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Epitope-specific immunotherapy targeting CD4-positive T cells in coeliac disease: two randomised, double-blind, placebo-controlled phase 1 studies

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Summary

Background

A gluten-free diet is the only means to manage coeliac disease, a permanent immune intolerance to gluten. We developed a therapeutic vaccine, Nexvax2, designed to treat coeliac disease. Nexvax2 is an adjuvant-free mix of three peptides that include immunodominant epitopes for gluten-specific CD4-positive T cells. The vaccine is intended to engage and render gluten-specific CD4-positive T cells unresponsive to further antigenic stimulation. We assessed the safety and pharmacodynamics of the vaccine in patients with coeliac disease on a gluten-free diet.

Methods

We did two randomised, double-blind, placebo-controlled, phase 1 studies at 12 community sites in Australia, New Zealand, and the USA, in HLA-DQ2·5-positive patients aged 18–70 years who had coeliac disease and were on a gluten-free diet. In the screening period for ascending dose cohorts, participants were randomly assigned (1:1) by central randomisation with a simple block method to a double-blind crossover, placebo-controlled oral gluten challenge. Participants with a negative interferon γ release assay to Nexvax2 peptides after the screening oral gluten challenge were discontinued before dosing. For the biopsy cohorts, the screening period included an endoscopy, and participants with duodenal histology who had a Marsh score of greater than 1 were discontinued before dosing. Participants were subsequently randomly assigned to either Nexvax2 or placebo in ascending dose cohorts (2:1) and in biopsy cohorts (1:1) by central randomisation with a simple block method. In the three-dose study, participants received either Nexvax2 60 μg, 90 μg, or 150 μg weekly, or placebo over 15 days; in a fourth biopsy cohort, patients received either Nexvax2 at the maximum tolerated dose (MTD) or placebo. In the 16-dose study, participants received Nexvax2 150 μg or 300 μg or placebo twice weekly over 53 days; in a third biopsy cohort, patients also received either Nexvax2 at the MTD or placebo. In the 4-week post-treatment period, ascending dose cohorts underwent a further double-blind crossover, placebo-controlled oral gluten challenge, which had a fixed sequence, and biopsy cohorts had a gastroscopy with duodenal biopsies and quantitative histology within 2 weeks without oral gluten challenge. Participants, investigators, and study staff were masked to the treatment assignment, except for the study pharmacist. The primary endpoint was the number and percentage of adverse events in the treatment period in an intention-to-treat analysis. Both trials were completed and closed before data analysis. Trials were registered with the Australian New Zealand Clinical Trials Registry, numbers ACTRN12612000355875 and ACTRN12613001331729.

Findings

Participants were enrolled from Nov 28, 2012, to Aug 14, 2014, in the three-dose study, and from Aug 3, 2012, to Sept 10, 2013, in the 16-dose study. Overall, 62 (57%) of 108 participants were randomly assigned after oral gluten challenge and 20 (71%) of 28 participants were randomly assigned after endoscopy. In the three-dose study, nine participants were randomly allocated to Nexvax2 60 μg and three to placebo (first cohort), nine were allocated to Nexvax2 90 μg and four to placebo (second cohort), eight were allocated to Nexvax2 150 μg and four to placebo (third cohort), and three were allocated to Nexvax2 150 μg and three to placebo (biopsy cohort). In the 16-dose study, eight participants were randomly allocated to Nexvax2 150 μg and four to placebo (first cohort), ten were allocated to Nexvax2 300 μg and three to placebo (second cohort), and seven were allocated to Nexvax2 150 μg and seven to placebo (biopsy cohort). The MTD for Nexvax2 was 150 μg because of transient, acute gastrointestinal adverse events with onset 2–5 h after initial doses of the vaccine, similar to those caused by gluten ingestion. In the ascending dose cohorts in the three-dose study, six (55%) of 11 placebo recipients, five (56%) of nine who received Nexvax2 60 μg, seven (78%) of nine who received Nexvax2 90 μg, and five (63%) of eight who received Nexvax2 150 μg had at least one treatment-emergent adverse event, as did all three (100%) placebo recipients and one (33%) of three Nexvax2 150 μg recipients in the biopsy cohort. In the ascending dose cohorts of the 16-dose study, five (71%) of seven placebo-treated participants, six (75%) of eight who received Nexvax2 150 μg, and all ten (100%) who received Nexvax2 300 μg had at least one treatment-emergent adverse event, as did six (86%) of seven placebo recipients and five (71%) of seven Nexvax2 150 μg recipients in the biopsy cohort. Vomiting, nausea, and headache were the only treatment-emergent adverse events that occurred in at least 5% of participants in either study. Among participants given the MTD, eight gastrointestinal treatment-emergent adverse events occurred in four (50%) of eight participants in the third cohort and none (0%) of three participants in the biopsy cohort in the three-dose study, and five events occurred in five (63%) of eight participants in the first cohort and three events in two (29%) of seven participants in the biopsy cohort of the 16-dose study. Median villous height to crypt depth ratio in distal duodenal biopsies was not significantly different between those who received the vaccine at the MTD on either schedule and those who received placebo. Of the participants who completed the post-treatment oral gluten challenge per protocol, interferon γ release assay to Nexvax2 peptides was negative (responders to treatment) in two (22%) of nine placebo-treated participants in the three-dose study versus two (33%) of six who received Nexvax2 60 μg, five (63%) of eight who received Nexvax2 90 μg, and six (100%) of six who received Nexvax2 150 μg (p=0·007); in the 16-dose study, none (0%) of five placebo-treated participants had a negative assay versus six (75%) of eight who received Nexvax2 150 μg (p=0·021).

