Targeting IL-13 with tralokinumab normalizes type 2 inflammation in atopic dermatitis both early and at 2 years.

BACKGROUND: Tralokinumab is a monoclonal antibody that specifically neutralizes interleukin (IL)-13, a key driver of skin inflammation and barrier abnormalities in atopic dermatitis (AD). This study evaluated early and 2-year impacts of IL-13 neutralization on skin and serum biomarkers following tralokinumab treatment in adults with moderate-to-severe AD. METHODS: Skin biopsies and blood samples were evaluated from a subset of patients enrolled in the Phase 3 ECZTRA 1 (NCT03131648) and the long-term extension ECZTEND (NCT03587805) trials. Gene expression was assessed by RNA sequencing; protein expression was assessed by immunohistochemistry and immunoassay. RESULTS: Tralokinumab improved the transcriptomic profile of lesional skin by Week 4. Mean improvements in the expression of genes dysregulated in AD were 39% at Week 16 and 85% at 2 years with tralokinumab, with 15% worsening at Week 16 with placebo. At Week 16, tralokinumab significantly decreased type 2 serum biomarkers (CCL17/TARC, periostin, and IgE), reduced epidermal thickness versus placebo, and increased loricrin coverage versus baseline. Two years of tralokinumab treatment significantly reduced expression of genes in the Th2 (IL4R, IL31, CCL17, and CCL26), Th1 (IFNG), and Th17/Th22 (IL22, S100A7, S100A8, and S100A9) pathways as well as increased expression of epidermal differentiation and barrier genes (CLDN1 and LOR). Tralokinumab also shifted atherosclerosis signaling pathway genes (SELE, IL-37, and S100A8) toward non-lesional expression. CONCLUSION: Tralokinumab treatment improved epidermal pathology, reduced systemic markers of type 2 inflammation, and shifted expression of key AD biomarkers in skin towards non-lesional levels, further highlighting the key role of IL-13 in the pathogenesis of AD. CLINICAL TRIAL REGISTRATION: NCT03131648, NCT03587805.

Systemic Biologic Management of Atopic Dermatitis.

Dupilumab and tralokinumab are currently the only FDA-approved biologic therapies for the treatment of moderate-to-severe atopic dermatitis. Tralokinumab is approved for patients greater than 18 years old, and dupilumab is approved for patients as young as 6 months old. Both medications are effective in clinical trials at improving atopic dermatitis. With a good safety profile and low-risk adverse events, dupilumab and tralokinumab are generally excellent treatment options for patients with severe or refractory atopic dermatitis.

Successful treatment of dyshidrotic palmoplantar eczema with tralokinumab.

Dyshidrotic palmoplantar eczema or pompholyx is considered to be a part of the spectrum of atopic dermatitis with a significant impact on the quality of life and limited treatment options. Tralokinumab is a new fully human monoclonal antibody which neutralizes interleukin 13, a chief cytokine in itch pathogenesis and skin barrier defects. Tralokinumab is FDA-approved for the treatment of atopic dermatitis in adults and EMA-approved for the treatment of atopic dermatitis in adults and adolescents. We, hereby, report a 40-year-old female with severe dyshidrotic palmoplantar eczema who was successfully treated with tralokinumab. To the best of our knowledge, this is the first report of the efficacious use of tralokinumab in dyshidrotic eczema.

Efficacy and safety of tralokinumab in the treatment of head and neck pattern atopic dermatitis: A multicentre study of 12 patients.

