Targeting autocrine HB-EGF signaling with specific ADAM12 inhibition using recombinant ADAM12 prodomain.

Miles A Miller,Douglas A Lauffenburger,Aaron S Meyer,Linda G Griffith,Lori Edwards,Marcia L Moss,Robert Petrovich,Gary Powell

doi:10.1038/srep15150

Abstract

Dysregulation of ErbB-family signaling underlies numerous pathologies and has been therapeutically targeted through inhibiting ErbB-receptors themselves or their cognate ligands. For the latter, “decoy” antibodies have been developed to sequester ligands including heparin-binding epidermal growth factor (HB-EGF); however, demonstrating sufficient efficacy has been difficult. Here, we hypothesized that this strategy depends on properties such as ligand-receptor binding affinity, which varies widely across the known ErbB-family ligands. Guided by computational modeling, we found that high-affinity ligands such as HB-EGF are more difficult to target with decoy antibodies compared to low-affinity ligands such as amphiregulin (AREG). To address this issue, we developed an alternative method for inhibiting HB-EGF activity by targeting its cleavage from the cell surface. In a model of the invasive disease endometriosis, we identified A Disintegrin and Metalloproteinase 12 (ADAM12) as a protease implicated in HB-EGF shedding. We designed a specific inhibitor of ADAM12 based on its recombinant prodomain (PA12), which selectively inhibits ADAM12 but not ADAM10 or ADAM17. In endometriotic cells, PA12 significantly reduced HB-EGF shedding and resultant cellular migration. Overall, specific inhibition of ligand shedding represents a possible alternative to decoy antibodies, especially for ligands such as HB-EGF that exhibit high binding affinity and localized signaling.

Highlights

The ErbB family of four closely related receptor tyrosine kinases (RTKs) – the epidermal growth factor receptor (ERBB1/EGFR), ERBB2/HER2, ERBB3/HER3, and ERBB4/HER4 – is implicated in various invasive diseases for promoting aberrant cell survival, proliferation, and migration
A computational model of autocrine signaling accurately predicts that heparin-binding epidermal growth factor (HB-EGF) signaling is more localized than the low-affinity ligand, amphiregulin (AREG)
To study how varied physicochemical properties of ErbB-ligands influence overall autocrine signaling behavior, we developed a model of EGFR signaling based on ordinary differential equations (ODEs) that described receptor production and internalization, ligand shedding and localized diffusion, and ligand-receptor binding (Fig. 1A)

Summary

Introduction

The ErbB family of four closely related receptor tyrosine kinases (RTKs) – the epidermal growth factor receptor (ERBB1/EGFR), ERBB2/HER2, ERBB3/HER3, and ERBB4/HER4 – is implicated in various invasive diseases for promoting aberrant cell survival, proliferation, and migration. These two effects are combined in the case of heparin-binding epidermal growth factor (HB-EGF), which activates both EGFR and ERBB4 at high affinity and leads to cetuximab resistance[6] This evidence has motivated the development of complimentary strategies for targeting ErbB-family signaling that extends beyond direct binding and inhibition of EGFR and HER2. We found that ADAM12 activity correlated closely with HB-EGF shedding in endometriosis; we developed a specific inhibitor of ADAM12 based on its recombinant prodomain (PA12) to reduce HB-EGF shedding, and demonstrated it as effective Taken together, these results i) provide a quantitative explanation of limiting factors in using decoy antibodies against growth-factor ligands, relevant to high affinity ligands such as HB-EGF; ii) demonstrate ADAM12 as a relevant sheddase of HB-EGF in endometriosis; and iii) present a novel, specific ADAM12 inhibitor to reduce HB-EGF shedding and resulting cell migration behavior

Methods

Results

Conclusion