AGR2, a unique tumor-associated antigen, is a promising candidate for antibody targeting

Abstract

Anterior gradient 2 (AGR2), a protein disulfide isomerase, shows two subcellular localizations: intracellular (iAGR2) and extracellular (eAGR2). In healthy cells that express AGR2, the predominant form is iAGR2, which resides in the endoplasmic reticulum. In contrast, cancer cells secrete and express eAGR2 on the cell surface. We wanted to test if AGR2 is a cancer-specific tumor-associated antigen. We utilized two AGR2 antibodies, P3A5 and P1G4, for in vivo tumor localization and tumor growth inhibition. The monoclonal antibodies recognized both human AGR2 and mouse Agr2. Biodistribution experiments using a syngeneic mouse model showed high uptake of P3A5 AGR2 antibody in xenografted eAgr2 + pancreatic tumors, with limited uptake in normal tissues. In implanted human patient-derived eAGR2 + pancreatic cancer xenografts, tumor growth inhibition was evaluated with antibodies and Gemcitabine (Gem). Inhibition was more potent by P1G4 + Gem combination than Gem alone or P3A5 + Gem. We converted these two antibodies to human:mouse chimeric forms: the constructed P3A5 and P1G4 chimeric mV L hC κ and mV H hC γ (γ1, γ2, γ4) genes were inserted in a single mammalian expression plasmid vector, and transfected into human 293F cells. Expressed human:mouse chimeric IgG1, IgG2 and IgG4 antibodies retained AGR2 binding. Increase in IgG yield by transfected cells could be obtained with serial transfection of vectors with different drug resistance. These chimeric antibodies, when incubated with human blood, effectively lysed eAGR2 + PC3 prostate cancer cells. We have, thus, produced humanized anti-AGR2 antibodies that, after further testing, might be suitable for treatment against a variety of eAGR2 + solid tumors.

Inhibitors of ERp44, PDIA1, and AGR2 induce disulfide-mediated oligomerization of Death Receptors 4 and 5 and cancer cell death

“…functions are under development [51,52], but agents that block the essential role of AGR2 in the folding of client proteins are not available.…”

“…However, AGR2 may also be secreted and promote tumor angiogenesis [49, 50] and act on cell surface receptors [51]. Monoclonal antibodies designed to block extracellular AGR2 functions are under development [51, 52], but agents that block the essential role of AGR2 in the folding of client proteins are not available.…”

Antibody Therapy Targeting Cancer-Specific Cell Surface Antigen AGR2

“…Each cDNA contained a Kozak box. Plasmids of chimeric P1G4 hIgG1, hIgG2, hIgG4; P3A5 hIgG1, hIgG2, hIgG4 were generated [22]. The different Cγ plasmids could be distinguished by Sac II and Eco RI digestion.…”

“…Mouse antibodies, besides being immunogenic in human, do not interact efficiently with human immune system components. To overcome these drawbacks, we have replaced the constant domains of AGR2 antibodies by the analogous human constant domains via recombinant DNA technology [21], generating human:mouse chimeric hIgG1, hIgG2, hIgG4 for both P1G4 and P3A5 for potential therapeutic development [22].…”

Secretion of pro-oncogenic AGR2 protein in cancer

“…This is the first study that showed such promising result of antibody-based therapy targeting AGR2, suggesting that this molecule is an important target in cancer development. Another rather exciting finding is the production of two humanized monoclonal anti-AGR2 antibodies, when incubated with human blood, resulting in cell lysis of prostate cancer cells containing eAGR2 [ 113 ].…”