BACKGROUND/OBJECTIVES: The evaluation of the efficacy and safety of new molecules for atopic dermatitis (AD) in real clinical practice is very important to obtain information that clinical trials (EECC) lack. The pattern of AD in the head and neck (H&N) continues to be a challenge in treatment today, despite the new molecules, and real-life data on the use of tralokinumab is still missing. This is the first daily practice study of tralokinumab treatment in patients with H&N AD pattern. The objective is to evaluate the efficacy and safety of tralokinumab in the short term (16 weeks) in patients with AD with H&N pattern, for the first time. METHODS: A multicentre prospective observational study was conducted, including patients with moderate-severe AD and H&N pattern who started tralokinumab treatment in four hospitals in Andalusia. Values of severity and quality of life scales, as well as patient-reported outcomes (PROs), were collected at baseline and at Weeks 4 and 16. Safety events were also recorded. RESULTS: Twelve patients were included. An improvement was observed in all efficacy and quality of life parameters evaluated at 16 weeks with respect to the baseline. No serious adverse events were recorded. CONCLUSIONS: In real clinical practice, tralokinumab is demonstrated to be an effective and safe treatment for patients with AD and H&N pattern at short term.

Systemic treatments for atopic dermatitis (eczema): Systematic review and network meta-analysis of randomized trials.

BACKGROUND: Atopic dermatitis (AD) is an inflammatory skin condition with multiple systemic treatments and uncertainty regarding their comparative impact on AD outcomes. OBJECTIVE: We sought to systematically synthesize the benefits and harms of AD systemic treatments. METHODS: For the 2023 American Academy of Allergy, Asthma & Immunology and American College of Allergy, Asthma, and Immunology Joint Task Force on Practice Parameters AD guidelines, we searched MEDLINE, EMBASE, CENTRAL, Web of Science, and GREAT databases from inception to November 29, 2022, for randomized trials addressing systemic treatments and phototherapy for AD. Paired reviewers independently screened records, extracted data, and assessed risk of bias. Random-effects network meta-analyses addressed AD severity, itch, sleep, AD-related quality of life, flares, and harms. The Grading of Recommendations Assessment, Development and Evaluation approach informed certainty of evidence ratings. This review is registered in the Open Science Framework (https://osf.io/e5sna). RESULTS: The 149 included trials (28,686 patients with moderate-to-severe AD) evaluated 75 interventions. With high-certainty evidence, high-dose upadacitinib was among the most effective for 5 of 6 patient-important outcomes; high-dose abrocitinib and low-dose upadacitinib were among the most effective for 2 outcomes. These Janus kinase inhibitors were among the most harmful in increasing adverse events. With high-certainty evidence, dupilumab, lebrikizumab, and tralokinumab were of intermediate effectiveness and among the safest, modestly increasing conjunctivitis. Low-dose baricitinib was among the least effective. Efficacy and safety of azathioprine, oral corticosteroids, cyclosporine, methotrexate, mycophenolate, phototherapy, and many novel agents are less certain. CONCLUSIONS: Among individuals with moderate-to-severe AD, high-certainty evidence demonstrates that high-dose upadacitinib is among the most effective in addressing multiple patient-important outcomes, but also is among the most harmful. High-dose abrocitinib and low-dose upadacitinib are effective, but also among the most harmful. Dupilumab, lebrikizumab, and tralokinumab are of intermediate effectiveness and have favorable safety.

Tralokinumab therapy for moderate-to-severe atopic dermatitis: Clinical outcomes with targeted IL-13 inhibition.

Atopic dermatitis (AD) is a chronic, inflammatory, intensely pruritic skin disorder associated with significant patient burden. Interleukin (IL)-13 is a cytokine that acts as a driver of immune dysregulation, skin-barrier dysfunction, and microbiome dysbiosis that characterizes AD, and is consistently overexpressed in AD skin. Tralokinumab is a fully human immunoglobulin (Ig) G4 monoclonal antibody that binds specifically to IL-13 with high affinity, thereby inhibiting subsequent downstream IL-13 signaling. Three pivotal phase 3 clinical trials demonstrated that tralokinumab 300 mg every other week, as monotherapy or in combination with topical corticosteroids as needed, provides significant improvements in signs and symptoms of moderate-to-severe AD, as measured by Investigator's Global Assessment 0/1 (clear/almost clear) and Eczema Area and Severity Index-75 at Week 16. Improvements were observed soon after tralokinumab initiation and were maintained over 52 weeks of therapy. Tralokinumab significantly improved patient-reported outcomes such as itch and sleep, and demonstrated a safety profile comparable with placebo; conjunctivitis during tralokinumab therapy was generally mild. Similar results were observed in a phase 3 adolescent trial. The role of IL-13 in the pathophysiology of AD justifies a targeted approach and a wealth of clinical data supports tralokinumab as a new therapeutic option for people with moderate-to-severe AD.

