SYSTEMATIC REVIEW PROTOCOL FOR ANIMAL INTERVENTION STUDIES FORMAT BY SYRCLE (WWW.SYRCLE.NL) VERSION 2.0 (DECEMBER 2014) Item Section/Subsection/Item # A. General 1.

Title of the review

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Authors (names, affiliations, contributions)

Description Systematic Review and Meta-analysis of the Efficacy of Interleukin-1 Receptor Antagonist in Animal Models of Stroke: An Update Sarah K McCann, University of Edinburgh Emily S Sena, University of Edinburgh Fala Cramond, University of Edinburgh Malcolm R Macleod, University of Edinburgh

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Other contributors (names, affiliations, contributions) Contact person + e-mail address

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Funding sources/sponsors

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Conflicts of interest Date and location of protocol registration Registration number (if applicable) Stage of review at time of registration Literature search and first screening phase complete B. Objectives Background Interleukin-1 receptor antagonist (IL-1 RA) has been identified as a candidate stroke drug. We previously carried out a systematic review and meta-analysis of the preclinical literature assessing the efficacy of IL-1 RA in ischaemic stroke (Banwell et al 2009), identifying several What is already known about this weaknesses in these data. Preclinical research into IL-1 RA disease/model/intervention? Why is it has continued in the 10 years since our original literature important to do this review? search and early phase clinical trials in both ischaemic and haemorrhagic stroke patients have been initiated. This timely update will establish the current range and quality of preclinical evidence supporting the translation of IL-1 RA into clinical application. Research question Specify the disease/health problem of interest Focal ischaemic stroke Specify the population/species All non-human animals studied Specify the intervention/exposure IL-1 RA Specify the control population Animals receiving vehicle or no treatment

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Emily Sena: [email protected] National Centre for the Replacement, Refinement & Reduction of Animals in Research None

Check for approval

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Specify the outcome measures State your research question (based on items 11-15) C. Methods Search and study identification Identify literature databases to search (e.g. Pubmed, Embase, Web of science)

Primary outcome measure: infarct area or volume Secondary outcome measures: neurobehavioural score and mortality What is the effect of IL-1 RA on infarct size in animal models of focal ischaemic stroke?

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MEDLINE via PubMed Web of Science EMBASE Other, namely: BIOSIS

[(interleukin 1 receptor antagonist) OR (IL-1 RA) OR (IL1 RA) OR (IL1-RA) OR (Anakinra)] AND [(stroke) OR (ischemia) OR (cerebrovascular) OR (middle cerebral artery) OR (MCA) OR (ACA) OR (anterior cerebral artery) OR (MCAO)] AND ( Hooijmans et al 2010 Pubmed animal filter OR de Vries et al 2014 Embase animal filter update) NOT (coronary OR myocardial)

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Define electronic search strategies (e.g. use the step by step search guide15 and animal search filters20, 21)

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Identify other sources for study identification

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Define search strategy for these other sources

(a) 2 independent reviewers will screen reference lists of studies included in the review update (b) Personal communication

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Study selection Define screening phases (e.g. prescreening based on title/abstract, full text screening, both)

Reference lists of included studies Other, namely: Stuart Allan

(a) Screening by title and abstract using the SyRF screening application (b) Full text screening (a) First screen requires 0.66 agreement completed by up to 3 reviewers Specify (a) the number of reviewers (b) Second screen will be completed independently per screening phase and (b) how by 2 reviewers with discrepancies resolved discrepancies will be resolved through discussion and if necessary consultation with a third reviewer Define all inclusion and exclusion criteria based on: Inclusion criteria: Any primary study comparing treatment and control groups Type of study (design) Exclusion criteria: No control group, review, protocol paper, editorial Inclusion criteria: Whole live animal models of ischaemic occlusive stoke of the middle cerebral or anterior cerebral Type of animals/population (e.g. age, arteries or their branches gender, disease model) Exclusion criteria: Models of haemorrhagic stroke, global ischaemia; in vitro studies Inclusion criteria: Any mode of delivery of IL-1 RA (e.g. Type of intervention (e.g. dosage, transgenic, viral, peripheral) at any time point and timing, frequency) frequency Exclusion criteria: None

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Outcome measures

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Language restrictions

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Publication date restrictions

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Other

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Sort and prioritize your exclusion criteria per selection phase

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Inclusion criteria: Infarct, neurobehavioural or mortality outcomes where the mean, variance and number of animals per group is reported or can be calculated Exclusion criteria: Outcomes missing critical information e.g. variance or number of animals per group that cannot be obtained through contacting authors; outcomes reported as medians Inclusion criteria: All languages Exclusion criteria: None Inclusion criteria: Studies published 2006 – 2016 that were not included in our original review Exclusion criteria: Studies published pre-2006 Inclusion criteria: Exclusion criteria: Selection phase: screening title and abstract 1. No focal ischaemia model 2. No IL-1 RA treatment 3. In vitro model 4. No original data (e.g. review article)

Selection phase: full text screening 1. Exclusion criteria listed above 2. No infarct quantification or neurobehavioural outcomes 3. No control group 4. Outcome reported as median 5. No variance or number of animals per group reported or retrieved through contacting authors Study characteristics to be extracted (for assessment of external validity, reporting quality) Author (first, corresponding), year, journal, DOI, Study ID (e.g. authors, year) publication type, source of funding Number of animals per group - if the exact number per Study design characteristics (e.g. group is not reported (but for example only a range) the experimental groups, number of lowest number of animals will be used for data analysis; animals) number of experimental groups served by a single control group; co-treatments Species, strain, sex, age, weight, anaesthetic for model induction, use of ventilation, method of model induction, Animal model characteristics (e.g. type of model (permanent, temporary), duration of species, gender, disease induction) ischaemia, co-morbidity, injury confirmation, monitoring of physiological variables Mode of delivery (transgenic, viral, peripheral), dose, unit, Intervention characteristics (e.g. single/multiple administrations, time and number of intervention, timing, duration) administrations, route of delivery, co-treatments Type, time of assessment, mean, variance – where it is unclear if a measure of variance is SD or SEM, the variance will be recorded as SEM (the more conservative measure); Outcome measures where multiple time points are reported for the same animals, the latest time point will be extracted as this has the most clinical relevance; were infarct outcomes corrected for oedema Other (e.g. drop-outs)

