PB0805 - Development of Personalized CRISPR/Cas9 Based Gene Correction Therapy for Von Willebrand Disease

Monday, July 11, 2022

6:30 PM – 7:30 PM

Location: Exhibition

Presentation Format: In-Person

Presenting Author(s)

Isabel Bär

PhD candidate
Department of Hematology, Erasmus University Medical Centre, Rotterdam, the Netherlands
Rotterdam, Zuid-Holland, Netherlands

Co-Author(s)

PB

Petra E. Bürgisser

Research Technician
Department of Hematology, Erasmus University Medical Center Rotterdam
Rotterdam, Zuid-Holland, Netherlands
Ruben Bierings, PhD

Principal investigator
Department of Hematology, Erasmus MC, University Medical Center Rotterdam, the Netherlands
Rotterdam, Zuid-Holland, Netherlands
JE

Jeroen C.J. Eikenboom

Professor, Internist, Head of Div. T&H
Department of Internal Medicine, Division of Thrombosis and Hemostasis, Einthoven Laboratory for Vascular and Regenerative Medicine, Leiden University Medical Center, Leiden, The Netherlands
Leiden, Zuid-Holland, Netherlands
Cv

Calvin B. van Kwawegen

PhD-candidate
Department of Hematology, Erasmus MC, Erasmus University Medical Center Rotterdam, Rotterdam, the Netherlands
Rotterdam, Zuid-Holland, Netherlands
Frank W.G. Leebeek, MD, PhD

Professor and Head of Department of Hematology
Erasmus MC, University Medical Center Rotterdam, The Netherlands
Rotterdam, Zuid-Holland, Netherlands
JV

Jan Voorberg

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

Background: Von Willebrand Disease (VWD) is the most common inherited bleeding disorder. Current treatment options, including desmopressin/DDAVP administration or VWF-concentrate therapy, provide no sufficient long-term solutions. CRISPR/Cas9 gene targeting approaches present the opportunity to selectively target and correct causative mutations in VWF. Endothelial colony forming cells (ECFCs) from VWD patients are a powerful translational cell model of VWD and could potentially be used for demonstrating VWF gene correction ex vivo.

Aims: The aim of this study is to set a ground for personalized gene therapy in VWD by developing CRISPR/Cas9 gene correction methods in VWD patient-derived ECFCs.

Methods: The Willebrand in the Netherlands (WiN) cohort study characterized >800 VWD patients. ECFCs were isolated from venous blood of selected VWD patients who enrolled in the BOEC-MK study. Cellular phenotypes are determined using biochemical assays for VWF synthesis, secretion, and multimer composition and using confocal imaging of VWF storage in Weibel-Palade-bodies. VWD mutations will be corrected using a Cas9-based cytosine base editor in conjunction with specific guide RNAs directed against the site of mutation. Single cell sorted ECFCs will be clonally expanded and will be evaluated for rescue of the disease phenotype. Correction of the mutation will be confirmed using DNA sequencing.

Results: A panel of causative VWF mutations has been selected for gene correction. Corresponding patient ECFCs have been isolated and their baseline disease phenotypes are being characterized using morphological and biochemical assays. ECFCs are currently being immortalized to overcome their limited proliferation ability.

Conclusion(s): Gene therapy is a promising treatment alternative for patients suffering from VWD. More studies are needed to evaluate which correction strategies are suitable for efficient and durable correction of VWF mutations in VWD.

Funding was received from the Netherlands Organization for Scientific Research (NWO), Domain Applied and Engineering Sciences (TTW), ‘Connecting Innovators’ Open Technology Programme, Project#18712