Ocular Safety: A Silent (In Vitro) Success Story

Abstract

Ocular irritation testing has been one of the animal test methods most criticised by animal welfare advocates. Additional criticism has arisen from within the scientific community, based on the variability of the animal test results and the questionable relevance of the extremely high dose levels employed. As a result, the Draize eye irritation test has been one of the main targets for in vitro replacement. Despite extensive efforts, however, there is still no in vitro method that is fully validated as a regulatory replacement. In spite of this, many individual companies are using diverse in vitro ocular irritation tests to gain important safety and efficacy information about their products and raw materials, eliminating the need for animal testing in the process. This is done in a safe fashion by applying intelligent testing paradigms. ECVAM has played a major role in this success, through its many programmes that have emphasised the importance of understanding the true toxicological need, and then using in vitro tests to provide that information. Thus, even in the absence of a successfully validated regulatory assay, the desired result of reducing animal testing is being met.

An overview of current techniques for ocular toxicity testing

“…Despite the adoption of the GHS, Draize testing is often criticized due to its subjective and time consuming nature, lack of repeatability, variable estimates, insufficient relevance of test chemical application (Davila et al, 1998), high dosages (Curren and Harbell, 2002) and over-prediction of human responses (Jester et al, 2001), primarily due to interspecies differences. In addition, for most routine and acute toxicity tests, for example skin toxicity tests, there are standardized exposure times and/or delivery methods in place.…”

“…The test is used to provide qualitative information on the potential effects occurring in the conjunctiva following exposure to a substance, whilst evaluation of coagulation can be used to reflect potential corneal damage (NICEATM, 2006). Although CAM models are usually classified alongside ICE, BCOP and IRE models, they differ in evaluation criteria used (Barile, 2010) since they have the addition of vasculature (Curren and Harbell, 2002). The general protocol involves exposing the CAM (Fig.…”

“…Epithelial models can be constructed from animal cells (commonly SIRC cells (Ubels and Clousing, 2005)) such as in the STE test, human epidermal cells, or human corneal cells, which are usually cultured in defined medium on cell culture membranes using air-lifting techniques (Alépée et al, 2013;Cotovio et al, 2007;Kaluzhny et al, 2011;Matsuda et al, 2009) to create a 3D stratified epithelium. Cytotoxicity following topical exposure is generally used as an endpoint (Curren and Harbell, 2002), and epithelial models have the potential to identify non-classified/nonirritating substances from mild irritants (Scott et al, 2010). Time-totoxicity measurements (ET 50 ), which account for the time required for a 50% reduction in cell or tissue viability following exposure when compared to a negative control (Kaluzhny et al, 2011;Osborne et al,1995), are often used as an endpoint.…”

“…Despite the lack of formal validation, in vitro tests still are commonly used by industry. For example, industrial toxicologists often use in vitro protocols for prioritizing products and ingredients for further development (Curren and Harbell, 2002). However, use of the Draize test is still permitted worldwide, with the exception of the cosmetics section within Europe.…”

Development of a hemicornea from human primary cell cultures for pharmacotoxicology testing

“…Significant levels of variability have been observed since the test is based on a subjective (visual) scoring procedure (Weil and Scala, 1971). Besides the fact that this test causes considerable discomfort and pain to animals, it is also recognized that the response in the rabbit is not always predictive of that found in humans (Curren and Harbell, 2002). Taken together, both ethical and scientific arguments have stimulated the development and validation of several alternative in vitro methods for assessing eye irritation (Curren and Harbell, 1998;Balls et al, 1999;Worth and Balls, 2002).…”