OC 58.5 - N-glycan mediated shielding of the CUB domains of ADAMTS13 prevents binding of C-terminal antibodies in patients with immune-mediated TTP

Tuesday, June 25, 2024

10:30 – 10:45 ICT

Room: 211 A-D

Presenting Author(s)

Tim Postmus, MSc

PhD
Department of Molecular Hematology, Sanquin Research and Landsteiner Laboratory, Amsterdam, the Netherlands
Duivendrecht, Noord-Holland, Netherlands

Co-Author(s)

NS

Nelly Schilder

Bachelor Student
Department of Molecular Hematology, Sanquin Research and Landsteiner Laboratory, Amsterdam, the Netherlands
Volendam, Noord-Holland, Netherlands
JF

Juliana Ferreira de Santana

Post-Doc
Department of Molecular Hematology, Sanquin Research and Landsteiner Laboratory, Amsterdam, the Netherlands
Amsterdam, Noord-Holland, Netherlands
Paul Coppo

Professor
Service d'Hématologie et CNR-MAT, Hôpital Saint-Antoine, AP-HP.6, Paris, France
Paris, Ile-de-France, France
BJ

Bérangère Joly

PharmD, PhD
Université Paris Cité, EA3518, Hôpital Lariboisière
Paris, Ile-de-France, France
AV

Agnès Veyradier

Professor
Service d'Hématologie Biologique and EA3518-Institut Universitaire d'Hématologie, Groupe Hospitalier Saint Louis-Lariboisière, Assistance Publique-Hôpitaux de Paris (AP-HP), Université Paris Diderot, Paris, France
Paris, Ile-de-France, France
Karen Vanhoorelbeke

Full professor
Laboratory for Thrombosis Research, IRF Life Sciences, KU Leuven Campus Kulak Kortrijk, Kortrijk, Belgium
Kortrijk, West-Vlaanderen, Belgium
JV

Jan Voorberg

Professor
Department of Molecular Hematology, Sanquin Research and Landsteiner Laboratory, Amsterdam, the Netherlands
Amsterdam, Noord-Holland, Netherlands

Background: Immune Thrombotic Thrombocytopenic Purpura (iTTP) is a rare thrombotic disorder, with an incidence of 1.5-6 cases per million per year. It has been well established that the majority of inhibitory auto-antibodies target the spacer domain. Non-inhibitory antibodies binding to the carboxy-terminal CUB domains have been suggested to enhance the clearance of ADAMTS13 in iTTP. Furthermore, anti-CUB domains antibodies have been shown to open the conformation of ADAMTS13. An open conformation has been identified as a biomarker for acute and relapsing iTTP, further highlighting the potential relevance of anti-CUB domain antibodies for the pathogenesis of iTTP.

Aims: Evaluate whether introduction of an N-glycan in the CUB1/2 domains of ADAMTS13 can prevent binding of pathogenic anti-CUB antibodies.

Methods: A panel of N-glycan modified ADAMTS13 variants was screened against a library of 7 iTTP patient derived monoclonal antibodies that target the CUB-domains of ADAMTS13. In parallel, these N-glycan modified ADAMTS13 variants were screened against 9 plasma samples derived from iTTP patients which primarily contain antibodies directed towards the carboxy-terminal domains of ADAMTS13.

Results: Introduction of N-glycans at amino acid positions of 1251, 1255 and 1368 in the CUB1/2 domains of ADAMTS13 can effectively reduce binding of iTTP patient derived monoclonal antibodies directed towards the CUB domains. Subsequently, the reactivity of 9 plasma samples of iTTP patients containing antibodies against the carboxy-terminal domains was analyzed. To exclude the influence of anti-spacer antibodies, CUB-N-glycans were used together with previously described auto-antibody resistant spacer domain variants, ADAMTS13-NGLY3 and ADAMTS13-5ALA. For 8 out of 9 patients, introduction of CUB-N-glycans further reduced binding of auto-antibodies. On average, binding was decreased to 51% (from 83% for NGLY3) and to 31% (from 64% for FL-5ALA).

Conclusion(s): Collectively, our findings show that introduction of an N-glycan within the CUB-domain of ADAMTS13 is able to prevent the binding of anti-CUB antibodies in patients with iTTP.