Transcriptome changes during peanut oral immunotherapy and omalizumab treatment

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Abstract

To the Editor, Peanut allergy is a common food allergy and the main cause of anaphylaxis among children.1 In recent years, oral immunotherapy has emerged as a promising treatment for children with different IgE-mediated food allergies, although safety issues must be considered.2 The main aim of immunotherapy is to induce tolerance or desensitization to an allergen, which otherwise causes an allergic reaction. For oral immunotherapy, this means ingesting the allergen in a controlled manner with gradually increasing dosages. Specifically, peanut oral immunotherapy (pOIT) is able to induce tolerance/desensitization.3 While the pathogenesis of food allergy is relatively well-studied,4 mechanisms of OIT-induced tolerance are not well understood. Omalizumab (anti-IgE), which is used as treatment for severe allergic asthma and other IgE-driven allergies, can also facilitate OIT initiation5; however, little is known about the involved mechanisms, including possible changes at the transcriptional level. We therefore investigated transcriptional changes in whole blood using RNA-sequencing profiles during omalizumab treatment and pOIT in participants from the FASTX (Food Allergen Suppression Therapy with Xolair®) study previously described.5, 6 17 peanut allergic adolescents (age 12–18 years) were included in this study (Table S1). Participants were sampled at treatment start (Baseline) and were then treated with omalizumab at recommended dosages for allergic asthma for an initial 8-week period and for some participants additional periods (total of 16 or 24 weeks) until basophil activation test (BAT/CD-sens)7 analysis showed suppressed reactivity to peanut. All participants then underwent an open peanut challenge (pOIT start), before starting peanut OIT (pOIT) in combination with continued omalizumab treatment. The peanut dose was gradually increased until reaching a maintenance dose. Guided by a BAT/CD-sens after 8 weeks on the maintenance dose, participants decreased the omalizumab dose by 50% (Maintenance) and continued to decrease the dose if pOIT was tolerated. Eleven participants were able to tolerate pOIT without omalizumab for >8 weeks and then passed an open peanut food challenge (Final); six participants could not discontinue omalizumab; however, blood samples were obtained after 2–3 years on omalizumab (Final); six participants dropped out of the study (Figure S1). RNA-sequencing was performed on whole blood (n = 17) at Baseline, pOIT start, Maintenance and Final timepoints using the NovaSeq 6000 platform. DESeq2 was used for differential expression analysis of the omalizumab effect and a linear mixed-effect model for analyses during pOIT in combination with omalizumab (pOIT + O) after adjustment for treatment outcome and cell types estimates via deconvolution (Figure S2). For further details on the treatment protocol, methods and statistical analysis, see the Online Appendix and Table S1. For study participant characteristics at Baseline, see Table S1. To elucidate whether omalizumab treatment alone induced alterations in peripheral blood gene expression, we investigated the two first timepoints, Baseline and pOIT start; however, no significant differences were observed (Figure S3). In the longitudinal analysis (pOIT start to Final), 680 genes associated with pOIT + O at nominal p < .005 (Table S2). The Gene Ontology (GO) biological process of these 680 genes is presented in Figure 1A,B. Upregulation of 337 genes was linked to GO terms ‘protein regulation and modification’, while ‘neutrophil degranulation, immune response, phagocytosis and metabolic process’ were among the top terms for the 343 downregulated genes. Out of the 680 genes, 16 were differentially expressed at false discovery rate (FDR) adjusted p < .05 (Table 1, Figure S4). The three genes with the largest negative and positive coefficients, respectively, are displayed in Figure 1C,D; downregulation of ASGR2, GPBAR1 and HM13, and upregulation of USP44, ICOS and CDKN2AIP. Finally, we evaluated the overlap between our 680 pOIT + O-associated genes and peripheral blood gene expression changes associated with acute peanut allergic reactions in a recently published clinical study by Watson et al using the same p-value cut-off (p < .005).8 Out of our 680 significant genes, 108 genes overlapped with differentially expressed genes found by Watson et al,8 mostly with opposite direction, Penrichment = 0.0095 (Figure 2). Our results demonstrate that omalizumab treatment alone does not induce alterations in whole blood gene expression in patients with severe food allergy. This is not surprising given that these patients were unexposed to peanut allergen at the time of Baseline blood sampling, and any concomitant asthma, rhinitis