The marked propensity of insulin to self-associate into large aggregates causes significant mechanical problems in insulin delivery devices and may also stimulate production of a tissue-amyloid A precursor in some patients. Although conventionally prepared sulfated insulin (SI) resists aggregation, clinical application has been limited by major insulin bioactivity losses that occur during synthesis.

To eliminate this problem, insulin sulfation was carried out in the organic solvent dimethylformamide in the presence of condensing agents such as N,N'-dicyclohexyl carbodiimide (DCC) and a sulfate donor. With this new procedure, the degree of sulfation could be controlled over an eightfold range by varying the amount of condensing agent. The bioactivity of these new SI derivatives varied between 78% and 87% of unmodified insulin. Insulin aggregation, induced by passage through a syringe and needle, did not occur with derivatives having two or more sulfate moieties per insulin molecule. Diffusion velocity studies using “non-aggregated” insulin solutions demonstrated that aggregates were present in crystalline zinc and sodium porcine insulin. In contrast, SI having more than 0.5 mole sulfate per mole of insulin dialyzed as if it were predominantly in the monomeric form.

Results from the studies described in this report now provide the means for selectively designing and preparing specific high-potency, non-aggregating insulins, which may be necessary for optimal use of current and future insulin delivery devices.

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