IL-13, periostin and dipeptidyl-peptidase-4 reveal endotype-phenotype associations in atopic dermatitis.

BACKGROUND: The heterogeneous (endo)phenotypes of atopic dermatitis (AD) require precision medicine. Currently, systemic therapy is recommended to patients with an Eczema Area and Severity Index (EASI)≥16. Previous studies have demonstrated an improved treatment response to the anti-interleukin (IL)-13 antibody tralokinumab in AD subgroups with elevated levels of the IL-13-related biomarkers dipeptidyl-peptidase (DPP)-4 and periostin. METHODS: Herein, 373 AD patients aged≥12 years were stratified by IL-13high , periostinhigh and DPP-4high endotypes using cross-sectional data from the ProRaD cohort Bonn. "High" was defined as >80th quantile of 47 non-atopic controls. We analyzed endotype-phenotype associations using machine-learning gradient boosting compared to logistic regression. RESULTS: AD severity and eosinophils correlated with IL-13 and periostin levels. Correlations of IL-13 with EASI were stronger in patients with increased (rs=0.482) than with normal (rs=0.342) periostin levels. We identified eosinophilia>6% and an EASI range of 5.5-17 dependent on the biomarker combination to be associated with increasing probabilities of biomarkerhigh endotypes. Also patients with mild-to-low-moderate severity (EASI<16) featured increased biomarkers (IL-13high : 41%, periostinhigh : 48.4%, DPP-4high : 22.3%). Herthoge sign (adjusted Odds Ratio (aOR)=1.89, 95% Confidence Interval (CI) [1.14-3.14]) and maternal allergic rhinitis (aOR=2.79-4.47) increased the probability of an IL-13high -endotype, "dirty neck" (aOR=2.83 [1.32-6.07]), orbital darkening (aOR=2.43 [1.08-5.50]), keratosis pilaris (aOR=2.21 [1.1-4.42]) and perleche (aOR=3.44 [1.72-6.86]) of a DPP-4high -endotype. CONCLUSIONS: A substantial proportion of patients with EASI<16 featured high biomarker levels suggesting systemic impact of skin inflammation already below the current cut-off for systemic therapy. Our findings facilitate the identification of patients with distinct endotypes potentially linked to response to IL-13-targeted therapy.

Tralokinumab treatment improves the skin microbiota by increasing the microbial diversity in adults with moderate-to-severe atopic dermatitis: Analysis of microbial diversity in ECZTRA 1, a randomized controlled trial.

BACKGROUND: Atopic dermatitis (AD) is characterized by microbial dysbiosis, immune dysregulation, and an impaired skin barrier. Microbial dysbiosis in AD involves a reduction in diversity primarily driven by an increased abundance of Staphylococcus aureus. Tralokinumab, an approved treatment for adults with moderate-to-severe AD, improves the skin barrier and immune abnormalities by specifically targeting the interleukin 13 cytokine, but its impact on the skin microbiome is unknown. OBJECTIVE: To investigate how tralokinumab affects the skin microbiome by examining the lesional skin of adults with moderate-to-severe AD from the phase 3 ECZTRA 1 trial (NCT03131648). METHODS: Microbiome profiling, S aureus abundance, and biomarker data were assessed in a subset of ECZTRA 1 participants (S aureus abundance at baseline and week 16; microbiome profiling at baseline, and week 8/16; and serum sampling before dose and week 4/8/16/28/52). RESULTS: Tralokinumab treatment led to increased microbial diversity, reduced S aureus abundance, and increased abundance of the commensal coagulase-negative Staphylococci. LIMITATIONS: Limitations include a lack of S aureus abundance data at week 8, sampling site variation between participants, and possible influence from concomitant systemic antiinfectives. CONCLUSION: Our findings indicate specific targeting of the interleukin 13 cytokine with tralokinumab can directly and/or indirectly improve microbial dysbiosis seen in AD skin.

