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    Oral Communication

    Hemophilia and Rare Bleeding Disorders

    Rare Bleeding Disorders I

    OC 07.5 - Modeling and correcting FVII deficiency using human stem cell-derived hepatic organoids

    Saturday, June 24, 2023
    14:00 – 14:15 ET
    Room: 524
    Session Available As: On-Demand

      Presenting Author(s)

    • Maria Eugenia Chollet Dugarte, MD, PhD (she/her/hers)

      Post doctoral researcher
      Department of Hematology, Research Institute of Internal Medicine, Oslo University Hospital
      Oslo, Oslo, Norway

      • Co-Author(s)

    • EA

      Elisabeth Andersen

      Post doc
      Oslo University Hospital
      Oslo, Oslo, Norway

    • SH

      Sean Harrison

      Post doc
      Oslo University Hospital
      Oslo, Oslo, Norway

    • Maria Eugenia de la Morena-Barrio, PhD

      Reseacher
      Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, CIBERER, Murcia, Spain
      Murcia, Murcia, Spain

    • SB

      Saphira Baumgarten

      PhD student
      University of Oslo
      Oslo, Oslo, Norway

    • AB

      Anindita Bhattacharya

      Post doc
      University of Oslo
      Oslo, Oslo, Norway

    • Giacomo Roman, MSc

      PhD student
      Department of Hematology, Oslo University Hospital, Oslo, Norway
      Oslo, Oslo, Norway

    • MA

      Mariane Andresen

      Senior laboratory technician
      Oslo University Hospital
      Oslo, Oslo, Norway

    • CB

      Carlos Bravo

      Post doc
      Department of Hematology and Oncology, University Hospital Morales Meseguer, Centro Regional de Hemodonación, University of Murcia, IMIB, CIBERER, Murcia, Spain
      Murcia, Murcia, Spain

    • MM

      Marie-Christine Mowinckel

      Senior laboratory technician
      Oslo University Hospital
      Oslo, Oslo, Norway

    • Javier Corral, PhD

      PhD
      Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
      Murcia, Murcia, Spain

    • PS

      Per Morten Sandset

      MD, PhD, Vice-rector University of Oslo
      University of Oslo
      Oslo, Oslo, Norway

    • GS

      Gareth J Sullivan

      Group leader
      Department of Pediatric Research, Oslo University Hospital, Oslo, Norway
      Oslo, Oslo, Norway

    • BS

      Benedicte Stavik

      Group leader
      Department of Hematology, Oslo University Hospital, Oslo, Norway
      Oslo, Oslo, Norway

    Background: Characterizing coagulation factors and their deficiencies is crucial for understanding the disease and for developing new treatments, but physiological relevant models are lacking. We have developed a protocol for generation of autologous liver organoids from patient-derived induced pluripotent stem cells (iPSCs).

    Aims: To evaluate the iPSCs-derived hepatic organoids (HOs) as a model for the study of coagulation factors, Factor (F) VII deficiency, and treatment development.

    Methods: Ethically sourced control and patient derived iPSCs carrying the FVII variant p.Q160R, were differentiated into HOs using our small molecule-based protocol. Expression of coagulation factors was characterized using qPCR, ELISA, WB, proteomics and digestion with N-glycosylase F (PNGaseF). FVII activity was assessed by chromogenic and thrombin generation assays (TGA) benchmarked against primary hepatocytes (PH). The p.Q160R variant was corrected using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR/CAS9) in patient’s iPSCs.

    Results: The HOs expressed hepatic coagulation factors FII, FVII, FX, Protein C, Protein S and Antithrombin at both mRNA and protein level. Secreted FVII levels measured around 70% of that secreted by PH, and comparable FVII activity was detected. TGA showed that the HOs cell medium shortened the lag time of FVII deficient plasma (24,7 vs 4,7 min). Digestion with the PNGaseF indicated that FVII is glycosylated. HOs carrying the p.Q160R variant showed a reduced secretion of FVII and undetectable FVII activity, similar to the clinical phenotype ( < 1%), and did not shorten the lag time of FVII deficient plasma. Homozygous correction of the p.Q160R mutation was successfully performed in the patient’ derived iPSCs.

    Conclusion(s): This iPSCs-HO model showed the production of FVII at physiologically relevant levels with similarities to plasma FVII. The iPSCs-HOs carrying the FVII mutation mimic the patients’ phenotype. The successful correction of the FVII mutation in the patient’ iPSCs provides a therapeutic potential.

    Learning Objectives:

    • Upon completion, participants will be able to evaluate the applicability of human stem cell-derived hepatic organoids in modeling FVII deficiency
    • Upon completion, participants will be able to identify the characteristics of FVII protein synthetized by human stem cell-derived hepatic organoids
    • Upon completion, participants will be able to estimate the therapeutic potential of human stem cell-derived hepatic organoids in factor VII deficiency