Therapeutic hormones - Pharmaceutical Biotechnology: Concepts and Aapplications - page 303

11.2.7 Engineered insulins

Recombinant DNA technology facilitates not only production of human insulin in microbial
systems, but also facilitates generation of insulins of modifi ed amino acid sequences. The major
aims of generating such engineered insulin analogues include:

• Identifi cation of insulins with altered pharmacokinetic properties, such as faster-acting or

slower-acting insulins.

• Identifi cation of super-potent insulin forms (insulins with higher receptor affi nities). This is due

to commercial considerations, namely the economic benefi ts that would accrue from utilizing
smaller quantities of insulin per therapeutic dose.

The insulin amino acid residues that interact with the insulin receptor have been identifi ed (A1,
A5, A19, A21, B10, B16, B23-25), and a number of analogues containing amino acid substitutions
at several of these points have been manufactured. Conversion of histidine to glutamate at the B10
position, for example, yields an analogue displaying fi vefold higher activity in vitro . Other substi-
tutions have generated analogues with even higher specifi c activities. However, increased in vitro
activity does not always translate to increased in vivo activity.
Attempts to generate faster-acting insulins have centred upon developing analogues that do not
dimerize or form higher polymers at therapeutic dose concentrations. The contact points between
individual insulin molecules in insulin dimers/oligomers include amino acids at positions B8, B9,
B12-13, B16 and B23-28. Thus, analogues with various substitutions at these positions have been
generated. The approach adopted generally entails insertion of charged or bulky amino acids, in
order to promote charge repulsion or steric hindrance between individual insulin monomers. Sev-
eral are absorbed from the site of injection into the bloodstream far more quickly than native solu-
ble (fast-acting) insulin. Such modifi ed insulins could thus be injected at mealtimes rather than
1 h before, and several such fast-acting engineered insulins have now been approved for medical
use (Table 11.3). 'Insulin lispro' (tradename 'Humalog') was the fi rst such engineered short-acting
insulin to come to market (Box 11.1 and Figure 11.5).
'Insulin Aspart' is a second fast-acting engineered human insulin analogue now approved for
general medical use. It differs from native human insulin in that the proline B28 residue has been
replaced by aspartic acid. This single amino acid substitution also decreases the propensity of
individual molecules to self-associate, ensuring that they begin to enter the bloodstream from the
site of injection immediately upon administration.
A number of studies have also focused upon the generation of longer-acting insulin analogues.
The currently used Zn-insulin suspensions, or protamine-Zn-insulin suspensions, generally
display a plasma half-life of 20-25 h. Selected amino acid substitutions have generated insulins
which, even in soluble form, exhibit plasma half-lives of up to 35 h.
Optisulin or Lantus are the tradenames given to one such analogue that gained general market-
ing approval in 2000 (Table 11.3). The international non-proprietary name for this engineered
molecule is 'insulin glargine'. It differs from native human insulin in that the C-terminal aspargine
residue of the A-chain has been replaced by a glycine residue and the -chain has been elongated

(again from its C-terminus) by two arginine residues. The overall effect is to increase the mol-
ecule's pI (the pH at which the molecule displays a net overall zero charge and, consequently, at

β