Fate of insulin analogs in intact and nephrectomized rats determined by their receptor binding constants

Abstract

After intravenous injection of 125I-labeled human insulin and analogs in normal and nephrectomized rats, we examined their kinetic fate by Q-Sepharose separation into intact ligand, "fragments" (genuine fragments and protein-bound radioactivity), and iodide. Receptor binding association and dissociation constants (kass and kdis, respectively) of the analogs were estimated dynamically in vitro by BIAcore. The very fast disappearance of intact ligand from serum was found to be determined by 1) both kass and kdis of receptor-bearing tissue, thus substantiating our primary hypothesis; 2) elimination by kidneys, and 3) fast extravascularization. The rate of appearance of degradation products from receptor-mediated intracellular processing seems determined by kdis. With the possible exception of a truncated analog, ligand appears protected against degradation while the intracellular receptor-ligand complex remains intact. Non-receptor-mediated processing in kidneys is slow, compared with the receptor-mediated uptake and degradation of ligands with rate constants comparable to those of insulin. We observed binding of insulin and analogs putatively to serum proteins; binding capacity and affinity appeared insignificant for insulin but considerable for some analogs.

Insulin Receptor Binding Kinetics: Modeling and Simulation Studies

“…There are also investigators who have done both the kinetic and the steady-state experiments on the same preparation in order to validate their results (Gammeltoft et al, 1978;Gliemann et al, 1985;Lipkin et al, 1986a, b). In addition to these labeled-ligand studies, BIAcore surface plasmon resonance has recently been employed to dynamically measure association and dissociation rate constants in vitro (Kruse et al, 1997). Reported values for K ?…”

Mathematical modeling and analysis of insulin clearance in vivo

“…Simulated insulin concentrations for low amounts of injected insulin [ 59 ] are relatively close to the experimental data set (Figure 2 ). When examining Figure 2 on a log-linear scale, it can be seen that the relatively low absolute errors at later points in time correspond to large relative errors (not shown).…”

“…• Labeled and unlabeled insulin show the same physiological characteristics . Labeling of the insulin molecules was performed with 125 I [ 57 - 59 ]. The size of this modification is small compared to the size of the insulin molecule and should not change its binding characteristics, the effect on receptor phosphorylation, the rate of nonspecific insulin binding or the rate of renal insulin filtration.…”

“…The dynamic insulin degradation behavior of the model was compared to experimentally determined time courses of insulin concentration in plasma after an intravenous insulin injection. Experimental data sets with extremely high [ 58 ] and extremely low [ 59 ] amounts of injected insulin were used. The extremely low amount (1.65·10 -6 nmol ) was radioactively labeled.…”

“…Therefore, the dynamics of injected insulin can be tracked though the amount of injected insulin is small compared to endogenous insulin. Measured concentrations of labeled insulin were about 0.5·10 -4 nM [ 59 ] (see Figure 2 ). The extremely high amount of injected insulin (47.5 nmol ) resulted in measured insulin concentrations above 1600 nM [ 58 ] (see Figure 3 ).…”