title: Cytochrome P450 Activity in Ex Vivo Cornea Models and a Human Cornea Construct.
abstract: The pharmacokinetic behaviors of novel ophthalmic drugs are often preliminarily investigated in preclinical studies using ex vivo animal cornea or corneal cell culture models. During transcorneal passage, topically applied drugs may be affected by drug metabolizing enzymes. The knowledge regarding the functional expression of metabolic enzymes in corneal tissue is marginal; thus, the aim of this study was to investigate cytochrome P450 activity in an organotypic three-dimensional human cornea construct and to compare it with porcine and rabbit corneas, which are commonly used ex vivo cornea models. The total cytochrome P450 activity was determined by measuring the transformation of 7-ethoxycoumarin. Furthermore, the expression of the cytochrome P450 enzyme 2D6 (CYP2D6) was investigated at the protein level using immunohistochemistry and western blotting. CYP2D6 activity measurements were performed using a d-luciferin-based assay. In summary, similar levels of the total cytochrome P450 activity were identified in all 3 cornea models. The protein expression of CYP2D6 was confirmed in the human cornea construct and porcine cornea, whereas the signals in the rabbit cornea were weak. The analysis of the CYP2D6 activity indicated similar values for the human cornea construct and porcine cornea; however, a distinctly lower activity was observed in the rabbit cornea.
authors.affiliation: Institut für Pharmazeutische Technologie, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Kölln
authors.affiliation: Institut für Pharmazeutische Technologie, Technische Universität Braunschweig, Braunschweig, Germany. Electronic address: s.reichl@tu-braunschweig.de.
authors.forename: Stephan
authors.initials: S
authors.lastname: Reichl
mesh_terms: D015660:7-Alkoxycoumarin O-Dealkylase / Q000378:metabolism; D060433:Administration, Ophthalmic; D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D003315:Cornea / Q000201:enzymology*; D019389:Cytochrome P-450 CYP2D6 / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D016503:Drug Delivery Systems; D006801:Humans; D007150:Immunohistochemistry; D008954:Models, Biological; D011817:Rabbits; D013045:Species Specificity; D013552:Swine
chemical_list: D003577:Cytochrome P-450 Enzyme System; D015660:7-Alkoxycoumarin O-Dealkylase; D019389:Cytochrome P-450 CYP2D6
keywords: cell culture; cell lines; cytochrome P450; drug metabolizing enzymes; in vitro models; ophthalmic drug delivery; phase I metabolism; tissue engineering