Targeting Pathogenic Postischemic Self-Recognition by Natural IgM to Protect Against Posttransplantation Cardiac Reperfusion Injury

Abstract

Background Natural IgM antibodies represent a class of innate pattern recognition receptors that recognize danger associated molecular patterns expressed on stressed or dying cells. They play important roles in tissue homeostasis by disposing of pre-necrotic cells and suppressing inflammation. However, ischemic insult leads to a pathogenic level of IgM binding and complement activation, resulting in inflammation and injury. We investigate the role of self-reactive IgM in the unique setting of transplantation, where the donor organ undergoes both cold and warm ischemia, and global ischemic insult. Methods and Results By transplanting hearts from wild-type donor mice into antibody-deficient mice reconstituted with specific self-reactive IgM mAbs, we identified neoepitopes expressed post-transplant, and demonstrated a key role for IgM recognition of these epitopes in graft injury. With this information, we developed and characterized a therapeutic strategy that exploited the post-ischemia recognition system of natural antibodies. Based on neoepitope identification, we constructed an anti-annexin-IV single chain antibody (scFv) and an scFv linked to Crry, an inhibitor of C3 activation (scFv-Crry). In an allograft transplant model, in which recipients contain a full natural antibody repertoire, both constructs blocked graft IgM binding and complement activation, and significantly reduced graft inflammation and injury. Furthermore, scFv-Crry specifically targeted to the transplanted heart and, unlike complement deficiency, did not affect immunity to infection, an important consideration for immunosuppressed transplant recipients. Conclusions We identified pathophysiologically important epitopes expressed within the heart post-transplant, and describe a novel translatable strategy for targeted complement inhibition that has several advantages over currently available approaches.

Targeted Complement Inhibition Protects Vascularized Composite Allografts From Acute Graft Injury and Prolongs Graft Survival When Combined With Subtherapeutic Cyclosporine A Therapy

“…Complement-dependent injury will also result in the expression of DAMPs, such as heat shock proteins, reactive oxygen species and fibrinogen 5,37 , that can play a role in the activation of APCs and the augmentation of effector T cell responses 5,37 . In this context, our data also indicate IgM-mediated activation of complement occurs in composite allografts, and for cardiac grafts we have shown previously that DAMPs exposed after transplant are recognized by self-reactive natural IgM Abs that activate complement in the transplanted heart 38 . Taken together, the activities of complement activation products provide a basis for how complement may augment graft allogenicity via IRI 5 …”

New therapeutic and diagnostic opportunities for injured tissue-specific targeting of complement inhibitors and imaging modalities

“…One neoepitope identified as being expressed post-transplant was ANX4, and this molecule was recognized by mAb B4 [58]. Subsequently, an anti-ANX4 scFv and the same scFv linked to Crry (B4-Crry) was constructed.…”

Innate immune mechanisms in transplant allograft vasculopathy

“…Besides the MyD88 signaling pathway, complement inhibition has also emerged as a putative strategy for blocking AV lesion formation. Two recent studies targeted endogenous complement regulators CD59 [58] and Crry [59], a murine complement regulatory ortholog inhibiting C3, using a CR2 fusion protein and single chain antibody, respectively, to relevant sites in vivo in murine hosts to block complement activation in response to IRI. Another recent study used mAb to functionally block proximal activation with anti-C1s [60].…”