KLRG1 can be induced to produce IL-10 upon activation with IL-33 and retinoic acid (RA).

When IL-10⁺ ILC2s were cultured with CD4⁺ T cells, IL-10-producing ILC2s blocked all T-cell responses, including those of Th2, Th1, and Th17 cells.

Finally, when they cultured nasal epithelium cells with ILCs previously polarized to secrete IL-10, followed by exposure to grass pollen allergen, they saw that IL-10-producing ILC2s prevented the barrier from disintegrating or helped restore the epithelial integrity. Further culture studies showed that this works by affecting expression of genes involved in the inflammatory response.

Different transcriptomic profiles of ILCs of grass pollen allergics and controls

To study possible gene expression differences in ILCs of people with grass pollen allergy, they performed an ex vivo cross-sectional study, taking blood samples from both allergic and control subjects and performing single cell Cellular Indexing of Transcriptomes and Epitopes by Sequencing (scCITE-seq) to profile 1,994 and 2,044 ILCs in grass pollen allergics (GPAs) and control patients, respectively. They found a total of 2,345 genes were differentially expressed and that, overall, many were related to immune functions, such as upregulation of pro-inflammatory chemokines and cytokine receptors. Notably, genes involved in RA synthesis were expressed more in controls compared to allergics.

Allergic people have low numbers of IL-10-producing ILC2s

In the in vitro part of the same cross-sectional study, Dr. Golebski and team aimed to learn whether allergic people and controls had the same capability to produce IL-10-secreting ILCs. After taking blood from GPAs, house dust mite allergics, and controls, and then stimulating the ILCs with IL-2, IL-7, IL-33, and RA, they found that the proportion of IL-10⁺ ILC2s and the amounts of secreted IL-10 were significantly lower in both allergic subsets versus controls.

Sublingual allergen immunotherapy induces IL-10⁺ ILC2s

They wondered if allergen immunotherapy could restore these missing ILC2 cells. Using a combination of in vitro assays and scCITE-seq to analyze the clinical trial data testing grass pollen subcutaneous immunotherapy against a placebo, they found that allergic patients receiving the therapy had a significantly higher proportion of IL-10⁺ ILC2s and amount of secreted IL-10 after treatment, while untreated grass pollen allergics had no changes. Importantly, after 12 months of treatment, patients’ symptoms decreased in proportion to the increase of ILCs and secreted IL-10.

Transcriptomic profiling revealed that the genes found to be differentially expressed between the two patient groups are associated with retinol metabolism, cytokine–cytokine receptor interaction, and the JAK-STAT signaling pathway. Their findings suggest that this type of immunotherapy changes the transcriptional program in ILCs to both increase production of IL-10 and increase activity of genes that help regulate retinol metabolism.

Predicting efficacy and response with a new biomarker

Allergen immunotherapy is an effective treatment for allergic rhinitis, but the underlying mechanisms behind the resulting immune tolerance are not fully understood. While increased amount of IL-10 following immunotherapy has mainly been attributed to the adaptive immune response, particularly Treg and Breg cells, this intriguing study lends evidence that ILC2 cells can also produce IL-10 as part of a successful response to immunotherapy. Future studies can leverage the restoration of IL-10-producing ILC2 cells as biomarkers of allergen immunotherapy success, including efficacy and patient response.

References:

1. Golebski K, et al. Induction of IL-10-producing type 2 innate lymphoid cells by allergen immunotherapy is associated with clinical response. Immunity 54(2): 291–307.e7, 2021.
2. Rosser EC & Mauri C. Regulatory B cells: origin, phenotype, and function. Immunity 42: 607–612, 2015.
3. van de Veen W, et al. IgG4 production is confined to human IL-10-producing regulatory B cells that suppress antigen-specific immune responses. J Allergy Clin Immunol 131: 1204–1212, 2013.
4. Wang S, et al. Regulatory innate lymphoid cells control innate intestinal inflammation. Cell 171: 201–216, 2017.
5. Seehus CR, et al. Alternative activation generates IL-10 producing type 2 innate lymphoid cells. Nat Commun 8: 1900, 2017.
6. Steveling EH, et al. Protocol for a randomised, double-blind, placebo-controlled study of grass allergen immunotherapy tablet for seasonal allergic rhinitis: time course of nasal, cutaneous and immunological outcomes. Clin Transl Allergy 5: 43, 2015. https://clinicaltrials.gov/ct2/show/NCT02005627