or eczema were well controlled. However, since omalizumab can facilitate OIT, this drug likely allows for biological changes related to allergen tolerance during OIT. Our longitudinal analysis during pOIT + O identified up- and downregulation of several immune-related genes. CD278/ICOS (Inducible T-cell costimulatory) is expressed on activated T cells and appears to play a role in directing effector T-cell differentiation and responses during inflammatory conditions.9 ICOS expression on T regulatory cells and T follicular helper cells may be involved in the allergic disease mechanism.10 In the pathway analyses, we observed significant enrichment for several GO biological process terms related to T-cell function and immune responses. Notably, we have previously described alterations in T-cell polyclonal in vitro activation during pOIT + O in the FASTX study,11 and our cell fraction analyses also identified immunological changes (Figure S2). Comparing our findings with data described by Watson et al8 suggests that pOIT + O may alter the expression level of genes that were found affected during an acute peanut allergic reaction. The main limitations of this study are small sample size and lack of control subjects without omalizumab treatment, which may mask changes only associated with pOIT. Further, whole blood contains a highly heterogeneous cell population, which can only be partly accounted for by estimated cell fractions and our analysis may have failed to detect important treatment-related changes connected specifically to peanut antigen-specific cells. In conclusion, omalizumab treatment alone does not alter the transcriptional signature in peripheral blood of peanut allergic patients, but during pOIT + O, several immune-related signatures were observed. These results may provide insights into mechanisms of allergen tolerance. We thank all FASTX patients for their participation and the research staff at Forskningscentrum, Södersjukhuset. None. Sophia Björkander: Conceptualization (equal); data curation (equal); investigation (equal); methodology (equal); visualization (lead); writing–original draft (equal); writing–review and editing (equal). Simon Kebede Merid: Conceptualization (equal); data curation (equal); formal analysis (lead); investigation (equal); methodology (lead); software (lead); visualization (equal); writing–original draft (equal); writing–review and editing (equal). David Brodin: Conceptualization (equal); data curation (equal); methodology (equal); software (equal); writing–review and editing (equal). Josef Brandström: Conceptualization (equal); data curation (equal); investigation (equal); writing–review and editing (equal). Fredrik Fagerström-Billai: Conceptualization (equal); data curation (equal); formal analysis (equal); methodology (equal); software (equal); writing–review and editing (equal). Marieke van der Heiden: Conceptualization (equal); methodology (equal); writing–review and editing (equal). Jon R. Konradsen: Conceptualization (equal); data curation (equal); writing–review and editing (equal). Michael Kabesch: Conceptualization (equal); funding acquisition (equal); writing–review and editing (equal). Cornelis M. van Drunen: Conceptualization (equal); funding acquisition (equal); writing–review and editing (equal). Korneliusz Golebski: Conceptualization (equal); writing–review and editing (equal). Anke H. Maitland-van der Zee: Conceptualization (equal); funding acquisition (equal); writing–review and editing (equal). Uroš Potočnik: Conceptualization (equal); funding acquisition (equal); writing–review and editing (equal). Susanne J. H. Vijverberg: Conceptualization (equal); funding acquisition (equal); writing–review and editing (equal). Anna Nopp: Conceptualization (equal); data curation (equal); investigation (equal); resources (equal); writing–review and editing (equal). Caroline Nilsson: Conceptualization (equal); funding acquisition (equal); investigation (equal); resources (equal); writing–review and editing (equal). Erik Melén: Conceptualization (equal); data curation (equal); funding acquisition (equal); investigation (equal); methodology (equal); project administration (equal); supervision (lead); writing–original draft (equal); writing–review and editing (equal). The study was approved by the Ethics Committee in Stockholm: 2013/827-31/3, 2014/1980-32, 2016/1390-32, 2020–00807 and the Swedish Drug Agency: 5.1–2013–46183; the trial is registered at EudraCT: 2012–005625–78, ClinicalTrails.gov; NCT02402231. Patients and caregivers provided written informed consent. The peer review history for this article is available at https://publons.com/publon/10.1111/pai.13682. The peer review history for this article is available at https://publons.com/publon/10.1111/pai.13682. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

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