In patients with severe asthma with eosinophilia in reslizumab clinical trials, high peripheral blood eosinophil levels are associated with low FEV1 reversibility.

Background A post hoc analysis of two randomized, placebo–controlled, Phase 3 trials of intravenous reslizumab, an anti-interleukin-5 (IL-5) biologic for severe eosinophilic asthma. Methods Relationships between baseline blood eosinophil levels (EOS), forced expiratory volume in 1 s (FEV1) reversibility to β2-agonists and treatment outcomes were assessed. Results Mean baseline FEV1 reversibility was numerically lower among patients with high (≥ 400 cells/µL) versus low baseline EOS. Reslizumab produced clinically significant improvement in FEV1, exacerbation rates and patient-reported outcomes after 52 weeks, including in patients with high EOS and low FEV1 reversibility (≤ 14%) to β2-agonists at baseline. Conclusions Clinical trial eligibility criteria stipulating minimum FEV1 reversibility to β2-agonists of ≥ 12% might exclude patients who would benefit from treatment with anti-IL-5 biologics.


Introduction
Severe non-allergic, adult-onset or intrinsic asthma is frequently associated with pronounced eosinophilia [1][2][3][4]. Interleukin-5 (IL-5) is a potent activator of eosinophils and enhances their viability [5]. IL-5 activity has been demonstrated to inversely correlate with pulmonary function in patients with asthma [4], and anti-IL-5 treatment has been shown to improve asthma control in patients with severe asthma and eosinophilia [6].
Reslizumab is an IgG4-kappa humanized monoclonal antibody targeting IL-5 [7]. In Phase 3 clinical trials intravenous (IV) reslizumab dosed at 3 mg/kg once every 4 weeks (q4w) was associated with a significant reduction in the risk of clinical asthma exacerbations (CAEs) and improved asthma control, lung function, and quality of life in patients with inadequately controlled asthma with blood eosinophil levels (EOS) ≥ 400 cells/µL and a history of CAEs [8]. Reslizumab has been indicated as add-on maintenance treatment for adult patients with severe eosinophilic asthma [9].
Among the entry criteria for these trials, patients had to have EOS ≥ 400 cells/µL at screening. In addition, based on traditional concepts to substantiate the presumed diagnosis of asthma, all patients had to demonstrate a reversibility of their forced expiratory volume in 1 s (FEV 1 ) in response to the inhalation of a β 2agonist (albuterol 200 µg) of ≥ 12%. A recent longitudinal cohort study in young adults, with and without asthma,

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Allergy, Asthma & Clinical Immunology has shown that elevated EOS are associated with airflow obstruction [10]. We therefore hypothesized that any substantial increase in FEV 1 following the administration of a β 2 -agonist might reflect asthma dominated by smooth muscle contraction rather than eosinophilic inflammation. On the other hand, a poor response to β 2agonists in patients with severe asthma and eosinophilia might predict a better response to anti-IL-5 therapy. To substantiate this we performed a post hoc analysis of data from two Phase 3 trials of IV reslizumab (NCT01287039 and NCT01285323) [8] to assess the relationship between EOS, reversibility of airway obstruction and treatment response to reslizumab therapy in a well-characterized population of patients with inadequately controlled moderate-to-severe asthma with EOS ≥ 400 cells/µL.

Study design and patients
The two duplicate trials enrolled patients aged 12-75 years with inadequately controlled asthma (Asthma Control Questionnaire-7 [ACQ-7] score ≥ 1.5) on medium-to-high doses of inhaled corticosteroids (ICS), and who had screening EOS ≥ 400 cells/μL, ≥ 1 CAE in the previous year, and FEV 1 reversibility of ≥ 12% with albuterol [8]. The selection criteria for FEV 1 reversibility were chosen based on National Asthma Education and Prevention Program guidelines available at the time of study design [11].
Both trials were conducted in accordance with Good Clinical Practice guidelines, the Declaration of Helsinki, and local regulatory requirements. All patients provided written informed consent, and the relevant health authorities and local ethics committees or institutional review boards approved the study protocols.
Following a 2-4-week screening period, patients were randomized (1:1) to receive IV reslizumab (3.0 mg/kg) or matching placebo q4w for 52 weeks. Patients continued their usual asthma treatment during the screening, run-in and treatment periods. Pre-bronchodilator spirometry, Asthma Symptom Utility Index (ASUI), and ACQ-7 were assessed q4w at the scheduled clinic visits, from day of randomization to the end of treatment. Possible cases of CAEs were assessed by questioning of the patient at every scheduled monthly visit. Asthma Quality of Life Questionnaire (AQLQ) score was assessed at baseline and weeks 16, 32, and 52.

Outcome measures
FEV 1 reversibility at baseline (during screening) was assessed according to EOS category at baseline (day of first dose). Categories for baseline FEV 1 reversibility were arbitrarily set at < 14%, 14 to < 16%, 16 to < 20% and ≥ 20%, with baseline EOS categories set arbitrarily at < 150 cells/µL, 150 to < 400 cells/µL, 400 to < 700 cells/ µL and ≥ 700 cells/µL. Given that blood eosinophil counts are known to be variable over time, assessment of blood eosinophil count at baseline allowed for selection of patients with persistently elevated blood eosinophils ≥ 400 cells/µL at two timepoints (screening and baseline).

