Selective Elimination of Autoreactive B Cells Using Multivalent Polyisocyanopeptide-Based Antigen-Toxin Conjugates

Biomacromolecules, vol. 27, pp. 4198–4212

Abstract

Rheumatoid arthritis (RA) is a debilitating autoimmune disease characterized by chronic inflammation and joint damage. Current treatments rely on systemic immune suppressors, which increases infection risk. Autoreactive B cells directed against citrullinated antigens and producing anticitrullinated protein antibodies (ACPAs) are thought to play a key role in RA pathogenesis, making their selective elimination a promising therapeutic strategy. Previous research demonstrated that a synthetic multivalent citrullinated antigen, cyclic citrullinated peptide 4 (CCP4), is efficiently internalized by these autoreactive B cells through an antigen-specific B cell receptor (BCR), positioning it as a potential delivery vehicle for targeted therapy. Here, we explored the elimination of autoreactive B cells using multivalent antigen-cleavable linker–toxin conjugates. Initial evaluation of diCCP4-VCP-MMAE revealed that it lacked sufficient toxicity toward ACPA BCR-expressing Ramos cells despite cathepsin-mediated linker cleavage and the absence of steric hindrance from diCCP4 or the BCR. Therefore, we investigated polyisocyano peptide (PIC) polymers, which allow for a higher CCP4 and toxin density. PIC 13 (containing 13 CCP4 units), functionalized with 0.5 equiv of MMAE per CCP4, selectively eliminated Ramos 3F3 cells with a midnanomolar IC 50 . Increasing MMAE to 1, 2, or 5 equiv further reduced the IC 50 to low nanomolar. PIC 13 + 5 equiv MMAE demonstrated selective toxicity in Ramos cells expressing various ACPA BCRs, as well as patient-derived, immortalized ACPA-expressing B cells. These findings underscore the importance of optimizing delivery modules and toxin ratios and highlight PIC-based antigen-toxin conjugates as a promising strategy for selectively eliminating autoreactive B cells in RA.

Authors 9

1. Kevin R. Venrooij

   Radboud University Nijmegen

   Affiliation as printed

   Institute for Molecules and Materials

   Radboud University

2. Margot J. van Weijsten

   Radboud University Nijmegen

   Affiliation as printed

   Institute for Molecules and Materials

   Radboud University

3. Sanne Kroos Aachen

   Leiden University Medical Center

   Affiliation as printed

   Department of Rheumatology

   Leiden University Medical Center

4. Roel Hammink

   Radboud University Medical Center

   Affiliation as printed

   Department of Medical BioSciences

   Radboudumc

5. F. Turlings Aachen

   Leiden University Medical Center

   Affiliation as printed

   Department of Immunology

   Leiden University Medical Center

6. Carl Gustav Figdor

   Radboud University Medical Center

   Affiliation as printed

   Department of Medical BioSciences

   Radboudumc

7. Martijn Verdoes Aachen

   Leiden University Medical Center · Radboud University Medical Center

   Affiliation as printed

   Department of Immunology

   Department of Medical BioSciences

   Leiden University Medical Center

   Radboudumc

8. René E. M. Toes Aachen

   Leiden University Medical Center

   Affiliation as printed

   Department of Rheumatology

   Leiden University Medical Center

9. Kimberly M. Bonger corresponding Aachen Leiden Institute of Chemistry

   Leiden University · Radboud University Nijmegen

   Affiliation as printed

   Institute for Molecules and Materials

   Leiden Institute of Chemistry

   Leiden University

   Radboud University