PP-016 Effect of tacrolimus eye drops on human primary corneal epithelial cells

Background Ensuring the safety of products made in pharmacy departments is an essential parameter in the development of any ophthalmic formulation. Purpose The aim of this study was to show the cytotoxic effect of tacrolimus 0.03 mg/mL eye drops on human primary corneal epithelial cells using real time monitoring of dynamic changes based on bioimpedance measurements induced by cell–toxicant interactions. Material and methods Initially, tacrolimus 0.03 mg/mL eye drops (TED) were prepared as a pharmaceutical compound in the department of pharmacy. The effect of the TED on the viability of cells was studied on ATCC normal human primary corneal epithelial cells. We used the xCELLigence real time cell analyser system (ACEA Biosciences, San Diego, California, USA) for monitoring the growth of cell cultures in real time. The cell index , based on the measured electric impedance across the cell culture, was used to represent the number of cells. 3000 cells/well (16 wells E-plates) were incubated for 20 hours. Subsequently, the original culture medium was aspirated and different TED concentrations (7.5 µg/mL, 15 µg/mL, 22 µg/mL, 30 µg/mL, 37 µg/mL and 45 µg/mL) were added to different wells. The results are represented as dose response curves versus time, and IC50 was calculated in different times. Results Surviving rate kinetic curves showed that TED induced a gradual decline in the cell surviving rate over a 20 hour exposure period. This behaviour suggests that TED cause significant corneal cellular toxicity. An analysis of the kinetic curve of the cell surviving rate showed that these effects were time and dose dependent. The IC50 at 30 min was 0.0139 mg/mL, at 4 hours 0.0125 mg/mL and at 16 hours 0.00836 mg/mL. Conclusion These results may be particularly relevant to estimate the optimal TED concentration in clinical situations to avoid a toxic effect. References and/or acknowledgements Acknowledgements: Fundacion Mutua Madrilena and Fundacion Espanola Farmacia Hospitalaria. No conflict of interest