Review of Tralokinumab in the Treatment of Atopic Dermatitis.

OBJECTIVE: To review pharmacokinetics, efficacy, and safety of tralokinumab in treatment of atopic dermatitis (AD). DATA SOURCES: Literature review was conducted using MEDLINE (PubMed), EMBASE, and ClinicalTrials.gov for articles published between January 2010 and May 2022. STUDY SELECTION AND DATA EXTRACTION: Articles in English discussing tralokinumab in AD were included. DATA SYNTHESIS: In one phase 2 trial, more subjects treated with tralokinumab 150 and 300 mg achieved an Investigator's Global Assessment (IGA) of 0/1 with minimum ≥2 point IGA reduction (23%), versus placebo (11.8%, P = 0.10). During 2 phase 3 trials, more subjects treated with tralokinumab achieved IGA success (ECZTRA 1: 15.8% and ECZTRA 2: 22.2%), versus placebo (7.1% and 10.9%, respectively; P = 0.002 and P < 0.001). During one phase 3 trial, in conjunction with topical corticosteroids (TCS), more subjects treated with tralokinumab 300 mg achieved IGA success (ECZTRA 3: 38.9%), versus placebo (26.2%, P = 0.015). During another phase 3 trial in subjects with resistance or contraindication to oral cyclosporine, more subjects treated with tralokinumab 300 mg achieved an Eczema Area Severity Index 75 (64.2%), versus placebo (50.5%, P = 0.018). RELEVANCE TO PATIENT CARE AND CLINICAL PRACTICE: Tralokinumab is efficacious for moderate-to-severe AD, as monotherapy, in conjunction with TCS, and resistance or contraindication to cyclosporine. Although IL-4 and IL-13 are both implicated in AD's pathogenesis, IL-13 is overexpressed, and head-to-head trials are needed to assess efficacy of tralokinumab, versus dupilumab. Compared with upadacitinib and abrocitinib, tralokinumab is not associated with black-box warnings. CONCLUSIONS: Tralokinumab is an efficacious and safe systemic treatment for moderate-to-severe AD.

Tralokinumab for Moderate-to-Severe Atopic Dermatitis in Adults.

Atopic dermatitis (AD) is a common, chronic, recurrent, immune-mediated inflammatory skin disease. Targeted treatment options remain limited. Tralokinumab (Adtralza®) is a promising, new systemic therapy that inhibits interleukin-13. It was recently approved by Health Canada and the US FDA for the treatment of moderate-to-severe AD in adults and may be used alone or with topical corticosteroids. Herein, we review the efficacy and safety of tralokinumab in adults, as demonstrated in clinical trials.

Long-term 2-year safety and efficacy of tralokinumab in adults with moderate-to-severe atopic dermatitis: Interim analysis of the ECZTEND open-label extension trial.

