Mepolizumab does not alter levels of eosinophils, T cells, and mast cells in the duodenal mucosa in eosinophilic esophagitis
2010; Elsevier BV; Volume: 126; Issue: 1 Linguagem: Inglês
10.1016/j.jaci.2010.04.029
ISSN1097-6825
AutoresSébastien Conus, Alex Straumann, Eva Bettler, Hans‐Uwe Simon,
Tópico(s)IL-33, ST2, and ILC Pathways
ResumoTo the Editor: There is evidence that IL-5 plays a pivotal role in the pathology of eosinophilic inflammation by regulating the generation, recruitment, activation, and survival of eosinophils.1Kato M. Kephart G.M. Talley N.J. Wagner J.M. Sarr M.G. Bonno M. et al.Eosinophil infiltration and degranulation in normal human tissue.Anat Rec. 1998; 252: 418-425Crossref PubMed Scopus (177) Google Scholar IL-5 is also crucial in the recruitment of physiological resident intestinal eosinophils.2Mishra A. Hogan S.P. Brandt E.B. Rothenberg M.E. IL-5 promotes eosinophil trafficking to the esophagus.J Immunol. 2002; 188: 2464-2469Google Scholar In contrast with the eosinophils involved in inflammatory processes, the resident eosinophils evoke neither inflammatory reactions nor tissue damage.3Straumann A. Kristl J. Conus S. Vassina E. Spichtin H.S. Beglinger C. et al.Cytokine expression in healthy and inflamed mucosa: probing the role of eosinophils in the digestive tract.Inflamm Bowel Dis. 2005; 11: 720-726Crossref PubMed Scopus (117) Google Scholar Nevertheless, a significant proportion of the resident eosinophils express the eosinophil activation markers CD25 and IL-13, neither of which is detected in circulating blood eosinophils.3Straumann A. Kristl J. Conus S. Vassina E. Spichtin H.S. Beglinger C. et al.Cytokine expression in healthy and inflamed mucosa: probing the role of eosinophils in the digestive tract.Inflamm Bowel Dis. 2005; 11: 720-726Crossref PubMed Scopus (117) Google Scholar In the intestinal mucosa, an intense interplay between different subsets of T cells and eosinophils exists, and it is believed that these mucosa-dwelling cells exhibit homeostatic functions, including the modulation of local immune responses. For instance, eosinophils might be important in antibacterial defense mechanisms.4Yousefi S. Gold J.A. Andina A. Lee J.J. Kelly A.M. Kozlowski E. et al.Catapult-like release of mitochondrial DNA by eosinophils contributes to antibacterial defense.Nat Med. 2008; 14: 949-953Crossref PubMed Scopus (737) Google Scholar Some lymphocyte subsets are also known to be involved in the maintenance of intestinal barrier function as well as in the suppression of chronic inflammation.5Izcue A. Coombes J.L. Powrie F. Regulatory lymphocytes and intestinal inflammation.Annu Rev Immunol. 2009; 27: 313-338Crossref PubMed Scopus (415) Google Scholar Similarly, gastrointestinal mast cells have been shown to play a protective role in defense against parasitic and microbial infections.6Penissi A.B. Rudolph M.I. Piezzi R.S. Role of mast cells in gastrointestinal mucosal defense.Biocell. 2003; 27: 163-172Google Scholar Mepolizumab is a fully humanized monoclonal antibody IgG1 specific for human IL-5. Mepolizumab blocks the binding of human IL-5 to the α-chain of the IL-5 receptor (R)–α complex, which is mainly expressed on the eosinophil cell surface. The use of mepolizumab for any indication could interfere with the function of the physiological resident intestinal eosinophils, particularly because the drug reduces blood eosinophils by more than 90%.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar, 8Conus S. Straumann A. Simon H.U. Anti-IL-5 (mepolizumab) therapy does not alter IL-5 receptor α levels in patients with eosinophilic esophagitis.J Allergy Clin Immunol. 2008; 123: 269-270Abstract Full Text Full Text PDF Scopus (17) Google Scholar Therefore, we were interested to investigate whether IL-5 blockade would reduce the physiologic eosinophil infiltration of the duodenum. We have recently reported that mepolizumab, in a clinical, randomized, double-blind, placebo-controlled study, reduced mean eosinophil numbers in the esophageal epithelial layer of adult patients with eosinophilic esophagitis (EoE) by approximately 50%.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Here we report the results of the analysis of duodenal biopsies, which were collected within this study.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Besides eosinophils, we analyzed the numbers of T cells and mast cells in the same duodenal biopsies because these 2 cell types also appear to play important physiologic functions in the gastrointestinal tract.5Izcue A. Coombes J.L. Powrie F. Regulatory lymphocytes and intestinal inflammation.Annu Rev Immunol. 