In vitroevaluation of the effects of 4-aminopyridine on cytochrome P450 enzymes

BACKGROUND: Dalfampridine extended release tablets (dalfampridine-ER, known as prolonged-, modified, or sustained-release fampridine tablets in some countries) are approved for the improvement of walking in patients with multiple sclerosis (MS). Dalfampridine-ER is an extended release formulation of 4-aminopyridine (4-AP). Dalfampridine-ER is incorporated into MS management strategies that may include disease-modifying and symptomatic therapies. Since several symptomatic therapies are partially or fully metabolized by enzymes of the hepatic cytochrome P450 system (CYP450) it is important to evaluate drug-drug interactions through potential effects of dalfampridine-ER on CYP450. METHODS: The ability of 4-AP to inhibit CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4/5 in a direct and time-dependent manner was evaluated using pooled human liver microsomes. 4-AP concentrations were 0.03, 0.1, 0.3, 1, 3, 10, and 30 μM, representing 0.1-100-times the average plasma 4-AP concentration (30 ng/mL; 0.32 μM) at therapeutic dosing; the concentration inhibiting 50% of each enzyme activity (IC50) was determined. The ability of 4-AP (0.025, 0.25, 2.5, and 25 μM) to induce the expression of CYP1A2, 2B6, 2C9, 2C19, 2E1, and 3A4/5 enzymes was evaluated using primary cultures of freshly isolated human hepatocytes from non-transplantable livers. The enzyme-inducing effects of 4-AP were compared with the prototypical inducers. Metabolites were assayed using high-performance liquid chromatography-tandem mass spectrometry techniques. All inhibition and induction assays included positive controls. RESULTS: 4-AP did not directly inhibit CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, or CYP3A4/5, but at a concentration of 30 μM, CYP2E1 was inhibited by 12%, resulting in an estimated IC50 value of 125 μM. None of the enzymes demonstrated time-dependent inhibition by 4-AP. There was little or no effect by 4-AP on enzyme induction, with enzyme activities approximately equivalent to vehicle control. A main limitation was the inability to estimate effectiveness of 4-AP relative to prototypical CYP450 inducers. CONCLUSION: The likelihood of drug-drug interactions is remote in patients with MS who may be taking dalfampridine-ER concomitantly with medications that are metabolized by CYP450 pathways.