BACKGROUND: Additional long-term treatments are needed for moderate-to-severe atopic dermatitis (AD). An ongoing, open-label, 5-year extension trial, ECZTEND (NCT03587805), assesses tralokinumab plus optional topical corticosteroids in participants from previous tralokinumab parent trials (PTs) with moderate-to-severe AD. OBJECTIVE: To evaluate the safety and efficacy of up to 2 years tralokinumab treatment in a post hoc interim analysis. METHODS: Safety analyses included adults from completed PTs enrolled in ECZTEND, regardless of tralokinumab exposure duration. Efficacy analyses included adult participants treated with tralokinumab in ECZTEND for ≥1 year and subgroup analyses of those on tralokinumab for 2 years (1 year from PT, 1 year in ECZTEND). Primary end point was the number of adverse events with additional efficacy end points. RESULTS: Participants on tralokinumab had an exposure-adjusted rate of 237.8 adverse events/100 patient-years' exposure (N = 1174) in the safety analysis set. Exposure-adjusted incidence rates of common adverse events were comparable to PTs, although at lower rates. With 2 years of tralokinumab, improvements in extent and severity of AD were sustained, with Eczema Area and Severity Index (EASI-75) in 82.5% of participants (N = 345). LIMITATIONS: Possible selection bias; no placebo arm; some participants experienced treatment gaps between PTs and ECZTEND. CONCLUSION: Over 2 years, tralokinumab was well tolerated and maintained long-term control of AD signs and symptoms.

Tralokinumab does not impact vaccine-induced immune responses: Results from a 30-week, randomized, placebo-controlled trial in adults with moderate-to-severe atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease. Interleukin (IL) 13 is a type 2 cytokine that is key to the inflammation underlying AD. Tralokinumab is a first-in-class, fully human, monoclonal antibody that specifically binds with high affinity to IL-13, neutralizing it in AD. Immunomodulatory treatments may impair vaccine-induced immune responses. OBJECTIVE: Assess the immune responses to standard vaccines in adults with moderate-to-severe AD who are undergoing treatment with tralokinumab. METHODS: ECZema TRAlokinumab Trial No. 5 (ECZTRA 5; NCT03562377) was a phase 2, double-blind, randomized, placebo-controlled trial taking place over 30 weeks. Eligible adults were randomized 1:1, with 107 patients receiving tralokinumab 300 mg and 108 patients receiving a placebo every 2 weeks for 16 weeks. All patients received Tdap (tetanus/diphtheria/pertussis) and meningococcal vaccines at week 12. The primary end points were positive antitetanus and antimeningococcal responses between weeks 12 and 16 (noninferiority margin, -25%; responder, >3-fold increase in IgG). RESULTS: The noninferiority of tralokinumab versus placebo for immune response to Tdap (91.9% vs 96.1%) and meningococcal (86.0% vs 84.2%) vaccines was demonstrated at week 16. During treatment, the rates of adverse events were lower for tralokinumab than for the placebo, with most events being mild or moderate. LIMITATIONS: Responses to other vaccines (including influenza) were not examined. CONCLUSIONS: Treatment with tralokinumab 300 mg every 2 weeks did not affect immune responses to Tdap and meningococcal vaccines. Treatment was well tolerated when administered concomitantly with the vaccines and demonstrated a safety profile comparable to phase 3 trials.

Inhibition of IL-13: A New Pathway for Atopic Dermatitis [Formula: see text].

Dupilumab, a monoclonal antibody against the common receptor of interleukin (IL)-4 and IL-13, was the first biologic therapy approved in Canada for treatment of moderate-to-severe atopic dermatitis (AD). While it is considered safe and effective, dupilumab is not universally effective and 8%-38% of patients develop conjunctivitis, while some patients develop head and neck dermatitis. Thus, new therapeutic options are warranted. While both IL-4 and IL-13 play important roles in the pathogenesis of AD, it has been recently demonstrated that IL-13 is the primary upregulated cytokine in AD skin biopsy samples. A placebo-controlled phase 2b clinical trial evaluating the efficacy and safety of lebrikizumab, an IL-13 inhibitor, in AD demonstrated that, at 16 weeks, Eczema Area and Severity Index (EASI) 75 and Investigator's Global Assessment (IGA) 0/1 were achieved by 60.6% and 44.6% of patients taking lebrikizumab at its highest dose (vs 24.3% and 15.3% of patients taking placebo, respectively). Moreover, treatment with lebrikizumab was associated with rapid improvement of pruritus and low rates of conjunctivitis (1.4%-3.8%). Another IL-13 monoclonal antibody, tralokinumab, was evaluated for safety and efficacy in moderate-to-severe AD. By week 12, among adults receiving 300 mg tralokinumab, 42.5% achieved EASI-75 and 26.7% achieved IGA 0/1 score (vs 15.5% and 11.8% in the placebo group, respectively). Both lebrikizumab and tralokinumab demonstrated acceptable safety profiles in AD (and non-AD) trials with adverse events often being comparable between treatment and control groups. Thus, IL-13 inhibitors may provide a safe and effective treatment alternative for patients with moderate-to-severe AD.

