warfarin and Hyperthyroxinemia

Two distinct genotypes that result in the amino acid substitutions R218P and R218H in subdomain 2A of human serum albumin (HSA) have been identified as the cause of familial dysalbuminemic hyperthyroxinemia (FDH). These substitutions increase the affinity of subdomain 2A for thyroxine by approximately 10-fold elevating plasma thyroxine levels in affected individuals. While many studies have examined the binding of thyroxine to FDH HSA, the binding of FDH HSA to drugs has not been widely investigated. The widely administered drug warfarin was selected as a model compound to study FDH HSA/drug interactions since it binds to subdomain 2A and its pharmacokinetics are dramatically influenced by HSA binding. Using two independent methods, fluorescence spectroscopy and equilibrium dialysis with radioactive warfarin, the binding of recombinant R218P, R218H, R218M and wild type HSA to warfarin was measured. Both methods showed an approximately 5-fold decrease in the affinity of R218P, R218H and R218M HSA for warfarin relative to wild type HSA. The Kd values determined by fluorescence spectroscopy for wild type, R218H, R218P and R218M HSA binding to warfarin were 1.35, 5.38, 5.61, and 8.34 microM, respectively. The values determined by equilibrium dialysis were 5.36, 29.5, 14.5, and 23.4 microM, respectively. Based on the above findings one would expect the free serum warfarin concentration in homozygous R218P and R218H FDH patients to be elevated about 5-fold, resulting in about a 5-fold reduction in the serum half-life of the drug.

Topics: Amino Acid Substitution; Arginine; Dialysis; Histidine; Humans; Hyperthyroxinemia; Kinetics; Mutagenesis, Insertional; Mutation; Protein Binding; Serum Albumin; Spectrometry, Fluorescence; Warfarin

The effects of pH and anionic binding inhibitors were used to test the hypothesis that the increased T4 binding affinity of the variant albumin (Alb-FDH) of familial dysalbuminemic hyperthyroxinemia (FDH) is due to an electrostatic bond with the ionized phenolic hydroxyl of the iodothyronine. As determined by charcoal adsorption from 2% serum in which binding to T4-binding globulin and transthyretin had been inhibited, increased T4 binding by Alb-FDH was pH dependent and proportional to the ionization of the phenolic hydroxyl. Increased T3 binding became apparent above physiological pH, as is consistent with the higher pK of the T3 phenolic hydroxyl. The iodothyroacetic analogs of T4 and T3 developed maximal increases in binding to Alb-FDH at about the same pH as the corresponding iodothyronines. Aspirin, salicylate, warfarin, and chloride, anions that have minimal stereochemical resemblance to the iodothyronines but bind to albumin cationic groups, inhibited T4 binding to FDH sera at concentrations that had little or no effect on binding in normal sera. Increased displacement of T4 from Alb-FDH by salicylate was also evident at therapeutic ratios to a 1:1 dilution of serum in a dialysis system. Aspirin displaced T4 at a lower pH than T3, as is consistent with competition with the ionized iodothyronine phenolic group. These findings suggest that an electrostatic bond between the iodothyronine phenolate and a cationic group on the protein is the basis for the increased affinity and specificity of Alb-FDH for T4.

Topics: Aspirin; Binding Sites; Chlorides; Female; Hot Temperature; Humans; Hydrogen-Ion Concentration; Hyperthyroxinemia; Pregnancy; Serum Albumin; Thyroxine; Triiodothyronine; Warfarin