In vitro testing methods

The effect of the device on air flow resistance and the ability of patients to achieve the necessary air flow rate for proper use of the product should be considered. The in vitro test method should also take this into account since the results obtained may be dependent on air flow characteristics. 

Brandon, E.F., Raap, C.D., Meijerman, I. etal. (2003) An update on in vitro test methods in human hepatic drug biotransformation research pros and cons. Toxicology and Applied Pharmacology, 189 (3), 233-246. 

Gad, S.C. (1989b). A tier testing strategy incorporating in vitro testing methods for pharmaceutical safety assessment. Flumane Innovations and Alternatives in Animal Experimentation 3 75-79. 

Gad, S.C. (1989). A Tier Testing Strategy Incorporating In Vitro Testing Methods for Pharmaceutical Safety Assessment. Humane Innovations Alternatives Anim. Exp. 3 75-79. 

There are adopted in vitro test methods (Section 4.5.3.2) under which a substance can be identified as corrosive. A negative result in these tests should, however, be supported by a weight of evidence assessment using other data. It should be noted that these tests do not provide information on skin irritation. 

ECVAM is the leading international center for alternative test method validation. Hartung et al. (29) summarized the modular steps necessary to accomplish stage 3 (test validation). The seven modular steps are (I) test definition, (2) within-laboratory variability, (3) transferability, (4) between-laboratory variability, (5) predictive capacity, (6) applicability domain, and (7) performance standards (29). Steps 2-4 evaluate the test s reliability steps 5 and 6 evaluate the relevance of the test. Successful completion of all seven steps is necessary to proceed to stage 4 (independent assessment or peer review). This modular approach allows flexibility for the validation process where information on the test method can be gathered either prospectively or retrospectively. The approach is applicable not only to in vitro test methods but also to in silico approaches (e.g., computer-based approaches such as quantitative structure-activity relationships or QSAR) and pattern-based systems (e.g., genomics and proteomics). 

The reported incidence of resistance to these drugs varies greatly, from less than 5% to 75%. In part this tremendous variation in incidence reflects the definition of resistance (recurrent thrombosis while on antiplatelet therapy vs in vitro testing), methods by which drug response is measured, and patient compliance. Several methods for testing aspirin and clopidogrel resistance in vitro are now FDA-approved however, their utility outside of clinical trials remains controversial. 

Application of in vitro test methods have become advantageous in specific cases, such as structurally defined compounds and delayed neuropathy, since target cell data and biochemical processes associated in delayed neuropathy are known. Microscopic studies reveal that cases of OPIDN have degeneration of axons followed by demyelination of the nervous system.25,26 Epidemiologic studies have indicated mild impairment of the brainstem, spinal cord, and peripheral nerve functions in Gulf War veterans.27 Such studies are consistent with the spectrum of OPIDN syndrome. The main nerve agents have been shown to inhibit NTE in vitro as well as in vivo. Sarin has been shown to produce delayed neurotoxicity when administered at higher doses in protected hens.25-27... 

National Institutes of Health (NIH), Corrositex an in vitro test method for assessing dermal corrosivity potential of chemicals, NIH Publication No. 99-4495, National Toxicology Program (NTP) Interagency Center for the Evaluation of Alternative Toxicological Methods (NICEATM), Research Triangle Park, North Carolina, 1999. 

Birkett DJ, Mackenzie PI, Veronese ME, Miners JO (1993) In vitro approaches can predict human drug metabolism. Trends Pharmacol Sci 14 292-294 Bloomer JC, Boyd HF, Hickey DMB et al. (2001) 1-(Arylpiperazinylamidoalkyl)-pyrimidones Orally Active Inhibitors of Lipoprotein-Associated Phospholipase A2. Bioorg Med Chem Lett 11 1925-1929 Brandon EFA, Raap CD, Meijerman I et al. (2003) An update on in vitro test methods in human hepatic drug biotransformation research pros and cons. Toxicol Applied Pharmacol 189 233-246... 

