Inhibition of lysosomal degradation increases expression of mutant ADA2 in DADA2 monocytes

Lisa Ehlers; Marjon Wouters; Bethany Pillay; Selket Delafontaine; Mariia Dzhus; Marco Baggio; Tim Niehues; Gregor Dückers; Lieve Sevenants; Kristina Casteels; Lien De Somer; Rik Schrijvers; Steven Vanderschueren; Maarten Jacquemyn; Dirk Daelemans; Anneleen Hombrouck; Eugene P Chambers; Thomas Tousseyn; Giorgia Bucciol; Patrizia Agostinis; Leen Moens; Isabelle Meyts

doi:10.1016/j.jaci.2025.06.009

Inhibition of lysosomal degradation increases expression of mutant ADA2 in DADA2 monocytes

J Allergy Clin Immunol. 2025 Jun 20:S0091-6749(25)00651-7. doi: 10.1016/j.jaci.2025.06.009. Online ahead of print.

Authors

Lisa Ehlers¹, Marjon Wouters², Bethany Pillay², Selket Delafontaine³, Mariia Dzhus², Marco Baggio⁴, Tim Niehues⁵, Gregor Dückers⁵, Lieve Sevenants⁶, Kristina Casteels⁶, Lien De Somer⁷, Rik Schrijvers⁸, Steven Vanderschueren⁹, Maarten Jacquemyn¹⁰, Dirk Daelemans¹⁰, Anneleen Hombrouck², Eugene P Chambers¹¹, Thomas Tousseyn¹², Giorgia Bucciol³, Patrizia Agostinis¹³, Leen Moens², Isabelle Meyts¹⁴

Affiliations

¹ Department of Microbiology, Immunology and Transplantation, Laboratory for Inborn Errors of Immunity, KU Leuven, Leuven, Belgium; Department of Pediatric Respiratory Medicine, Immunology and Critical Care Medicine, Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Berlin, Germany; Berlin Institute of Health at Charité - Universitätsmedizin Berlin, Berlin, Germany; German Center for Child and Adolescent Health, partner site Berlin, Berlin, Germany; Deutsches Rheuma-Forschungszentrum, an Institute of the Leibniz Association, Berlin, Germany.
² Department of Microbiology, Immunology and Transplantation, Laboratory for Inborn Errors of Immunity, KU Leuven, Leuven, Belgium.
³ Department of Microbiology, Immunology and Transplantation, Laboratory for Inborn Errors of Immunity, KU Leuven, Leuven, Belgium; Department of Pediatrics, University Hospitals Leuven, KU Leuven, Leuven, Belgium.
⁴ Department of Microbiology, Immunology and Transplantation, Laboratory for Inborn Errors of Immunity, KU Leuven, Leuven, Belgium; Laboratory of Computational and Developmental Biology, Berlin Institute for Medical Systems Biology, Max-Delbrück-Centrum for Molecular Medicine in the Helmholtz Association, Berlin, Germany.
⁵ Helios Children's Hospital, Krefeld, Germany.
⁶ Department of Pediatrics, University Hospitals Leuven, KU Leuven, Leuven, Belgium.
⁷ Department of Pediatrics, University Hospitals Leuven, KU Leuven, Leuven, Belgium; Laboratory of Immunobiology, Department of Microbiology and Immunology, KU Leuven, Leuven, Belgium.
⁸ Department of Microbiology, Immunology and Transplantation, Allergy and Clinical immunology Research Group, KU Leuven, Leuven, Belgium.
⁹ Department of General Internal Medicine, Research Department Microbiology, Immunology, and Transplantation, Laboratory of Clinical Infectious and Inflammatory Disorders, University Hospitals Leuven, Leuven, Belgium.
¹⁰ KU Leuven Department of Microbiology, Immunology and Transplantation, Molecular Genetics and Therapeutics in Virology and Oncology Research Group, Rega Institute for Medical Research, Leuven, Belgium.
¹¹ Vanderbilt University Medical Center, Nashville, Tenn; DADA2 Foundation, Nashville, Tenn.
¹² Department of Pathology, University Hospitals Leuven, Leuven, Belgium.
¹³ Cell Death Research and Therapy Lab, Department of Cellular and Molecular Medicine, Center for Cancer Biology, VIB-KU Leuven, Leuven, Belgium.
¹⁴ Department of Microbiology, Immunology and Transplantation, Laboratory for Inborn Errors of Immunity, KU Leuven, Leuven, Belgium; Department of Pediatrics, University Hospitals Leuven, KU Leuven, Leuven, Belgium. Electronic address: isabelle.meyts@uzleuven.be.

PMID: 40545182
DOI: 10.1016/j.jaci.2025.06.009

Abstract

Background: Deficiency of adenosine deaminase 2 (DADA2) is an inborn error of immunity causing vasculitis and bone marrow failure. Bone marrow failure is mostly unresponsive to TNF-α inhibitors. The limited understanding of the pathomechanisms driving the disease impedes the development of new treatment options. Unlike cellular model systems expressing pathogenic ADA2 variants, primary monocytes from patients with DADA2 lack ADA2 protein expression.

Objectives: This study aimed to analyze the role of protein degradation in the pathogenesis of DADA2 and the therapeutic potential of the lysosomotropic drug hydroxychloroquine in the treatment of patients with DADA2.

Methods: ADA2 protein expression in CD14⁺ monocytes from healthy controls (n = 8) and patients with DADA2 (n = 11) was determined by Western blot after inhibition of lysosomal and proteasomal degradation, as well as after hydroxychloroquine treatment in vivo in 1 patient with DADA2. Lipidation of microtubule associated protein 1 light chain 3 beta (LC3B) was analyzed as a measure of autophagic activity. Clinical and laboratory data were recorded in cytopenic patients with DADA2 treated with hydroxychloroquine, 200 mg per day.

Results: We demonstrated that inhibition of lysosomal degradation restores ADA2 protein expression in DADA2 monocytes in vitro. DADA2 monocytes exhibited increased autophagic activity. We observed clinical improvement in 2 cytopenic patients with DADA2 who were treated with hydroxychloroquine, and we showed a concomitant increase in ADA2 protein levels in monocytes from one of these patients in vivo.

Conclusion: We identified lysosomal protein degradation of ADA2 as a pathomechanism of DADA2 and introduced hydroxychloroquine as a potential treatment option in patients with DADA2 with refractory cytopenia.

Keywords: Deficiency of ADA2; TLR9 signaling; cytopenia; hydroxychloroquine; lysosomal degradation.