Impact of receptor downregulation on clearance of two human EGFs with different receptor binding activity.

Human epidermal growth factor [hEGF(1-53)] has been thought to be cleared mainly via an EGF receptor (EGFR) endocytosis pathway. Pretreatment of rats with hEGF(1-53) has been shown previously to cause a dramatic reduction in clearance of the peptide contributable to EGFR downregulation. The impact of receptor downregulation has raised concerns for rational design of dosage regimen for this potential wound-healing therapeutic peptide. However, following a similar protocol, we could not reproduce the dramatic reduction in clearance reported previously mediated by an i.v. bolus acute dose. As EGFR downregulation may be sensitive to the length of exposure and to the activation of the receptor tyrosine kinase activity, two other pretreatment protocols were also evaluated: a 4-h i.v. infusion (prolonged exposure) of the peptide and an i.v. bolus of a potent synthetic kinase inhibitor pretreatment were evaluated for effects on clearance. However, neither pretreatment affected the peptide's clearance profile. Further, no effects on clearance and other kinetic parameters were observed for any pretreatment paradigms with a truncated analogue hEGF (1-48), whose EGF receptor binding activity is much weaker but plasma clearance is much higher than hEGF (1-53). In addition, a study in a second rat strain showed no difference in clearance profile of hEGF-(1-53) following pretreatment. Results of the present investigation suggest that receptor binding does not have a direct relationship with plasma clearance, and that the EGF clearance mechanisms is highly refractory with EGF receptors possibly recovering rapidly from downregulation through the recycling process.