No formal international validation study of the in vitro methodology has been performed. Importantly, the same applies however to the in vivo studies. Despite the publication of several well-conducted comparative studies (Franz, 1975 Yang et al., 1986a,b Grissom et al, 1987 Scott and Ramsey, 1987 Hotchkiss et al., 1990, 1992 Scott et al., 1992 Ramsey et al., 1994 Roper et al., 1995 Dick et al., 1997a,b Wester et al., 1998), the Organization for Economic Cooperation and Development (OECD) concluded in 2000 that evaluation of in vitro test methods by means of data available from the public literature is very difficult, because these studies, comparing in vitro and in vivo test results, contained too many variables (different species, thickness and types of the... 

Predictive in vitro test methods are as well developed as nonanimal test alternatives, including specific cell-based assays.22 Such cellular assays include the culture of keratinocytes that represent very often the first cells in the skin to encounter potential reactive chemicals. Purified keratinocytes are cultured with a specific test chemical, and the production of proinflammatory cytokines or chemokines are then measured. Another cell population that can be used to test for chemicals inducing contact allergy are Langerhans cells (LCs). LCs are the... 

New in vitro test methods target the behavior of macromolecules, cells, tissues, and organs in well defined methods, which control experimental conditions and standardize experimentation. These tests provide more reproducible, rapid, and cost-effective results. In addition, more information at a basic mechanistic level can be obtained from these tests. Table 3 provides a summary of current test systems. 

Current research and development efforts have focused on the use of more biocompatible coatings to reduce the biological response of both intravascular and subcutaneous devices. These efforts are based on the expectation that such developments wfllbe critical to the ultimate success in developing implanted sensors that yield continuous analytical results that match closely with conventional in vitro test methods. One new approach in this direction employs novel nitric oxide (NO) release polymers to coat the surface of intravascular sensors.The potent antiplatelet activity of NO has been shown to greatly reduce the formation of thrombus on the surface of implantable electrochemical oxygen sensing catheters, and yield much more accurate continuous PO2 values in animal experiments. 

A Bottom-Up approach, which starts with using in vitro test methods that can accurately identify chemicals not classified for eye hazards according to the UN Globally Harmonized System (GHS) and the EU Classification, Labelling, and Packaging (EU CLP) system [15, 16]. 

Besides the test methods described in Section 3 as being able to identify non-classified materials (BCOP, ICE and CM), a number of advanced in vitro test methods have been proposed for the identification of non-classified materials from classified ones (GHS Cat. 1 and Cat. 2). These methods are considered useful within integrated test strategies and in particular for initiating the bottom-up approach to accurately identify non-classified materials. 

The lack of full reversibility within 21 days of application is a critical parameter to identify substances inducing serious eye damage. If the majority of the models currently accepted from a regulatory point of view (see Section 3) may allow to predict such effects, they were not specifically designed and/or do not specifically address the reversibility/persistence of effects. This may be particularly of relevance depending upon the types of materials to be evaluated, and if in vivo irreversible effects are expected. As such, a number of in vitro test methods have been developed and optimized to distinguish persistent from reversible ocular effects. These include the use of histopathology and Depth of Injury of effects (Dol), the Porcine Corneal Ocular Reversibility Assay (PorCORA) and the Ex-Vivo Eye Irritation test (EVEIT). 

Adriaens et al. [21] also showed that persistence of effects seems to play an important role in the classification of a chemical as a GHS Cat. 1 by the in vivo method. It may thus be necessary to have in vitro test methods capable of detecting persistence of effects. 

Adriaens E., Barroso J., Eskes C., Hoffmann S., McNamee P., Alepee N., Bessou-Touya S., De Smedt A., De Wever B., Pfannenbecker U., Magalie Tailhardat M. Zuang V. (2014). Retrospective analysis of the Draize test for serious eye damage/eye irritation importance of understanding the in vivo endpoints under UN GHS / EU CLP for the development and evaluation of in vitro test methods. Archives of Toxicology 88, 701-723. 

ICCVAM (2009). Background review document—current status of in vitro test methods for identifying mild/moderate ocular irritants Hen s egg test—chorioallantoic membrane test method. National Institute of Environmental Health Sciences, Resesarch Triangle Park, North Carolina, USA. Available at http //iccvam.niehs.nih.gov/ docs/ocutox docs/InVitro-2010/AppE-HET-Front-Body.pdf. Accessed 31 July 2013... 

In this context, this chapter is meant to review the existing in vitro testing methods and update on the emerging in vitro approaches for the assessment of genotoxicity and carcinogenicity. 

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