Therapeutic ACPA inhibits NET formation: a potential therapy for neutrophil-mediated inflammatory diseases

Abstract

Excessive release of neutrophil extracellular traps (NETs) is associated with disease severity and contributes to tissue injury, followed by severe organ damage. Pharmacological or genetic inhibition of NET release reduces pathology in multiple inflammatory disease models, indicating that NETs are potential therapeutic targets. Here, we demonstrate using a preclinical basket approach that our therapeutic anti-citrullinated protein antibody (tACPA) has broad therapeutic potential. Treatment with tACPA prevents disease symptoms in various mouse models with plausible NET-mediated pathology, including inflammatory arthritis (IA), pulmonary fibrosis, inflammatory bowel disease and sepsis. We show that citrulline residues in the N-termini of histones 2A and 4 are specific targets for therapeutic intervention, whereas antibodies against other N-terminal post-translational histone modifications have no therapeutic effects. Because citrullinated histones are generated during NET release, we investigated the ability of tACPA to inhibit NET formation. tACPA suppressed NET release from human neutrophils triggered with physiologically relevant human diseaserelated stimuli. Moreover, tACPA diminished NET release and potentially initiated NET uptake by macrophages in vivo, which was associated with reduced tissue damage in the joints of a chronic arthritis mouse model of IA. To our knowledge, we are the first to describe an antibody with NET-inhibiting properties and thereby propose tACPA as a drug candidate for NET-mediated inflammatory diseases, as it eliminates the noxious triggers that lead to continued inflammation and tissue damage in a multidimensional manner.

Anti-Citrullinated Protein Antibodies in Patients with Rheumatoid Arthritis: Biological Effects and Mechanisms of Immunopathogenesis

“…Appealingly, a therapeutic ACPA design to alter the binding of ACPAs, by targeting citrulline at a specific position, was documented to suppress NETosis from human neutrophils triggered by disease-relevant stimuli [121]. The changes in the neutrophil response upon ACPA alteration again strengthen the importance of ACPAs in triggering NETosis [121].…”

“…Just as Khandpur and others discovered that NETosis was enhanced in neutrophils in RA synovial fluid and systemic circulation, sera extracted from RA patients, particularly those rich in ACPAs, were shown to significantly stimulate NETosis [101]. Appealingly, a therapeutic ACPA design to alter the binding of ACPAs, by targeting citrulline at a specific position, was documented to suppress NETosis from human neutrophils triggered by disease-relevant stimuli [121]. The changes in the neutrophil response upon ACPA alteration again strengthen the importance of ACPAs in triggering NETosis [121].…”

“…Through grafting the citrullinated epitopes into a stable scaffold, the synthetic high-affinity peptide-based scavenger of ACPAs demonstrated superior stability with apparent low nanomolar affinity, which makes in vivo testing possible in the future [172]. Recently, therapeutic ACPAs specifically created to target citrulline residues in the N-terminus of histones 2A and 4 have been tested in various NET-relevant diseases, including inflammatory arthritis [121]. Although not specifically designed to treat diseases resulting from ACPA overproduction, the attenuation of NETosis and sequential inflammatory processes via citrulline blockade is a useful application referenced by the pathogenic mechanisms of ACPAs.…”

The Roles of Immune Cells in the Pathogenesis of Fibrosis

“…A clinical study has reported higher levels of NE in lung parenchyma, bronchoalveolar lavage fluid (BALF) and sera from patients with IPF when compared with those in normal subjects, suggesting an involvement of NE in the pathogenesis of IPF [ 33 ]. Furthermore, pharmacological inhibition of neutrophil extracellular trap formation may represent a promising therapy for the treatment of various inflammatory diseases [ 34 ].…”

Association Between Neutrophils and Renal Impairment of Rheumatoid Arthritis: A Retrospective Cross-sectional Study

“…NETs, a network structure consisted of DNA and granulose, could indirectly impair the endothelial function,promote blood vessel and glomerular damage, thus it would induce renal failure and even death [21]. Several previous studies respectively reported that the imbalance between the emergence and removal of NETs adversely affected kidney health [22,23].…”