Oral desensitization therapy for peanut allergy induces dynamic changes in peanut-specific immune responses

Abstract

Background: The PALISADE study, an international, phase 3 trial of peanut oral immunotherapy (POIT) with AR101, resulted in desensitization in children and adolescents who were highly allergic to peanut. An improved understanding of the immune mechanism induced in response to food allergen immunotherapy would enable more informed and effective therapeutic strategies. Our main purpose was to examine the immunological changes in blood samples from a subset of peanut-allergic individuals undergoing oral desensitization immunotherapy with AR101.

Methods: Blood samples obtained as part of enrollment screening and at multiple time points during PALISADE study were used to assess basophil and CD4+ T-cell reactivity to peanut.

Results: The absence of clinical reactivity to the entry double-blinded placebo-controlled peanut challenge (DBPCFC) was accompanied by a significantly lower basophil sensitivity and T-cell reactivity to peanut compared with DBPCFC reactors. At baseline, peanut-reactive TH2A cells were observed in many but not all peanut-allergic patients and their level in peripheral blood correlates with T-cell reactivity to peanut and with serum peanut-specific IgE and IgG4 levels. POIT reshaped circulating peanut-reactive T-cell responses in a subset-dependent manner. Changes in basophil and T-cell responses to peanut closely paralleled clinical benefits to AR101 therapy and resemble responses in those with lower clinical sensitivity to peanut. However, no difference in peanut-reactive Treg cell frequency was observed between groups.

Conclusion: Oral desensitization therapy with AR101 leads to decreased basophil sensitivity to peanut and reshapes peanut-reactive T effector cell responses supporting its potential as an immunomodulatory therapy.

Keywords: CD4+ T cells; Th2A cells; basophils; oral immunotherapy; peanut allergy.

Figure 1.. DBPCFC non-reactors have lower basophil sensitivity to Peanut allergen extract.

A. Peanut dose response of basophil activation between DBPCFC non-reactors (black dots; n=10) and reactors (orange dots; n=28) at baseline. B. A Bayesian modelling approach was used to obtain logarithmic fits to the data and determine the concentration at which basophil activation is half of the maximum activation (EC-50) for each patient. C-D. Statistical summary showing BAT EC-50 (C) and CD-max (D) values to peanut at baseline between DBPCFC non-reactors and reactors individuals. E-F. Statistical summary showing BAT EC-50 (E) and CD-max (F) values to peanut based on threshold sensitivity to peanut during baseline DBPCFC in reactor individuals. G-H. Statistical summary showing BAT EC-50 (E) and CD-max (F) values to peanut based on severity of allergic reaction during baseline DBPCFC in reactor individuals. Each dot represents distinct individuals. Differences between groups were analyzed by using the two-sided Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p <0.001.

Figure 2.. Ex vivo peanut-reactive T cell profile between DBPCFC reactor and non-reactors.

A-B. Ex vivo frequencies of circulating pTeff cells (A) and pTreg cells (B) in DBPCFC-reactor (orange box) and non-reactors (black box). C. Percentages of CCR6; CD27 and CRTH2 expression within pTeff cells between DBPCFC reactor (orange dots) and non-reactor individuals (black dots). D. Correlation between CRTH2+; CCR6+ and CD27+ pTeff cells in DBPCFC reactor (orange dots) and non-reactor individuals (black dots). A-D. Each dot represents distinct individuals. Differences between groups were analyzed by using the two-sided Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p <0.001

Figure 3.. Dichotomous pattern of CRTH2+ and CCR6+ pTeff cells in PA individuals.

A. Scatterplot of the average signal of CRTH2+ pTeff cell versus CCR6+ pTeff cell gene expression. Shown are genes whose transcription has been up-regulated (red) or down-regulated (blue) by a factor of 2. B. Heatmap of the top differentially expressed genes between the sorted pTeff cell subset. Data are shown in z score–scaled values. C. Expression levels of transcripts associated with TH2A and TH1/TH17 signaling in CRTH2+ (red) and CCR6+ (blue) pTeff cells. Each dot represents distinct individuals. Differences between groups were analyzed by using the two-sided Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p <0.001

Figure 4.. Level of circulating peanut reactive TH2A cells characterizes peanut allergic subjects with distinct immunological and clinical characteristics.

(A-F) Correlation between proportion of pTeff cell subsets and baseline serum peanut-specific IgE level (A), global pTeff cell frequency (B), serum peanut-specific IgG4 level (C), Skin prick test to peanut (D), severity of allergic reaction during DBPCFC (E) and threshold sensitivity to peanut during DBPCFC (F). (G) Heatmap of indicated baseline clinical and immune parameters by row; and proportion of CRTH2+ pTeff cells in each tested DBPCFC-reactor are indicated across the top. Data are shown in z score–scaled values. A-F. Each dot represents distinct individuals. Differences between groups were analyzed by using the two-sided Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p <0.001.

Figure 5.. POIT decreased peripheral basophil sensitivity to peanut.

A-D. Maximum tolerated dose (A), serum peanut-specific IgE level (B), serum peanut-specific IgG4 level (C) and basophil sensitivity to peanut (D) at screen and exit DBPCFC between active (red dots; n=30) and placebo (blue dots; n=12) arm during PALISADE trial. E. Peanut dose response curves of basophil activation at screen and exit visit between placebo (blue line) and active arm (red line). A Bayesian modelling approach was used to obtain logarithmic fits to the data and determine the concentration at which basophil activation is half of the maximum activation (EC-50) for each patient. F. Basophil sensitivity to peanut between peanut sensitized individuals who did not react to the entry DBPCFC (black dots), POIT-treated participants at exit visit (red dots) and placebo-treated participants at exit (blue dots). Each dot represents distinct individuals. Differences between groups were analyzed by the two-sided Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p <0.001

Figure 6.. POIT reshaped circulating peanut-reactive T cell responses in a subset dependent manner.

A-B. Dynamic of circulating pTeff (A) and pTreg (B) cell frequencies between active and placebo arm during PALISADE trial. C-D. Dynamic of CRTH2+ (red dots) and CCR6+ (blue dots) pTeff cell frequencies (C) and proportion (D) between active and placebo arm during PALISADE trial. E-H. Frequency of pTeff cells (E), proportion of CRTH2+ (F), CCR6+ (G) and CD27+ (H) pTeff cells between peanut sensitized individuals who did not react to the entry DBPCFC (black dots), POIT-treated participants at exit visit (red dots) and placebo-treated participants at exit (blue dots). I-K. Correlation between proportion of CRTH2+ pTeff cells at baseline and ratio Exit:Screen peanut specific IgG4 level (I), ratio Exit:Screen pTeff cell frequency (J), and ratio Exit:Screen proportion of CD27+ pTeff cells (K) between active and placebo arm during PALISADE trial. Each dot represents distinct individuals. Differences between groups were analyzed by the two-sided Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p <0.001.