2009; 27: 313-338Crossref PubMed Scopus (415) Google Scholar, 6Penissi A.B. Rudolph M.I. Piezzi R.S. Role of mast cells in gastrointestinal mucosal defense.Biocell. 2003; 27: 163-172Google Scholar Eleven adults with active EoE (>20 peak eosinophil number/high-power field [hpf] and dysphagia) were randomized to 750 mg mepolizumab (n = 5) or placebo (n = 6) by intravenous infusions at days 0 and 7.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Those not in complete remission (<5 peak eosinophil number/hpf) 4 weeks postinjection received 2 further doses 4 weeks apart (week 5 and 9) of 1500 mg mepolizumab or placebo according to their original randomization.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Patients underwent esophagogastroduodenoscopy, and biopsies and blood were taken before the first infusion (before therapy), 4 weeks after the first infusion (week 4), and 4 weeks after the last infusion (week 13).7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Patients then progressed to the short-term follow-up phase for 8 weeks, such that all patients were assessed 21 weeks after the start of treatment for blood eosinophil counts in particular.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Patients then continued into the long-term follow-up phase, which ended with an assessment at 34 weeks after the last infusion (week 43). Infiltration of cells in the subepithelial layers of duodenal specimens was assessed by immunofluorescence analysis at pretreatment (before therapy) and at 13 weeks after the first infusion of mepolizumab or placebo.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Indirect immunostaining was carried out on 4-μm-thick, paraformaldehyde-fixed, paraffin-embedded sections by using the following primary antibodies: antieosinophil cationic protein (EG1; Pharmacia & Upjohn Diagnostics AB, Uppsala, Sweden), anti-CD3 (DakoCytomation, Glostrup, Denmark), antitryptase (DakoCytomation), anti–IL-5Rα (Proteintech Europe Ltd, Manchester, United Kingdom), anti–IL-13 (Santa Cruz Biotechnology, distributed by LabForce AG, Nunningen, Switzerland) and anti-CD25 (Immunotech, Marseille, France). At least 4 biopsy specimens were sampled endoscopically from the duodenum of 11 adult patients with EoE. On each biopsy specimen, cells were randomly counted in 10 consecutive hpfs (area of 1 hpf, 0.538 mm2), according to a predetermined sequence (eg, left part of the lowest piece of tissue) that was similar for all slides, such that 40 counts were conducted (10 × 4 specimens) for each patient at each time point. The mean numbers of positive cells were derived by confocal laser scanning microscopy (LSM 510; Carl Zeiss MicroImaging GmbH, Jena, Germany) equipped with argon and helium-neon lasers. IL-13 and CD25 expression was determined on autofluorescent eosinophils.3Straumann A. Kristl J. Conus S. Vassina E. Spichtin H.S. Beglinger C. et al.Cytokine expression in healthy and inflamed mucosa: probing the role of eosinophils in the digestive tract.Inflamm Bowel Dis. 2005; 11: 720-726Crossref PubMed Scopus (117) Google Scholar The baseline characteristics of the patients (Table I) illustrate that the treatment groups were largely similar with the exception that there was a higher proportion of men in the mepolizumab group (80%) compared with placebo (50%). Although mepolizumab significantly reduced eosinophil numbers in the blood and esophageal tissues of patients with EoE,7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar mean numbers of eosinophils and mean of IL-5Rα expression levels on eosinophils in the duodenal tissues did not differ between the mepolizumab and placebo groups (Table II). The latter result is in agreement with our previous report in which we did not observe changes in IL-5Rα expression levels on blood eosinophils of the same patients.8Conus S. Straumann A. Simon H.U. Anti-IL-5 (mepolizumab) therapy does not alter IL-5 receptor α levels in patients with eosinophilic esophagitis.J Allergy Clin Immunol. 2008; 123: 269-270Abstract Full Text Full Text PDF Scopus (17) Google Scholar However, it should be noted that in both groups, the numbers of IL-5Rα–positive cells are smaller compared with the numbers of eosinophil cationic protein–positive cells (Table II), suggesting that a proportion of duodenal eosinophils exhibits low or no IL-5Rα surface expression. Similar findings were reported earlier regarding eosinophils infiltrating nasal polyp tissues.9Gevaert P. Hellman C. Lundblad L. Lundahl J. Holtappels G. van Cauwenberge P. et al.Differential expression of the interleukin 5 receptor alpha isoforms in blood and tissue eosinophils of nasal polyp patients.Allergy. 