Immunological origin and functional properties of catalytic autoantibodies to amyloid beta peptide

Abstract

Objectives: Objectives The objectives of this study are to (1) evaluate the ability of the immune system to synthesize specific antibodies that catalyze the degradation of amyloid beta peptide (Abeta) and to (2) evaluate the prospect of developing a catalytic IVIG (CIVIG) formulation for therapy of Alzheimer's disease (AD).

Conclusions: Polyclonal autoantibodies from humans without dementia hydrolyzed Abeta specifically. The catalytic activity improved as a function of age. Patients with AD produced catalytic antibodies at increased levels. IgM-class antibodies expressed the activity at levels superior to IgGs. Production of catalytic autoantibodies appears to be an innate immunity function with adaptive improvements occurring upon Abeta overexpression, which suggests a beneficial function of the catalytic activity. The catalytic autoantibodies impeded Abeta aggregation, dissolved preformed Abeta aggregates, and inhibited Abeta cytotoxicity in tissue culture. Recombinant catalytic antibodies from a human library have been identified, validating the phenomenon of antibody-catalyzed Abeta cleavage. As a single catalyst molecule inactivates multiple Abeta molecules, catalytic antibodies may clear Abeta efficiently. IVIG did not cleave Abeta, indicating the importance of purification procedures that maintain catalytic site integrity. Traditional Abeta-binding antibodies form immune complexes that can induce inflammatory reaction and vascular dysfunction. Catalysts do not form stable immune complexes, minimizing these risks. Criteria appropriate for developing a CIVIG formulation with potential therapeutic utility are discussed, including isolation of the Abeta-specific catalytic subsets present in IgM and IgG from human blood.

Fig. 1

Potential for treatment of Alzheimer’s disease (AD): passive immunotherapy with Aβ binding and Aβ hydrolyzing Igs. a Aβ binding IgG. Reversibly binding IgG injected into peripheral blood can enter the brain in small amounts and help clear Aβ by mechanisms described in the text. Following binding of IgG to Aβ monomers (a) or aggregates (b), the immune complexes are ingested by an Fcγ receptor-dependent uptake process by microglia (c). The IgG bound by Fcn receptors at the blood-brain barrier (BBB) may help clear Aβ from the brain by facilitating efflux to the periphery through a transcytosis process (d). Microglial ingestion of the immune complexes is accompanied by release of inflammatory mediators, which could exacerbate the already inflamed state of the AD brain. In mouse AD models, clearance of amyloid plaques from the brain parenchyma induced by Aβ-binding IgGs is accompanied by Aβ deposition in the blood vessels and microhemorrhages. b Aβ hydrolyzing Ig. Sufficient amounts of a catalytic antibody administered into peripheral blood are predicted to gain entry into the brain and hydrolyze brain Aβ deposits and soluble oligomers, reducing the brain Aβ burden. Like the IgG in panel a, catalytic Ig containing the Fc domain may mediate an FcRn-mediated Aβ transcytosis. LRP1/RAGE transporters are known to help maintain the equilibrium between brain and peripheral Aβ. c Aβ hydrolyzing IgM. Hydrolysis of Aβ by peripheral IgM increases the Aβ brain-periphery concentration difference and thereby enhances Aβ efflux from the brain. Catalytic IgM bound to FcRμ/α on the luminal (blood) side of the BBB are hypothesized to enhance the local concentration difference at the BBB, thus inducing an overcompensatory depletion effect and facilitating sustained removal of brain Aβ into the periphery. FcRn neonatal Fc receptor; FcRμ/α Fc receptor for IgM; LRP-1 low-density lipoprotein receptor related protein 1, RAGE receptor for advanced glycation end products

Fig. 2

Cleavage of Aβ40 by human IgM and IgG. a Aβ40 (100 μM) was incubated for 3 days at 37°C with IgG (1.6 μM) or IgM (34 nM) pooled from six non-AD subjects each of age <35 years (young) or >72 years (old). Reactions were analyzed by reversed phase HPLC (gradient of 10% to 80% acetonitrile in trifluoroacetic acid, 45 min; detection at 220 nm). Hydrolysis rates were computed from the area of the Aβ1-28 product peak interpolated from a standard curve that was constructed using increasing amounts of synthetic Aβ1-28 (means ± SD; n=4 assays). *P<0.0044; **P<0.035. Two-tailed unpaired t test. b Molecular basis for epitope specificity of proteolytic antibodies: split-site model. Two different antibody subsites are responsible for the initial noncovalent antigen binding and the subsequent peptide bond hydrolysis process. In the initial immune complex (left), the antigen region not involved in noncovalent antibody binding enjoys conformational flexibility. Consequently, peptide bonds remote from the noncovalent binding site that are in register with the antibody nucleophilic subsite can be hydrolyzed (right). Triangle nucleophile, circle neighboring general base that activates the nucleophile

Fig. 3

Failure of IVIG preparations to cleave Aβ40. a IVIG Carimune, 2.2 mg/mL; control monoclonal IgM Yvo, 0.18 mg/mL. Catalytic activity measured as in Fig. 2a. b IVIG Intratect and IgM Yvo, 0.18 mg/mL. Catalytic activity measured as in [11] using ¹²⁵I-Aβ40 as substrate (mean ± SD; two replicates each)