THU0193 Determination of the potential for an<i>in vitro</i>protein binding interaction between the leflunomide (lef) metabolite, a77 1726 and the sulfasalazine (ssz) metabolite, sulfapyridine (sp)
Authors: DL Larsen, J Holsapple
Journal: Annals of the Rheumatic Diseases (2001)

<h3>Background</h3> LEF is a relatively new DMARD licensed for the treatment of rheumatoid arthritis (RA). When orally administered, it is rapidly and almost completely converted into its active metabolite, A77 1726, which is highly bound to plasma proteins (&gt;99%). A77 1726 may therefore be subject to interactions with other highly bound drugs. SSZ is also a DMARD used in the treatment of RA. It is metabolised to 5-aminosalicylate (5-ASA) and SP with the latter thought to be responsible for the toxicity of SSZ. SP may also bind to plasma protein, thus there is a possibility of a protein binding interaction. Competitive displacement of bound drug molecules from the binding site may alter the pharmacokinetics of A77 1726 or SP and possibly alter the tolerance of a combined regimen. <h3>Objectives</h3> This study investigated the potential for a protein binding interaction between the LEF metabolite, A77 1726 and the SSZ metabolite, SP. <h3>Methods</h3> The free and bound fractions of 14C-labelled A77 1726 and 14C-labelled SP were determined by dialysis of drug fortified human plasma against phosphate buffer (pH 7.4) at 37 degrees celcius. The extent of equilibration was determined after 4, 6, 8, 10 and 12 h of dialysis and the time to equilibration was evaluated at concentrations of 10 and 50μg/ml for both A77 1726 and SP. The extent of binding of 14C-labelled A77 1726, alone, and in the presence of 10, 20 and 50μg/ml of non-labelled SP was determined at nominal A77 1726 concentrations of 10, 20 and 50μg/ml. Conversely the extent of 14C-labelled SP binding alone, or in the presence of 10, 20 and 50μg/ml of non-labelled A77 1726 was determined at nominal SP concentrations of 10, 20 and 50μg/ml. <h3>Results</h3> No time-dependent changes in A77 1726 and SP free fractions were observed over the 4 to 12-hour sampling period so a dialysis time of six hours was chosen for the subsequent interaction experiments. Under study conditions, 14C-labelled A77 1726 was 99.2% to 99.3% protein-bound over the concentration range 10 to 50μg/ml. The free fractions of A77 1726 tended to be slightly greater at 50μg/ml (0.78%) than at 10 or 20μg/ml (0.73% and 0.74%, respectively). Addition of SP to the plasma had no effect on the observed binding of A77 1726. In contrast to A77 1726, 14C-labelled SP was only 24 to 28% bound over a concentration range of 10 to 50μg/ml. Free fractions of SP were relatively constant (73% to 76%) as the concentration was increased from 10 to 50μg/ml. Addition of A77 1726 to plasma to provide concentrations of 10, 20 and 50μg/ml had no effect on the observed binding of SP. In addition, the free fractions of SP did not change (72?76%) in the presence of 0, 10, 20 and 50μg/ml A77 1726. <h3>Conclusion</h3> Free fractions of both A77 1726 and SP did not change when each underwent dialysis in the presence of the other at concentrations of 10, 20 and 50μg/ml. The low binding of SP in human plasma suggests there is a low potential for a protein binding interaction between A77 1726 and SP over the range of concentrations examined. A clinical study to investigate the pharmacokinetics of A77 1726 and SP in healthy subjects is underway.