Treatment of atopic dermatitis with tralokinumab, an anti-IL-13 mAb.

BACKGROUND: IL-13 has an important role in atopic dermatitis (AD) pathogenesis. Tralokinumab is a fully human mAb that potently and specifically neutralizes IL-13. OBJECTIVE: We sought to evaluate the efficacy and safety of tralokinumab in adults with moderate-to-severe AD. METHODS: In this phase 2b study (NCT02347176), 204 adults were randomized 1:1:1:1 to receive 45, 150, or 300 mg of subcutaneous tralokinumab, or placebo, every 2 weeks for 12 weeks with concomitant topical glucocorticoids. Coprimary end points were change from baseline in Eczema Area Severity Index score and percentage of participants with an Investigator's Global Assessment response (0/1 score and reduction of ≥2 grades from baseline) at week 12. RESULTS: At week 12, 300 mg of tralokinumab significantly improved change from baseline in Eczema Area Severity Index score versus placebo (adjusted mean difference, -4.94; 95% CI, -8.76 to -1.13; P = .01), and a greater percentage of participants achieved an Investigator's Global Assessment response (26.7% vs 11.8%). Greater responses were found in participants with greater concentrations of biomarkers of increased IL-13 activity. Participants treated with 300 mg of tralokinumab demonstrated improvements in SCORAD, Dermatology Life Quality Index, and pruritus numeric rating scale (7-day mean) scores versus placebo. Upper respiratory tract infection was the most frequent treatment-emergent adverse event reported as related to study drug in the placebo (3.9%) and pooled tralokinumab (3.9%) groups. CONCLUSIONS: Tralokinumab treatment was associated with early and sustained improvements in AD symptoms and an acceptable safety and tolerability profile, thereby providing evidence for targeting IL-13 in patients with AD.

Understanding the immune landscape in atopic dermatitis: The era of biologics and emerging therapeutic approaches.

Atopic dermatitis (AD) is a chronic, systemic, inflammatory disease that affects the skin and is characterized by persistent itch and marked redness. AD is associated with an increased risk of skin infections and a reduced quality of life. Most AD treatment options to date were not designed to selectively target disease-causing pathways that have been established for this indication. Topical therapies have limited efficacy in moderate-to-severe disease, and systemic agents such as corticosteroids and immunosuppressants present with tolerability issues. Advances in the understanding of AD pathobiology have made possible a new generation of more disease-specific AD therapies. AD is characterized by the inappropriate activation of type 2 T helper (Th2) cells and type 2 innate lymphoid (ILC2) cells, with a predominant increase in type 2 cytokines in the skin, including interleukin (IL)-13 and IL-4. Both cytokines are implicated in tissue inflammation and epidermal barrier dysfunction, and monoclonal antibodies targeting each of these interleukins or their receptors are in clinical development in AD. In March 2017, dupilumab, a human anti-IL-4Rα antibody, became the first biologic to receive approval in the United States for the treatment of moderate-to-severe AD. The anti-IL-13 monoclonal antibodies lebrikizumab and tralokinumab, which bind different IL-13 epitopes with potentially different effects, are currently in advanced-stage trials. Here, we briefly review the underlying pathobiology of AD, the scientific basis for current AD targets, and summarize current clinical studies of these agents, including new research to develop both predictive and response biomarkers to further advance AD therapy in the era of precision medicine.

