title: Validation study on the Ocular Irritection assay for eye irritation testing.
abstract: Both a prospective and a retrospective validation study were undertaken to assess the suitability of the Ocular Irritection assay to discriminate ocular hazards as defined by the OECD and UN Globally Harmonized System (UN GHS) for classification. The primary focus of the study was to evaluate the usefulness of the Ocular Irritection assay to reliably discriminate chemicals not requiring classification (UN GHS non-classified), from classified chemicals (UN GHS Categories 1 and 2). Furthermore a post-hoc evaluation was carried out to evaluate the usefulness of the assay to discriminate chemicals inducing serious eye damage (UN GHS Category 1) from other classes. The prospective validation study was conducted between 2009 and 2012 following internationally agreed principles. A set of 56 coded test chemicals for which quality and/or peer-reviewed in vivo data were available were used to obtain prospective data on the assay's reliability (reproducibility within and between laboratories) and relevance (predictive capacity). The assay showed good within-laboratory variability, transferability including to a naïve laboratory, and between-laboratory concordance of classifications (82% for the discrimination of non-classified from classified chemicals, and 83% for the discrimination of Category 1 from other classes). The obtained prospective data were then combined with existing data on the Ocular Irritection collected from various sources, totaling 88 chemicals with parallel in vivo and in vitro data to obtain a comprehensive assessment of the test method performances. The enlarged dataset comprised 43 non-classified, 25 Category 2 and 20 Category 1 chemicals according to the UN GHS classification. When used for the identification of UN GHS non-classified versus classified materials (based on the existing cut-off of 12.5) the Ocular Irritection assay showed an overall a sensitivity of 93% and a specificity of 58%. An evaluation on possible reasons for misclassification identified some organic functional groups (acrylate, carboxamide and cycloalkene) to correlate with the observed mispredictions. If these functional groups were excluded from the Ocular Irritection applicability domain, the obtained dataset (n=79 chemicals distributed as 41 UN GHS Classified and 38 Non-Classified chemicals) had an overall sensitivity of 98%, and specificity of 63%, which is in line with currently adopted test methods. When used for the identification of UN GHS Category 1 versus other categories (based on the existing cut-off of 30.0) the Ocular Irritection assay showed an overall specificity of 81% and a sensitivity of 50% which is again in line with currently adopted test methods. The Ocular Irritection assay appeared therefore as a useful test method to predict chemicals not requiring classification for eye hazards according to the UN GHS classification system. Furthermore the method was found suitable to identify serious/irreversible eye damage (UN GHS Category 1). The detailed documentation and results of the study have been submitted to an internationally recognized validation centre for peer-review.
authors.affiliation: SeCAM Services & Consultation on Alternative Methods, Agno, Switzerland. Electronic address: chantra.eskes@secam-ce.eu.
authors.forename: Chantra
authors.initials: C
authors.lastname: Eskes
authors.affiliation: seh consulting + services, Paderborn, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Hoffmann
authors.affiliation: INT.E.G.RA, Milan, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: Facchini
authors.affiliation: InVitro International, Irvine, USA.
authors.forename: Rich
authors.initials: R
authors.lastname: Ulmer
authors.affiliation: InVitro International, Irvine, USA.
authors.forename: Amy
authors.initials: A
authors.lastname: Wang
authors.affiliation: SeCAM Services & Consultation on Alternative Methods, Agno, Switzerland.
authors.forename: Manuela
authors.initials: M
authors.lastname: Flego
authors.affiliation: FAR.CO.S., Garbagnate Milanese, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Vassallo
authors.affiliation: ResPharma, Milano, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Bufo
authors.affiliation: SeCAM Services & Consultation on Alternative Methods, Agno, Switzerland.
authors.forename: Erwin
authors.initials: E
authors.lastname: van Vliet
authors.affiliation: INT.E.G.RA, Milan, Italy.
authors.forename: Federica
authors.initials: F
authors.lastname: d'Abrosca
authors.affiliation: IIVS, Gaithersburg, USA.
authors.forename: Nathan
authors.initials: N
authors.lastname: Wilt
mesh_terms: D000826:Animal Testing Alternatives; D001681:Biological Assay; D003315:Cornea; D005123:Eye / Q000187:drug effects*; D005136:Eye Proteins / Q000378:metabolism; D007509:Irritants / Q000145:classification / Q000633:toxicity*; D008954:Models, Biological; D011489:Protein Denaturation; D015203:Reproducibility of Results; D023382:Toxicity Tests, Acute / Q000379:methods*
chemical_list: D005136:Eye Proteins; D007509:Irritants
keywords: Eye hazards; Ocular Irritection®; Regulatory testing; Validation