Structural Basis for Eculizumab-Mediated Inhibition of the Complement Terminal Pathway

Abstract

Eculizumab is a humanized mAb approved for treatment of patients with paroxysmal nocturnal hemoglobinuria and atypical hemolytic uremic syndrome. Eculizumab binds complement component C5 and prevents its cleavage by C5 convertases, inhibiting release of both the proinflammatory metabolite C5a and formation of the membrane attack complex via C5b. In this study, we present the crystal structure of the complex between C5 and a Fab fragment with the same sequence as eculizumab at a resolution of 4.2 Å. Five CDRs contact the C5 macroglobulin 7 domain, which contains the entire epitope. A complete mutational scan of the 66 CDR residues identified 28 residues as important for the C5–eculizumab interaction, and the structure of the complex offered an explanation for the reduced C5 binding observed for these mutant Abs. Furthermore, the structural observations of the interaction are supported by the reduced ability of a subset of these mutated Abs to inhibit membrane attack complex formation as tested in a hemolysis assay. Our results suggest that eculizumab functions by sterically preventing C5 from binding to convertases and explain the exquisite selectivity of eculizumab for human C5 and how polymorphisms in C5 cause eculizumab-resistance in a small number of patients with paroxysmal nocturnal hemoglobinuria.

Long lasting neutralization of C5 by SKY59, a novel recycling antibody, is a potential therapy for complement-mediated diseases

“…CFA0322 cross-reacts with cynomolgus monkey C5 and has a highly similar profile to eculizumab in terms of neutralizing activity and binding epitope. Similar to eculizumab 21 , CFA0322 does not bind to C5 R885H, suggesting that binding epitope of eculizumab partially overlaps with that of CFA0322 (data not shown). CFA0322 therefore serves as a surrogate for eculizumab to enable comparison against SKY59.…”

“…recently demonstrated that these patients carry a common single nucleotide polymorphism (SNP) in C5 where arginine at position 885 is substituted with histidine 16 . Since arginine at 885 in the α-chain of C5 is within the binding epitope of eculizumab 11, 21 , this C5 variant cannot be neutralized by eculizumab. In contrast, SKY59 could be expected to neutralize the activity of this C5 variant because the epitope of SKY59 is distinct from that of eculizumab.…”

Complement and Bacterial Infections: From Molecular Mechanisms to Therapeutic Applications

“…Eculizumab forms a 1: 1 complex with C5, and sterically hinders convertases from binding and cleaving C5 into C5a and C5b [74]. Eculizumab successfully prevents intravascular hemolysis and thrombosis and has revolutionized disease management of PNH [75].…”

Incomplete inhibition by eculizumab: mechanistic evidence for residual C5 activity during strong complement activation

“…[6][7][8][9][10] Eculizumab binds C5 with picomolar affinity and inhibits its enzymatic activation by C5 convertases, possibly through steric hindrance. 11,12 However, a recent study indicates that eculizumab not only acts sterically, by blocking binding to the C5 convertase, but also prevents C5 to adopt a primed conformation that is susceptible to processing by the C5 convertase. 13 A similar mechanism has been suggested for the tick inhibitor OmCI (Ornithodoros moubata complement inhibitor) or its recombinant version, coversin, which binds C5 at the face opposite to the eculizumab epitope.…”