Sequential Prodrug Strategy To Target and Eliminate ACPA-Selective Autoreactive B Cells

Abstract

Autoreactive B cells are thought to play a pivotal role in many autoimmune diseases. Rheumatoid arthritis (RA) is an autoimmune disease affecting ∼1% of the Western population and is hallmarked by the presence of anticitrullinated proteins antibodies (ACPA) produced by autoreactive B cells. We intend to develop a method to target and selectively eliminate these autoreactive B cells using a sequential antigen prodrug targeting strategy. As ACPA-expressing B cells are thought to play essential roles in RA-disease pathogenesis, we used this B cell response as a prototype to analyze the feasibility to generate a construct consisting of a biologically silenced, that is, blocked, antigen connected to a cytotoxic prodrug. Blocking of the antigen is considered relevant as it is anticipated that circulating autoantibodies will otherwise clear the antigen-prodrug before it can reach the target cell. The antigen-prodrug can only bind to the autoantigen-specific B cell receptor (BCR) upon enzymatic removal of the blocking group in close proximity of the B cell surface. BCR binding ultimately induces antigen-specific cytotoxicity after internalization of the antigen. We have synthesized a cyclic citrullinated peptide (CCP) antigen suitable for BCR binding and demonstrated that binding by ACPA was impaired upon introduction of a carboxy-p-nitrobenzyl (CNBz) blocking group at the side chain of the citrulline residue. Enzymatic removal of the CNBz moiety by nitroreductase fully restored citrulline-specific recognition by both ACPA and ACPA-expressing B cells and showed targeted cell death of CCP-recognizing B cells only. These results mark an important step toward antigen-specific B cell targeting in general and more specifically in RA, as successful blocking and activation of citrullinated antigens forms the basis for subsequent use of such construct as a prodrug in the context of autoimmune diseases.

Multifunctional, Multivalent PIC Polymer Scaffolds for Targeting Antigen-Specific, Autoreactive B Cells

“…Combining PICs with antigen-shielding protection groups that can be locally unlocked by enzymes could solve this issue. 28 Further studies are needed to investigate the PIC inhibitory effects, activation and inhibitory kinetics, and antigen-shielding strategies. Together, our data illustrate the application of PIC conjugates in the immunophenotyping and immunomodulation of rare B-cell populations.…”

Cyclic Citrullinated Peptide Aptamer Treatment Attenuates Collagen-Induced Arthritis

“…Upon binding of ACPA ligand by B-cells, the carboxy- p -nitrobenzyl moiety was enzymatically removed by nitroreductase. This fully restored the cytotoxic activity of the drug, leading to targeted cell death of citrullinated peptide-recognizing B-cells specifically …”

“…Binding of an aptamer by ACPA is expected to prevent their mode of action . Utilizing citrullinated peptides as scavengers for ACPA has previously been demonstrated in vitro . In this study, ACPA-expressing B-cells were treated with a cyclic citrullinated peptide antigen connected to a cytotoxic prodrug.…”

Anti-citrullinated Protein Antibody Generation, Pathogenesis, Clinical Application, and Prospects

“…Therefore, the curative effects are uncertain. Moreover, these findings are based on animal studies ( 84 ).…”