In vitro and in vivo potency of insulin analogues designed for clinical use.

Analogues of human insulin designed to have improved absorption properties after subcutaneous injection have been prepared by recombinant DNA technology. Five rapidly absorbed analogues, being predominantly in mono- or di-meric states in the pharmaceutical preparation, and a hexameric analogue with very low solubility at neutral pH and slow absorption, were studied. Receptor binding assays with HEP-G2 cells showed overall agreement with mouse free adipocyte assays. Two analogues, B28Asp and A21Gly + B27Arg + B30Thr-NH2, had nearly the same molar in vitro potency as human insulin. Another two showed increased adipocyte potency and receptor binding, B10Asp 194% and 333% and A8His + B4His + B10Glu + B27His 575% and 511%, while B9Asp + B27Glu showed 29% and 18% and the B25Asp analogue only 0.12% and 0.05% potency. Bioassays in mice or rabbits of the analogues except B25Asp showed that they had the same in vivo potency as human insulin 1.00 IU = 6.00 nmol. Thus the variation had the same in vivo potency as human insulin 1.00 IU = 6.00 nmol. Thus the variation in in vivo potency reflects the differences in receptor binding affinity. Relative to human insulin a low concentration is sufficient for a high affinity analogue to produce a given receptor complex formation and metabolic response. In conclusion, human insulin and analogues with markedly different in vitro potencies were equipotent in terms of hypoglycaemic effect. This is in agreement with the concept that elimination of insulin from blood and its subsequent degradation is mediated by insulin receptors.

The absorption of insulin.

It is desirable to improve metabolic control in diabetics receiving insulin. Studies on insulin kinetics provide information that may be useful in the optimization of the injection scheme. Absorption has been measured using 125I-insulin as marker and counting the residual activity with a crystal detector. Plasma insulin has been measured (by radioimmunoassay) in the absence of insulin antibodies. The relationship between the rate of absorption and exogenous insulinemia was analyzed using a simple mathematical model. The time course of plasma insulin was calculated for various insulin preparations, including mixtures. The data suggest the significance of the injection site for the effect of insulin, e.g., a more rapid effect after injection into the abdominal subcutaneous tissue. The pattern of exogenous insulinemia in patients treated with insulin bears no resemblance to that in normal subjects. The calculated patterns of exogenous plasma insulin after injection of various preparations suggest that insulin therapy might be improved to obtain a better and more physiological metabolic control.