Prevention of experimental autoimmune myasthenia gravis by rat Crry-Ig: A model agent for long-term complement inhibition in vivo

Abstract

Despite its vital role in innate immunity, complement is involved in a number of inflammatory pathologies and has therefore become a therapeutic target. Most agents generated for anti-complement therapy have short half-lives in plasma, or have been of mouse or human origin, thereby limiting their use either to murine models of disease or to short-term therapy. Here we describe the generation of a long-acting rat therapeutic agent based on the rat complement inhibitor, Crry. Characterisation of various soluble forms of Crry demonstrated that the amino-terminal four short-consensus repeat domains were required for full regulatory and C3b-binding activities. Fusion of these domains to rat IgG2a Fc generated an effective complement inhibitor (rCrry-Ig) with a circulating half-life prolonged from 7 min for Crry alone to 53 h for rCrry-Ig. Systemic administration of rCrry-Ig over 5 weeks generated a weak immune response to the recombinant agent, however this was predominantly IgM in nature and did not neutralise Crry function or cause clearance of the agent from plasma. Administration of rCrry-Ig completely abrogated clinical disease in a rat model of myasthenia gravis whereas soluble Crry lacking the immunoglobulin Fc domain caused a partial response. rCrry-Ig not only ablated clinical disease, but also prevented C3 and C9 deposition at the neuromuscular junction and inhibited cellular infiltration at this site. The long half-life and low immunogenicity of this agent will be useful for therapy in chronic models of inflammatory disease in the rat.

Zymosan, but Not Lipopolysaccharide, Triggers Severe and Progressive Peritoneal Injury Accompanied by Complement Activation in a Rat Peritonitis Model

“…Thickness of peritoneum (A), LCApositive cell infiltration (B), and ED-1 positive cell infiltration (C) in the peritoneum were all significantly increased in group 2 compared with group 1. peritoneal inflammation. Inhibition of complement activation was found for up to 53 h after a systemic single injection of Crry-Ig (20 mg/kg) as described in our previous report (30); in the current study, we i.p. injected 1.5 mg of Crry-Ig per animal in 0.5 ml of sterile PBS mixed with the PDF in the scraped peritonitis model just after the operation and every 2 days thereafter, Rats also received daily zymosan injections for 5 days after scraping (group 8 in Table I, n ϭ 6).…”

“…Crry-Ig has a longer half-life in vivo compared with sCR1, reducing the number of i.p. administrations required (30). Both therapies were effective, local Crry-Ig administration rather better than local sCR1 administration, likely because of better pharmacokinetics.…”

A Novel Antibody against Human Factor B that Blocks Formation of the C3bB Proconvertase and Inhibits Complement Activation in Disease Models

“…Experimental autoimmune MG (EAMG) was passively induced in young adult female Lewis rats essentially as described (20,24). In brief, rats were weighed and prebled, then injected i.p.…”

“…The results represent the mean 6 SD of three independent experiments. EAMG can be induced actively by AChR immunization or passively by transfer of anti-AChR Abs in mice and rats (20,24). Strong experimental evidence supports the critical role of complement in the development of both the human disease and the rodent models (7,41).…”

Effect of complement and its regulation on myasthenia gravis pathogenesis

“…In the past few years, there has been an appreciation for the role of complement inhibitors in the autoimmune diseases, and several groups have reported beneficial results of complement inhibitors in reducing severity of EAMG [64,68,89]. Certainly, the ability for these reagents to treat MG will rely on the overall effect that these may have on the systemic immune system.…”