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IN VITRO ALTERNATIVES IN INHALATION TOXICOLOGY – WHERE DO WE START?

Greenwell1, L., Roggen2, E., Holder3, J., Westmoreland1, C., Cathew1, P., Fentem1, J. 1Safety and Environment Assurance Centre, Unilever Colworth, Sharnbrook, Bedford, U.K., 2Novozyme A/S, Smoermoseve, Bagsvaerd, Denmark, 3Glaxo-Smith-Kline, Ware, Hertfordshire, U.K.
Abstract

This study by researchers at Unilever’s Safety and Environment Assurance Centre (SEAC), Novozyme A/S and GlaxoSmithKline demonstrated that MatTek’s EpiAirway in vitro human tracheal/bronchial tissue equivalent is sufficiently in vivo-like to be considered as an in vitro Replacement (one of the 3 R’s) for use in inhalation toxicology studies. The 7th Amendment to EU directive 76/768/EEC stipulates that alternatives to repeat-dose toxicity animal studies must be in place by 2013. Addressing the challenges of developing a suitable battery of tests to replace in vivo inhalation testing is no simple task. A multitude of questions must be answered, ranging from the technically challenging ‘how do we emulate representative inhalation exposure in vitro?’ to the more ethereal ‘what exactly defines a NOAEL (“No-Observed-Adverse-Effect Level”) in vitro?’. Researchers at Unilever’s Safety and Environment Assurance Centre (SEAC), Novozyme A/S, and GlaxoSmithKline are currently investigating a range of approaches encompassing both ‘top down’ bridging studies and ‘bottom up’ investigative studies. These include the use of various models (e.g. mono- and co-culture of primary and immortalised cells), in silico systems (e.g. QSAR modelling) and leading edge technologies (e.g. the application of ‘omic technologies to risk assessment). MatTek’s EpiAirway™ Human Tissue Equivalent, a 3-D model of respiratory tract tissue, is being assessed as a Replacement (one of the “3R’s”) for existing in vivo techniques. One of the major issues in inhalation toxicology is the vast array of end-points currently used in toxicological assessment. These pose a variety of issues, such as consideration of the complex cell signalling events occurring between different tissue types in vivo (e.g. free cell recruitment and activation) and reproduction of long-term endpoints (e.g. initiation of fibrosis). The fact that the development of a single model suitable to assess all endpoints is unlikely, it is more conceivable that a barrage of tests will be necessary to fully characterise the toxicology of novel ingredients. Progress is being made – markers of specific endpoints are being characterised in vitro, and efficacy and limitations of different models are being assessed, along with their comparability to observed results in vivo. Research is being initiated into representative aerosol exposure in vitro and the development of mutually available databases.

Keywords

3R’s, 7th Amendment to the Cosmetics Directive, Aerosol dosimetry, Cell signaling, EU 76/768/EEC, EpiAirway, In vivo inhalation, Inhalation toxicology, International partnership for alternatives to animal testing (IPAAT), No observed adverse effect level (NOAEL), Omic Analysis, Respiratory toxicology , Screening tool aerosol exposure

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