LONG-ACTING GROWTH-HORMONES PRODUCED BY CONJUGATION WITH POLYETHYLENE-GLYCOL

Derivatives of human growth hormone (hGH) of in creasing size were pro duced by reaction with the N-hydroxysuccinimide ester of polyethylene glycol-5000 (PEG(5000)), a 5-kDa reagent that selectively conjugates t o primary amines, By adjusting the reaction conditions and purificatio n procedure, it was possible to isolate hGH derivatives containing up to seven PEG moieties that altered the Stokes radius and thereby the e ffective molecular masses of the unmodified hormone from 22 to 300 kDa . Fortunately, the most reactive amines were ones that did not lie in either of the two sites important for receptor binding, Nonetheless, i ncreasing the level of PEG modification linearly reduced the affinity of hGH for its receptor and increased the EC(50) in a cell-based assay up to 1500-fold. Most of the reduction in affinity was the result of slowing the association rate for the receptor, The clearance rate of h GH in rats was inversely proportional to effective molecular weight an d closely fit a filtration model, We have tested the potency of these analogs by injecting them daily or every 6 days into hypophysectomized rats and determining the effects on body and organ growth, The effica cy of these analogs was optimal for hGH conjugated with 5 eq of PEG(50 00), and the potency was increased by about 10-fold compared with unmo dified hGH. Such PEG-hGH derivatives show promise as long-acting alter natives to daily injections of hGH. More generally these studies show that improving hormone clearance properties, even at the expense of re ducing receptor binding affinity, can lead to dramatic increases in ho rmone efficacy.