Engineering Novel, Prolonged-Acting Insulins

  • Jan Markussen
Part of the New Horizons in Therapeutics book series (NHTH)


Insulin-dependent diabetic patients need a constant supply of basal insulin to control blood glucose between meals, in addition to meal-related bolus insulin to counter the peak in glucose following a meal. Intensive basal/bolus insulin regimens have been introduced in order to improve blood glucose control, particularly in Scandinavia and England. The long-acting insulins used to provide the basal supply come in the form of neutral, crystalline suspensions, obtained by crystallization using eitherZn2+ (Lente-type preparations) or protamine (NPHtype preparations). These suspensions require thorough shaking prior to injection in order to ensure homogeneity of the suspension and correct dosage. Furthermore, the inter-and intra-patient absorption of insulin suspensions is extremely variable: the T50% (i.e., the time taken for 50% of the insulin to disappear from the injection site) ranges from 8 to 72 hr (Lauritzen et al., 1979). Whereas the homogeneity problem of suspensions could be overcome if a soluble, prolonged-acting insulin was made available, the variability in absorption caused by the physiological conditions, like blood flow at the site of injection, could hardly be totally eliminated by applying new principles of prolongation.


Human Insulin Basal Insulin Basic Amino Acid Insulin Antibody Porcine Insulin 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Binder, C., 1969, Absorption of Injected Insulin, Munksgaard, Copenhagen.Google Scholar
  2. Blundell, T. L.. Cutfield, J. F., Cutfield, S. M., Dodson, E. J., Dodson, G. G., Hodgkin. D. C.. and Mercola, D. A.. 1972, Three-dimensional atomic structure of insulin and its relationship to activity, Diabetes 21(suppl. 2): 492–505.PubMedGoogle Scholar
  3. Bradbury, A. F., Finnic, M. D. A., and Smyth. D. G., 1982, Mechanism of C-terminal amide formation by pituitary enzymes. Nature 298: 686–688.PubMedCrossRefGoogle Scholar
  4. Brange, J., Skelbaek-Pedersen. B., Langkjaer, L. Damgaard. U., Ege, H., Havelund. S.. Heding, L. G., Jorgensen. K. H., Lykkeberg, J.. Markussen, J., Pingel, M. and Rasmussen. E., 1987, Galenics of Insulin, pp. 36–38. 54–57, Springer-Verlag. Berlin.Google Scholar
  5. Brange, J., Ribel. U., Hansen. J. F., Dodson, G., Hansen, M. T., Havelund, S., Melberg, S. G., Norris. F., Norris, K., Snel, L., Sorensen, A. R., and Voigt, H. O., 1988, Monomeric insulins obtained by protein engineering and their medical implications, Nature 333: 679–682.PubMedCrossRefGoogle Scholar
  6. British Pharmacopoeia, 1980, Her Majesty’s Stationery Office. London, A 142.Google Scholar
  7. European Pharmacopoeia, 1984, V.2.2.3. Assay of insulin method C, Her Majesty’s Stationery Office, London.Google Scholar
  8. Geiger, T., and Clarke, S.. 1987. Deamidation, isomerization, and racemization at asparaginyl and aspartyl residues in peptides, J. Biol. Chem. 262: 785–794.PubMedGoogle Scholar
  9. Gliemann. J. and Gammeltoft. S., 1974, The biological activity and the binding affinity of modified insulins determined on isolated rat fat cells, Diabetologia 10: 105–113.PubMedCrossRefGoogle Scholar
  10. Helbig, H-J., 1976, Insulindimere aus der B-Komponente von Insulinpräparationen, Rheinisch-Westfälischen Technischen Hochschule, Aachen.Google Scholar
  11. Hodgkin, D. C., Dodson, E., Dodson, G., and Reynolds. C., 1983, Insulin, Biochem. Soc. Trans. 11: 411–417.PubMedGoogle Scholar
  12. Hübinger, A., Becker, A., and Gries, F. A., 1988, Total insulin levels in Type 1 diabetic patients with insulin antibodies and their effect on insulin requirement and metabolic control, Diabetes Res. 7: 65–69.PubMedGoogle Scholar
  13. Kobayashi, M., Ishibashi, O., Takata, Y., Haneda, M., Maegawa, H., Watanabe, N., and Shigeta, Y., 1985, Prolonged disappearance rate of a structurally abnormal mutant insulin from the circulation in humans, J. Clin. Endocrinol. Metab. 61: 1142–1145.PubMedCrossRefGoogle Scholar
  14. Lauritzen, T., 1985, Pharmacokinetic and Clinical Aspects of Intensified Subcutaneous Insulin Therapy, Laegeforeningens Forlag, Copenhagen.Google Scholar
