Expression of Cytochrome P450-enzymes and Metabolism of Timolol in Human Ocular Tissue

Timolol has been used for decades to lower increased intraocular pressure (IOP). However, only limited information is available on the metabolism of the drug, especially that in the human eye. The aim of this work was to investigate the expression of drug-metabolizing cytochrome P450 (CYP) enzymes in the ocular tissues, the absorption of timolol into the aqueous humor from two commercially available products, the metabolism of timolol in the human eye and the interaction potential of timolol. The presence of CYP enzymes in the human ciliary epithelial cells at mRNA level was studied by a reverse transcriptase-polymerase chain reaction (RT-PCR) method. A dose response experiment was carried out for mRNAs of CYP1A1 and CYP1B1 with Northern blotting. The presence of CYP1B1 protein was analyzed by immunoblotting. The metabolism and interactions of timolol were investigated in incubations with human liver homogenate, microsomes and cryopreserved hepatocytes. A clinical trial was undertaken with planned cataract surgery subjects in order to measure timolol concentrations and metabolites of timolol in the aqueous humor. CYP1A1 and CYP1B1 mRNA were expressed in the human ciliary epithelial cells. CYP1B1 was also expressed at protein level, the expression being strongly induced by the known potent inducer TCDD. CYP1B1 induction is heed to be mediated by aryl hydrocarbon receptor (AHR). Timolol was confirmed to be metabolized mainly by CYP2D6 and potent CYP2D6 inhibitors, especially fluoxetine, paroxetine and quinidine, inhibited its metabolism. This inhibition may be of clinical significance in patients using ophthalmic timolol products. Low levels of CYP2D6 mRNA splice variants were expressed in the human ciliary epithelial cells and very low levels of timolol metabolites were detected in the human aqueous humor. A negligible amount of CYP2D6 protein is evidently expressed in the human ocular tissues. Timolol 0.1% eye gel leads to an aqueous humor concentration high enough to achieve a therapeutic effect. Inter-individual variation in concentrations is low and systemic safety can be