Abstract
The insulin-like growth factor (IGF) signaling pathway is involved in certain human cancers, and the feasibility of directly targeting the IGF receptor has been actively investigated. However, recent evidence from clinical trials suggests that this approach can be problematic. We have developed an alternative strategy to indirectly inhibit the IGF signaling by targeting the metalloproteinase, pregnancy-associated plasma protein-A (PAPP-A). PAPP-A associated with the cell surface cleaves IGF binding protein-4 (IGFBP-4), when IGF is bound to IGFBP-4, and thereby increases IGF bioavailability for receptor activation in an autocrine/paracrine manner. We hypothesized that inhibition of PAPP-A would suppress excessive local IGF signaling in tissues where this is caused by increased PAPP-A proteolytic activity. To test this hypothesis, we developed an inhibitory monoclonal antibody, mAb 1/41, which targets a unique substrate-binding exosite of PAPP-A. This inhibitor selectively and specifically inhibits proteolytic cleavage of IGFBP-4 with an inhibitory constant (Ki) of 135 pM. In addition, it inhibited intracellular signaling of the IGF receptor (AKT phosphorylation) in monolayers of A549 cells, an IGF-responsive lung cancer-derived cell line found to express high levels of PAPP-A. We further showed that mAb 1/41 is effective towards PAPP-A bound to cell surfaces, and that it is capable of inhibiting PAPP-A activity in vivo. Using a murine xenograft model of A549 cells, we demonstrated that mAb 1/41 administered intraperitoneally significantly inhibited tumor growth. Analysis of xenograft tumor tissue recovered from treated mice showed penetration of mAb 1/41, reduced IGFBP-4 proteolysis, and reduced AKT phosphorylation. Our study provides proof of concept that IGF signaling can be selectively reduced by targeting a regulatory proteinase that functions extracellularly, upstream of the IGF receptor. PAPP-A targeting thus represents an alternative therapeutic strategy for inhibiting IGF receptor signaling.
Highlights
The insulin-like growth factor receptor type I (IGFIR) and its native ligands, IGF-I and IGF-II, play crucial regulatory roles in many cellular processes, during normal development and growth [1, 2]
Selected candidates were screened for their ability to inhibit plasma protein-A (PAPP-A) cleavage of IGF binding protein-4 (IGFBP-4), and one antibody, mAb 1/41, was chosen for further characterization following production in milligram quantities
Use of IGFBPs to sequester IGF bioactivity is being attempted [15], but there are no prior reports in which the proteolytic activity of an IGFBP proteinase has been targeted
Summary
The insulin-like growth factor receptor type I (IGFIR) and its native ligands, IGF-I and IGF-II, play crucial regulatory roles in many cellular processes, during normal development and growth [1, 2]. Several different strategies have been developed, including the use of 1) anti-ligand antibodies to reduce the concentration of bioactive ligand capable of stimulating the receptor, 2) anti-receptor antibodies, which trigger receptor internalization and block binding of active IGF-I or -II, and 3) small molecule inhibitors of the intracellular receptor tyrosine kinase domain [11, 12]. All of these strategies have made the transition into clinical trials, each of them is connected with potential problems, and the outcomes have been less fruitful than anticipated [13, 14]
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