Statistical analysis
An analysis of covariance was used to model change from baseline at Week 52 in lung function and patientreported outcomes (ACQ-7, AQLQ and ASUI) with fixed factors for treatment arm, sex, oral corticosteroid use at baseline (Yes or No), region (USA or Other), and a continuous covariate for height.
CAEs counted are those which occurred between the completion of the first dose of study drug and 2 weeks after the end of treatment/early withdrawal visit. CAE rates, CAE rate ratio, and confidence intervals (CIs) and p values are based on a negative binomial regression model adjusted for baseline usage of oral corticosteroid (Yes or No) and region (USA or other).
All analyses were conducted using SAS version 9.4 (SAS Institute Inc., Cary, NC, USA).

Results
A total of 953 patients were randomized in the two duplicate Phase 3 studies (reslizumab: n = 477; placebo: n = 476). Patient demographics and clinical characteristics at baseline were similar between reslizumab and placebo groups (Table 1).

Baseline FEV 1 reversibility according to eosinophil group
Those patients who had low baseline EOS (< 150 cells/ µL or 150 to < 400 cells/µL) had a higher mean FEV 1 reversibility and a higher proportion of patients who were highly reversible to inhaled β 2 -agonists (≥ 20% reversibility, 60% and 62.6% of the subgroup populations) compared with patients with higher EOS (Fig. 1). The proportion of patients who responded relatively poorly to β 2 -agonists (< 14% improvement) was largest in the EOS ≥ 700 cells/µL group (17.4%) compared with other EOS groups, and this high EOS group had the numerically lowest mean reversibility (Fig. 2). Figure 3 shows the observed treatment effects for reslizumab on FEV 1 Table 2.  Table 3). The CAE rate ratio for reslizumab versus placebo was numerically lower in the remaining overall population excluding those with EOS ≥ 400 cells/µL and < 14% reversibility compared to the high EOS/low β 2 -agonist reversibility group, although numerical differences did not reach statistical significance between groups. Baseline values and treatment effects on ACQ, AQLQ and ASUI in the high EOS/low β 2 -agonist reversibility group and the remaining overall population excluding those with EOS ≥ 400 cells/µL and < 14% reversibility are shown in Table 3. Similar to the lung function findings, we observed a numerically greater treatment effect for reslizumab versus placebo across the asthma clinical outcomes in the high EOS/low β 2 -agonist reversibility group than in the remaining overall population excluding those with EOS ≥ 400 cells/µL and < 14% reversibility, with no statistically significant differences between groups.

Discussion
Eosinophilia is an important but often variable feature of bronchial asthma, which is highly responsive to corticosteroids and anti-IL-5 therapy. Based on our observation we can postulate that patients with the highest EOS were more likely to have a relatively poor reversibility to β 2 -agonists, but were as likely to have  a marked response to treatment with the anti-IL-5 antagonist reslizumab as the remaining population. Unfortunately, due to standard asthma inclusion criteria for reversibility used in this study, no patients with EOS ≥ 400 cells/µL and a β 2 -agonist reversibility of ≤ 12% were studied. Based on our observed results it can be speculated, however, that these patients might have benefited from treatment with reslizumab as the lack of FEV 1 increase following the administration of a β 2 -agonist might reflect asthma dominated by eosinophilic inflammation. Therefore, this population should be specifically investigated in a future study. At present, our data suggest that high β 2 -agonist reversibility might not be the best predictor of response to anti-IL-5 treatment while low β 2 -reversibility (in the presence of a marked eosinophilia) might be indicative of eosinophil-dependent asthma. Future studies should therefore test if the combination of the two 'biomarkers' , namely a low β 2 -agonist reversibility together with a high peripheral blood eosinophilia, might be a better predictor of response to anti-IL-5 therapy than either parameter alone or other markers such as fractioned exhaled nitric oxide currently used to predict treatment responses in asthma. Finally, our observation suggests that in asthma low β 2 -agonist reversibility in the presence of eosinophilia should not lead to the assumption of an irreversibility of airflow obstruction, but in contrast might be indicative of persistent eosinophilic inflammation rather than 'remodeling' , which might predict a marked responsiveness to antiinflammatory treatment with corticosteroids and/or anti-IL-5 treatments.
In this study, we noted a large increase in FEV 1 from baseline in the placebo groups, particularly in the EOS high/β 2 -agonist reversibility low group. The marked improvement in FEV 1 after 52 weeks in the EOS high/ β 2 -agonist reversibility low group receiving placebo may be due to a 'trial effect' including increased adherence to ICS, which might preferentially result in improvement in eosinophilic patients. However, the improvement on reslizumab was numerically better in this group than in the remaining patients, suggesting that a possibly good response in FEV 1 to ICS does not seem to interfere with the actions of reslizumab on airflow obstruction in eosinophilic patients.

Limitations of the study
Our study is limited by the post hoc nature of our analysis and the relatively small number of patients in the subgroup of interest (n = 52 in the group with the lowest reversibility to β 2 -agonists treated with reslizumab). Furthermore, our hypothesis that the biomarker combination of high peripheral blood eosinophilia/low β 2 -agonist reversibility might be an ideal predictor of response to reslizumab is limited by the inclusion criteria of FEV 1 reversibility of ≥ 12% precluding assessment of patients with lower baseline reversibility values.

Conclusion
Higher baseline EOS are associated with numerically lower FEV 1 reversibility in patients with inadequately controlled asthma with eosinophilia. Reslizumab treatment resulted in clinically significant improvements in asthma clinical outcomes in patients with high EOS and low FEV 1 reversibility at baseline. Therefore, reslizumab may preferentially reverse airway obstruction which appears 'fixed' to β 2agonists (albuterol) due to action on IL-5 and airway eosinophils. The exclusion of patients with airflow obstruction poorly responsive to β 2 -agonists from clinical trials of biologics for severe asthma may result in an under-representation of patients with enhanced inflammation with eosinophils who might in fact particularly benefit from this treatment.