2009; 64: 725-732Crossref Scopus (31) Google ScholarTable IPatient characteristicsMepolizumab (n = 5)Placebo (n = 6)Mean age (y)32.434.0Male (%)8050Caucasians (%)100100Mean weight (kg)74.075.3Mean duration of EoE (y)5.325.28Mean age of onset (y)26.628.7Atopic disease at screen Asthma, n (%)1 (20)2 (33) Eczema, n (%)1 (20)0 Food allergy, n (%)1 (20)0 Rhinitis, n (%)4 (80)4 (67)Mean peak esophageal eosinophils at screen (per hpf)201202 Open table in a new tab Table IIDuodenal and esophageal tissue infiltration by inflammatory cells before and after mepolizumab or placebo treatmentMeanSDPercent changeP valueDuodenum EosinophilsMepolizumabBefore17.02.3 per hpf(n = 5)After17.61.9+4NS (ECP+ cells)PlaceboBefore16.30.9(n = 6)After15.92.2−2NS EosinophilsMepolizumabBefore9.01.9 per hpf(n = 5)After10.03.0+11NS (IL-5Rα+ cells)PlaceboBefore8.51.4(n = 6)After8.22.2−4NS Mast cellsMepolizumabBefore16.64.4 per hpf(n = 5)After15.23.0−9NS (tryptase+ cells)PlaceboBefore16.73.8(n = 6)After16.02.4−4NS T cellsMepolizumabBefore86.313.3 per hpf(n = 5)After96.510.9+12NS (CD3+ cells)PlaceboBefore84.818.4(n = 6)After86.814.9+2NS Activated eosinophilsMepolizumabBefore16.75.1 per hpf(n = 5)After14.92.0−11NS (IL-13+ eosinophils)PlaceboBefore16.83.3(n = 6)After16.34.7−3NS Activated eosinophilsMepolizumabBefore8.47.0 per hpf(n = 5)After9.09.3+7NS (CD25+ eosinophils)PlaceboBefore13.82.6(n = 6)After13.25.0−5NSEsophagus EosinophilsMepolizumabBefore73.250.6 per hpf(n = 5)After32.828.3−55.011 (ECP+ cells)PlaceboBefore56.733.4(n = 6)After52.728.8−7NSBlood EosinophilsMepolizumabBefore492213 per mm3(n = 5)After5027−90.009PlaceboBefore532291(n = 6)After440338−17NSECP, Eosinophil cationic protein; NS, not significant. Open table in a new tab ECP, Eosinophil cationic protein; NS, not significant. The mean numbers of infiltrating mast cells (tryptase-positive cells) and T cells (CD3-positive cells) did not change significantly as a consequence of mepolizumab or placebo treatment (Table II). Because intestinal eosinophils have been shown to express, at least partially, CD25 and IL-13,3Straumann A. Kristl J. Conus S. Vassina E. Spichtin H.S. Beglinger C. et al.Cytokine expression in healthy and inflamed mucosa: probing the role of eosinophils in the digestive tract.Inflamm Bowel Dis. 2005; 11: 720-726Crossref PubMed Scopus (117) Google Scholar we also investigated the effect of mepolizumab treatment on the expression of these 2 markers. Again, the proportions of CD25 and IL-13–positive duodenal eosinophils did not change as a consequence of therapy (Table II). In summary, an anti–IL-5 treatment had no effect on the duodenal infiltration of eosinophils, T cells, and mast cells in subjects participating in the first placebo-controlled clinical study performed in adult patients with EoE.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar Whereas "inflammatory" eosinophils at the site of inflammation are reduced by approximately 50%, physiologic eosinophil infiltration, at least in the duodenum, is not affected, even at the high 1500-mg dose level used in this study.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar In addition, the anti–IL-5 therapy did not change the expression levels of IL-5Rα and the eosinophil activation markers CD25 and IL-13, suggesting that the relative proportions of potential functionally different eosinophil subgroups also did not change. Therefore, although the study is limited because of the small number of subjects studied, levels of duodenal eosinophils, T cells, and mast cells do not seem to be affected by anti–IL-5 antibody therapy, and our data support the concept that targeting IL-5 by using mepolizumab is a safe therapy.7Straumann A. Conus S. Grzonka P. Kita H. Kephart G. Bussmann C. et al.Anti-interleukin-5 antibody treatment (mepolizumab) in active eosinophilic oesophagitis: a randomized, placebo-controlled, double-blind trial.Gut. 2010; 59: 21-30Crossref PubMed Scopus (467) Google Scholar CorrectionJournal of Allergy and Clinical ImmunologyVol. 127Issue 3PreviewWith regard to the July 2010 article by Conus et al (J Allergy Clin Immunol 2010;126:175-7), the area of 1 hpf is incorrect as given. The correct area of 1 hpf is 0.0538 mm2. Full-Text PDF
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