Interpretation

The MTD of Nexvax2 was 150 μg for twice weekly intradermal administration over 8 weeks, which modified immune responsiveness to Nexvax2 peptides without deterioration in duodenal histology. The gastrointestinal symptoms that followed the first intradermal administration of the vaccine resembled those associated with oral gluten challenge. These findings support continued clinical development of this potential therapeutic vaccine for coeliac disease.

Funding

ImmusanT.

Introduction

Coeliac disease is an autoimmune-like response to dietary gluten that is characterised by the presence of highly specific autoantibodies to transglutaminase 2 and damage of epithelial cells in the small intestine.1 The community prevalence of coeliac disease is about 1% in children and adults in many regions, including Europe and North America.2 Clinical investigation is usually prompted by digestive symptoms, associated comorbidities such as iron deficiency, or screening family members of probands.2 Abnormal coeliac-specific serology and duodenal histology showing villous atrophy with crypt hyperplasia and intraepithelial lymphocytosis support diagnosis of coeliac diease.2 The only management is a life-long gluten-free diet.2 However, such diets are burdensome and restrictive in social situations, resulting in reduced quality of life and, ultimately, non-adherence.3, 4 Consequently, gluten-free diets seldom result in complete clinical and histological recovery.

Research in context

Evidence before this study

We searched PubMed with the terms “celiac disease” or “coeliac disease” and “non-dietary therapy”, “immunotherapy”, or “vaccine” for reviews of emerging treatments for coeliac disease published in English on or before Aug 24, 2016, the date of our final search. We identified 20 reviews of new treatments in development that had been published since 2009, but found no therapeutics that had been approved specifically for treatment of coeliac disease. We also searched ClinicalTrials.gov with the terms “celiac disease therapy”, “peptide-based immunotherapy”, “peptide immunotherapy”, “antigen-specific immunotherapy”, “epitope-specific immunotherapy”, and “specific immunotherapy” to identify clinical trials listed on or before Aug 24, 2016. We identified clinical trials assessing 16 different agents for coeliac disease, but none was at a stage of development more advanced than phase 2. Our trials were the only antigen-specific therapeutic trials listed for coeliac disease. Our searches identified only two studies of the effects of immunogenic peptides on autoimmune diseases, both for type 1 diabetes (NCT01536431 and NCT02837094, which has not begun). We are also aware of two other clinical trials of immunogenic peptides in multiple sclerosis (NCT01097668 and NCT01973491).

Added value of this study

The clinical effects and therapeutic potential of peptides recognised by disease-specific CD4-positive T cells have not been systematically assessed in clinical autoimmune diseases. Unlike other autoimmune diseases, the peptides that cause the disease-specific CD4-positive T-cell response to gluten associated with coeliac disease are well characterised. To the best of our knowledge, Nexvax2 is the first antigen-specific immunotherapy under development for coeliac disease and is an adjuvant-free solution of three peptides with immunodominant HLA-DQ2·5-restricted epitopes. Intradermal administration of Nexvax2 in HLA-DQ2·5-positive patients with coeliac disease initially caused gastrointestinal symptoms similar in timing and quality to those triggered by gluten ingestion. Adverse events after later administrations of Nexvax2 were no different from those after placebo. A maximum tolerated dose of Nexvax2 150 μg was established. No evidence of deterioration in duodenal histology was noted after twice weekly intradermal administrations over 8 weeks with Nexvax2 150 μg. The recall immune response to gluten was modified in people with coeliac disease receiving Nexvax2, consistent with T cells specific for epitopes in Nexvax2 being rendered unresponsive to further antigenic stimulation.

