Reversal of Advanced Digitoxin Toxicity and Modification of Pharmacokinetics by Specific Antibodies and Fab Fragments

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Abstract

The effects of Fab fragments of high-affinity specific antibodies have been studied in a canine experimental model of lethal digitoxin toxicity. Selected antiserum from sheep immunized and boosted with a digoxin-serum albumin conjugate contained antibodies that cross-reacted with digitoxin with an average intrinsic association constant of 1.4 × 1010 M−1 as determined by equilibrium dialysis. Rapid second-order association kinetics (kf = 3.7 × 106 M−1 per s) and slow dissociation kinetics (kr = 1.9 × 10−4 per s) were documented for the antibody-digitoxin complex. Eight dogs given 0.5 mg/kg digitoxin intravenously developed ventricular tachycardia after 23±4 (SEM) min. Control nonspecific Fab fragments were then given. All animals died an average of 101±36 min after digitoxin administration. Another eight dogs given the same digitoxin dose similarly developed ventricular tachycardia after 28±3 min. This group then received a molar equivalent dose of specific Fab fragments intravenously over 3 min, followed by a 30-min infusion of one-third of the initial dose. All dogs survived. Conducted sinus beats reappeared 18±4 min after initial Fab infusion, and stable normal sinus rhythm was present at 54±16 min. Plasma total digitoxin concentrations increased threefold during the hour after initial Fab infusion, while plasma free digitoxin concentration decreased to less than 0.1 ng/ml. Effects on digitoxin pharmacokinetics of these Fab fragments and the antibody population from which they were derived were further investigated in a primate species. Unlike common laboratory animals previously studied, the rhesus monkey was found to have a prolonged elimination half-life, estimated at 135 and 118 h by radioimmunoassay and [3H]digitoxin measurements, respectively, similar to man and thus providing a clinically relevant experimental model. Intravenous administration of 2 mol of specific Fab fragments per mole of digitoxin 6 h after 0.2 mg of digitoxin produced a rapid 4.3-fold increase in plasma total digitoxin concentration followed by a rapid fall (t½ 4 h) accompanied by a 14-fold enhancement of urinary digitoxin excretion over control values during the 6-h period after Fab was given. Analytical studies were consistent with increased excretion of native digitoxin rather than metabolites, and the glycoside was found in equilibrium dialysis studies to be excreted in the urine in Fab-bound form. Administration of 2 mol of specific antibody binding sites per mole of digitoxin as intact IgG caused a greater and more prolonged increase in plasma total digitoxin concentration, peaking 13-fold above control levels. In contrast to the effects of Fab, however, specific IgG reduced the rate of urinary digitoxin excretion substantially below control values. We conclude that Fab fragments of antibodies with high affinity for digitoxin are capable of rapid reversal of advanced, otherwise lethal digitoxin toxicity, and are capable of reducing the plasma half-life and accelerating urinary excretion of digitoxin.

Authors

Hermann R. Ochs, Thomas W. Smith