Relative efficacy of systemic treatments for atopic dermatitis.

BACKGROUND: Systemic medications are often required for severe atopic dermatitis (AD) refractory to topical therapies. Biologic medications are a recent advancement in the field and a comparison with standard systemic approaches would be beneficial. OBJECTIVE: To compare efficacies of systemic therapies for the treatment of AD. METHODS: A systematic literature review was performed using Medline, Ovid, and Embase. Randomized controlled trials looking at the efficacy of systemic treatments for AD in adults and children were included. RESULTS: A total of 41 studies met criteria and were included in our final analysis. Consistent improvements in Eczema Area and Severity Index and Scoring Atopic Dermatitis were reported with dupilumab and cyclosporine. Phase 2 clinical trials for lebrikizumab and tralokinumab were effective and would benefit from phase 3 trials. No study reported efficacy of biologic medications in pediatric patients; however, cyclosporine improved clinical severity by the greatest amount in this group. LIMITATIONS: A lack of well controlled comparison studies make direct comparisons between the treatments difficult. CONCLUSION: For treatment of severe AD, the strongest evidence currently exists for dupilumab and cyclosporine at improving clinical disease severity. Further research is required to determine long-term safety and efficacy of biologic medications.

Application of structured statistical analyses to identify a biomarker predictive of enhanced tralokinumab efficacy in phase III clinical trials for severe, uncontrolled asthma.

BACKGROUND: Tralokinumab is an anti-interleukin (IL)-13 monoclonal antibody investigated for the treatment of severe, uncontrolled asthma in two Phase III clinical trials, STRATOS 1 and 2. The STRATOS 1 biomarker analysis plan was developed to identify biomarker(s) indicative of IL-13 activation likely to predict tralokinumab efficacy and define a population in which there was an enhanced treatment effect; this defined population was then tested in STRATOS 2. METHODS: The biomarkers considered were blood eosinophil counts, fractional exhaled nitric oxide (FeNO), serum dipeptidyl peptidase-4, serum periostin and total serum immunoglobulin E. Tralokinumab efficacy was measured as the reduction in annualised asthma exacerbation rate (AAER) compared with placebo (primary endpoint measure of STRATOS 1 and 2). The biomarker analysis plan included negative binomial and generalised additive models, and the Subgroup Identification based on Differential Effect Search (SIDES) algorithm, supported by robustness and sensitivity checks. Effects on the key secondary endpoints of STRATOS 1 and 2, which included changes from baseline in standard measures of asthma outcomes, were also investigated. Prior to the STRATOS 1 read-out, numerous simulations of the methodology were performed with hypothetical data. RESULTS: FeNO and periostin were identified as the only biomarkers potentially predictive of treatment effect, with cut-offs chosen by the SIDES algorithm of > 32.3 ppb and > 27.4 ng/ml, respectively. The FeNO > 32.3 ppb subgroup was associated with greater AAER reductions and improvements in key secondary endpoints compared with the periostin > 27.4 ng/ml subgroup. Upon further evaluation of AAER reductions at different FeNO cut-offs, ≥37 ppb was chosen as the best cut-off for predicting tralokinumab efficacy. DISCUSSION: A rigorous statistical approach incorporating multiple methods was used to investigate the predictive properties of five potential biomarkers and to identify a participant subgroup that demonstrated an enhanced tralokinumab treatment effect. Using STRATOS 1 data, our analyses identified FeNO at a cut-off of ≥37 ppb as the best assessed biomarker for predicting enhanced treatment effect to be tested in STRATOS 2. Our findings were inconclusive, which reflects the complexity of subgroup identification in the severe asthma population. TRIAL REGISTRATION: STRATOS 1 and 2 are registered on ClinicalTrials.gov ( NCT02161757 registered on June 12, 2014, and NCT02194699 registered on July 18, 2014).