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Assessment risk of bias (internal validity) or study quality Specify (a) the number of reviewers (a) 2 reviewers will independently assess study assessing the risk of bias/study quality quality in each study and (b) how (b) Discrepancies will be resolved through discussion discrepancies will be resolved and if necessary a third reviewer will be consulted By use of the CAMARADES study quality checklist, adapted as follows:  Peer reviewed publication  Control of temperature  Randomization to treatment or control  Blinded induction of ischemia Define criteria to assess (a) the  Blinded assessment of outcome internal validity of included studies  Avoidance of anaesthetics with marked intrinsic (e.g. selection, performance, neuroprotective properties  Use of animals with comorbidities [e.g., detection and attrition bias) and/or hypertension or diabetes] (b) other study quality measures (e.g.  Sample size calculation reporting quality, power)  Statement of compliance with animal welfare requirements  Statement of possible conflicts of interest  Pre-specified inclusion and exclusion criteria  Reporting of animals excluded from analysis  Source of study funding reported Collection of outcome data For each outcome measure, define Primary outcome: Infarct area or volume reported on a the type of data to be extracted (e.g. continuous scale in any unit of measurement continuous/dichotomous, unit of Secondary outcomes: i) Neurobehavioural outcomes measurement) reported on any scale ii) Mortality Data will be extracted from text or tables. Where data are presented only in graphical format, digital ruler software Methods for data extraction/retrieval will be used. Where data cannot be extracted digitally or (e.g. first extraction from graphs using critical data are missing, authors will be contacted. a digital screen ruler, then contacting Additional data will be extracted from all studies included authors) in the original review where necessary. All data will be extracted to the CAMARADES Meta-analysis Data Manager (Microsoft Access). (a) 2 reviewers will independently extract data (b) i) Where data extracted digitally differ by <10%, Specify (a) the number of reviewers an average of the two values will be taken ii) Data extracting data and (b) how differing by >10% and any other discrepancies will discrepancies will be resolved be resolved through discussion and if necessary a third reviewer will be consulted Data analysis/synthesis For each outcome, unique data from the updated search Specify (per outcome measure) how will be combined with data from the original review and you are planning to combine/compare summarised descriptively. A minimum 30% increase in the the data (e.g. descriptive summary, number of independent comparisons will be required to meta-analysis) justify further meta-analysis. Specify (per outcome measure) how it For meta-analysis, at least 25 independent comparisons will be decided whether a metaper outcome measure will be required. analysis will be performed If a meta-analysis seems feasible/sensible, specify (for each outcome measure): The effect measure to be used (e.g. Infarct area or volume: normalised mean difference mean difference, standardized mean Neurobehavioural score: normalised mean difference difference, risk ratio, odds ratio) Mortality: odds ratio

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The statistical model of analysis (e.g. random or fixed effects model)

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The statistical methods to assess heterogeneity (e.g. I2, Q)

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Which study characteristics will be examined as potential source of heterogeneity (subgroup analysis)

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Any sensitivity analyses you propose to perform

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Other details meta-analysis (e.g. correction for multiple testing, correction for multiple use of control group)

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The method for assessment of publication bias

Final approval by (names, affiliations):

Data will be combined by weighted mean difference metaanalysis using DerSimonian and Laird random effects modelling or by weighted mean difference metaregression using REML random effects modelling. Where different neurobehavioral outcomes are reported from the same cohort of animals for the same time point, these (prenested) comparisons will be combined using fixed-effects meta-analysis (nesting) and this summary estimate will be used in the random-effects model. Statistical analysis will be performed using Stata Statistical Software: Release 13 (College Station, TX: StataCorp LP). Where sufficient data exist we will use meta-regression to investigate sources of heterogeneity. If there are insufficient data, stratified meta-analysis will be carried out. Tau2, I2, Q, and adjusted R2 statistics will be used as appropriate. Components of the study quality checklist will be analysed in the overall dataset. If meta-regression is carried out, year of publication will be investigated as a confounding factor in study quality analyses. If mode of IL1 RA delivery is a significant source of heterogeneity in the overall dataset, data will be grouped according to central or peripheral administration and all further study design analyses will be carried out in these subgroups. Study design characteristic analyses: drug dose, time of administration, route of delivery, presence of cotreatments, co-morbidities, method of induction of ischemia, type of ischaemia, anaesthetic used, use of mechanical ventilation, species and sex of animal used, outcome measure used, and interval to quantification of outcome. (a) If meta-regression is used as the primary method to investigate sources of heterogeneity, stratified meta-analysis will be carried out as a sensitivity analysis. (b) If there are sufficient data we will perform multivariate meta-regression. If stratified meta-analysis is used, stratifications will be considered in two domains: study design and study quality. Each domain will be tested at p<0.05 overall for infarct area/volume, neurobehaviour and mortality outcomes. A Holm-Bonferroni adjusted critical p value will be calculated to account for the number of parameters tested within each domain. Publication bias will be assessed with a funnel plot, Egger regression and trim and fill. SKM, MRM, ESS, University of Edinburgh

Date: 15/04/16