Implications of all the available evidence

Repeated small, fixed doses of adjuvant-free peptides, specifically immunodominant epitopes for gluten-specific CD4-positive T cells, are capable of modifying the recall immune response to gluten in patients with coeliac disease without causing duodenal injury. Further assessment and clinical development of antigen-specific immunotherapy for coeliac disease with immunogenic gluten peptides is justified.

Coeliac disease has a remarkably strong association with specific MHC haplotypes that accounts for almost half of the total heritable risk.5 About 90% of patients have the MHC class II genes HLA-DQA1*05 and HLA-DQB1*02, which together encode the HLA heterodimer HLA-DQ2·5.6 HLA-DQ2·5 homozygosity is associated with augmented T-cell activation by gluten peptides.7

MHC class II heterodimers have a central role in antigen presentation and the induction and maintenance of acquired cellular immune responses.8 The complex formed by short, antigen-derived peptides (epitopes) loaded into the binding groove of MHC class II molecules binds to the T-cell antigen receptor of CD4-positive T cells, resulting in highly specific antigen recognition and antigen-specific activation. CD4-positive T cells specific for HLA-DQ2·5-restricted gluten peptides that secrete pro-inflammatory cytokines such as interferon γ can be isolated from intestinal tissue.9 1 week after commencing a 3-day gluten food challenge, these same CD4-positive T cells circulate in blood at increased frequencies.10 The aminoacid sequences of immunodominant epitopes recognised by gluten-reactive CD4-positive T cells are well established, and are highly consistent among HLA-DQ2·5-positive people with coeliac disease.9, 10, 11, 12, 13, 14

Epitope-specific immunotherapy is a form of antigen-specific immunotherapy that uses peptides instead of whole antigens to target and modify CD4-positive T cells.15 In general, high doses and increased duration of antigen-specific immunotherapy are most clinically effective.16 Evidence suggests that clinical benefit is related to disease-specific CD4-positive T cells transitioning from a state of responsiveness to antigenic stimulation to a state of reversible functional unresponsiveness (anergy), induction of suppressive regulatory T cells, and, in the long term, deletion and durable immune tolerance.16

Gluten is not suitable for a therapeutic vaccine because it is insoluble, requires deamidation for full immunogenicity, and some gluten peptides and contaminants have direct innate immune activity.17, 18 We developed an adjuvant-free, particle-free solution of three highly soluble synthetic peptides with 15 or 16 aminoacids (NPL001, NPL002, and NPL003; appendix p 10). This vaccine (Nexvax2; ImmusanT, Cambridge, MA, USA) has been designed and developed as an epitope-specific immunotherapy for HLA-DQ2·5-positive coeliac disease (appendix pp 6–7). It encompasses at least five immunodominant epitopes that selectively bind to HLA-DQ2·5 and activate gluten-reactive CD4-positive T cells isolated from HLA-DQ2·5-positive patients with coeliac disease (appendix pp 10, 27).14 These peptides include sequences recognised by anti-gliadin antibodies,19 but are short enough to minimise the likelihood of complement activation by immune complex formation and antibody-mediated hypersensitivity.20

Our aim was to assess the safety and pharmacodynamics of repeated intradermal administrations of Nexvax2 in regimens that could potentially modify gluten-specific immunity in HLA-DQ2·5-positive patients with coeliac disease who are on a gluten-free diet.

Section snippets

Study design and participants

We did two separate randomised, double-blind, placebo-controlled phase 1 studies, one in which Nexvax2 was administered as three fixed doses 1 week apart over 15 days (ie, the three-dose study), and one in which Nexvax2 was administered every 3 or 4 days to a total of 16 doses over 53 days (ie, the 16-dose study; appendix p 29). The studies were designed to establish a maximum tolerated dose by testing incremental dose increases in a series of ascending dose cohorts. To mobilise gluten-specific

Results

Between Nov 28, 2012, and Aug 14, 2014, 102 volunteers were screened for eligibility for the three-dose study, of whom 81 were enrolled. 69 of the participants were screened for inclusion in the ascending dose cohorts and 12 for inclusion in the biopsy cohort (figure 1). 37 participants screened for ascending dose cohorts continued to treatment randomisation after oral gluten challenge, and six participants screened for the biopsy cohort continued to randomisation after gastroscopy (figure 1).

Discussion

These studies show the first potent clinical and immunomodulatory effects of a peptide-based therapeutic vaccine in patients with coeliac disease. The vaccine, which was designed to engage gluten-specific CD4-positive T cells, triggered acute clinical manifestations of coeliac disease after initial doses, but later doses were well tolerated without causing intestinal injury, and ex-vivo interferon γ release by Nexvax2 peptides was lost. Coeliac disease is unique among autoimmune diseases in

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