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Actin filaments play a critical role in the normal physiology of lenticular and retinal cells in the eye. Disruption of the actin cytoskeleton has been associated with retinal pathology and lens cataract formation. Ocular toxicity is an infrequent observation in drug safety studies, yet its impact to the drug development process is significant. Recognizing compounds through screening with a potential ocular safety liability is one way to prioritize development candidates while reducing development attrition. Lens epithelial cells from human, dog, and rat origins and retinal pigmented epithelium cells from human, monkey, and rat origins were cultured and investigated with immunocytochemical techniques. Cells were treated using noncytotoxic doses of the compound, fixed, stained for actin with rhodamine phalloidin, and counterstained for nuclei with TOTO-3, followed by confocal imaging. Tamoxifen and several experimental compounds known to be in vivo lens and retinal toxicants caused a reduction in F-actin fluorescence at noncytotoxic concentrations in all cells tested as observed by confocal microscopy. Developing an assay that predicts ocular toxicity helps the development process by prioritizing compounds for further investigation. Drug-induced cytoskeletal alterations may be useful as a potential safety-screening marker of retinal and lens toxicity. The knowledge of actin molecular biology and the application of other mechanistic screens to toxicology are discussed. Reducing this work to a high-throughput platform will enable chemists to select compounds with a reduced risk of ocular toxicity.