  15. Lauritzen, T., Faber, O. K., and Binder, C., 1979, Variations in 125I-insulin absorption and blood glucose concentration, Diabetologia 17: 291–295.PubMedCrossRefGoogle Scholar
  16. Leach, S. J., and Lindley, H., 1953, The kinetics of hydrolysis of the amide group in proteins and peptides, Trans. Faraday Soc. 49: 915–925.CrossRefGoogle Scholar
  17. Markussen, J., 1982, U.S. patent 4, 343, 898.Google Scholar
  18. Markussen, J., 1985, Comparative reduction/oxidation studies with single chain des-(B30) insulin and porcine proinsulin, Int. J. Peptide Protein Res. 25: 431–434.CrossRefGoogle Scholar
  19. Markussen, J., 1987, Human Insulin by Tryptic Transpeptidations of Porcine Insulin and Biosynthetic Precursors, MTP Press, Lancaster.Google Scholar
  20. Markussen, J., Jorgensen, K. H., Sorensen, A. R., and Thim, L., 1985, Single chain des-(B30) insulin, Int. J. Peptide Protein Res. 26: 70–77.CrossRefGoogle Scholar
  21. Markussen, J., Hougaard, P., Ribel, U., Sorensen, A. R., and Sorensen, E., 1987a, Soluble, Prolonged-acting insulin derivatives. I. Degree of protraction and crystallizability of insulins substituted in the termini of the B-chain, Protein Engineering 1(3): 205–213.Google Scholar
  22. Markussen, J., Diers, I., Engesgaard, A., Hansen, M. T., Hougaard, P., Langkjaer, L., Norris, K., Ribel, U., Sorensen, A. R., Sorensen, E., and Voigt, H. O., 1987b, Soluble prolonged-actingGoogle Scholar
  23. insulin derivatives. II. Degree of protraction and crystallizability of insulins substituted in positions A17, B8, B13, B27 and B30, Protein Engineering 1(3):215–223.Google Scholar
  24. Markussen, J., Damgaard, U., Diers, I., Fiil, N., Hansen, M. T., Larsen, P., Norris, F., Norris, K., Schou, O., Snel, L., Thim, L., and Voigt, H. O., 1987c, Biosynthesis of human insulin in yeast via single-chain precursors, in: Peptides 1986( D. Theodoropoulos, ed.), pp. 189–194, Walter de Gruyter, Berlin.Google Scholar
  25. Markussen, J., Diers, I., Hougaard, P., Langkjaer, L., Norris, K., Snel, L., Sorensen, A. R., Sorensen, E., and Voigt, H. O., 1988a, Soluble, prolonged-acting insulin derivatives. III. Degree of protraction, crystallizability and chemical stability of insulins substituted in positions A21, B13, B23, B27 and B30, Protein Engineering, 2(2): 157–166.Google Scholar
  26. Markussen, J., Hansen, M. T., Norris, K., and Sorensen, E., 1988b, Synthesis of insulins substituted in positions A17, B13, B27 and in the terminals of the B-chain, combining genetic engineering and Cryptic transpeptidation in organic-aqueous medium, in: Peptide Chemistry 1987( T. Shiba, and S. Sakakibara, eds.), Protein Research Foundation, Osaka pp. 417–422.Google Scholar
  27. Marshall, M. O., Heding, L. G., Villumsen, J., Akerblom, H. K., Baevre, H., Dahlquist, G., Kjaergaard, J-J., Knip, M, Lindgren, F., Ludvigsson, J., Persson, B., Rilva, A., Stenhammer, L., Stromberg, L., Sovik, O., Thalme, B., Vidnes, J., and Wefring, K., 1988, Development of insulin antibodies, metabolic control and B-cell function in newly diagnosed insulin dependent diabetic children treated with monocomponent human insulin or monocomponent porcine insulin, Diabetes Res. 9: 169–175.PubMedGoogle Scholar
  28. Moody, A. J., Stan, M. A., and Stan, M., 1974, A simple free fat cell bioassay for insulin, Horm. Metab. Res. 6: 12–16.PubMedCrossRefGoogle Scholar
  29. Schlichtkrull, J., 1958, Insulin Crystals, Munksgaard, Copenhagen.Google Scholar
  30. Schlichtkrull, J., Pingel, M., Heding, L. G., Brange, J., and Jorgensen, K. H., 1975, Insulin preparations with prolonged effect, in: Handbook of Experimental Pharmacology, Vol. XXXII/2 ( A. Hasselblatt and F. v. Bruchhausen, eds.), Springer-Verlag, Berlin pp. 729–777.Google Scholar
  31. Sundby, F., 1962, Separation and characterization of acid-induced insulin transformation products by paper electrophoresis in 7 M urea, J. Biol. Chem. 237: 3406–3411.PubMedGoogle Scholar
  32. Thim, L., Hansen, M. T., Norris, K., Hoegh, I., Boel, E., Forström, J., Ammerer, G., and Fiil, N. P., 1986, Secretion and processing of insulin precursors in yeast, Proc. Natl. Acad. Sci. USA 83: 6766–6770.PubMedCrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1990

Authors and Affiliations

  • Jan Markussen
    • 1
  1. 1.Novo Research InstituteBagsvaerdDenmark

Personalised recommendations