mdugarte

• Roman, Giacomo; Lauritzen, Knut Husø; Harrison, Sean Philip; Bhattacharya, Anindita; Andresen, Marianne Seierstad & Mowinckel, Marie-Christine [Show all 16 contributors for this article] (2025). Ex vivo correction of severe coagulation Factor VII deficiency in patient-derived 3D liver organoids. Haematologica. ISSN 0390-6078. doi: 10.3324/haematol.2025.288046. Full text in Research Archive Show summary

  Coagulation factor (F) VII deficiency is the most frequent among the rare, inherited bleeding disorders and is predominantly caused by missense mutations in the F7 gene. The disease phenotype ranges from asymptomatic cases to extremely severe hemorrhagic forms, requiring prophylactic injections with plasma-derived or recombinant FVII concentrates. In response, we have developed an autologous cell-based approach that corrects the disease-causing mutation in patient-derived induced pluripotent stem cells (iPSCs) and generates therapeutic, three-dimensional hepatic organoids (HOs). We report the CRISPRmediated correction of homozygous c.718G>C (p.G240R), a missense mutation associated with a severe, life-threatening bleeding phenotype. The HOs contain all liver cell types and exhibit key liver functions, including coagulation factor production. After correction, our data indicate that the patient-derived HOs secrete consistent amounts of functional FVII protein, resulting in improved thrombin generation times. These results represent a significant milestone toward the establishment of an autologous cell-based therapy for patients with FVII- and other coagulation factor deficiencies.

• Andresen, Marianne Seierstad; Mowinckel, Marie-Christine; Skarpen, Ellen; Andersen, Elisabeth; Sandset, Per Morten & Dugarte, Maria Eugenia Chollet [Show all 7 contributors for this article] (2025). Identification and evaluation of Pharmacological enhancers of the factor VII p.Q160R variant. Scientific Reports. 15(1). doi: 10.1038/s41598-025-98689-3. Full text in Research Archive
• Westerberg, Niklas Starck; Atneosen-Åsegg, Monica; Melheim, Maria; Dugarte, Maria Eugenia Chollet; Harrison, Sean Philip & Siller, Richard [Show all 8 contributors for this article] (2024). Effect of hypoxia on aquaporins and hepatobiliary transport systems in human hepatic cells. Pediatric Research. ISSN 0031-3998. doi: 10.1038/s41390-024-03368-0. Full text in Research Archive
• Harrison, Sean Philip; Baumgarten, Saphira Felicitas; Dugarte, Maria Eugenia Chollet; Stavik, Benedicte; Bhattacharya, Anindita & Almaas, Runar [Show all 7 contributors for this article] (2024). Parenteral nutrition emulsion inhibits CYP3A4 in an iPSC derived liver organoids testing platform. Journal of Pediatric Gastroenterology and Nutrition - JPGN. ISSN 0277-2116. 78, p. 1047–1058. doi: 10.1002/jpn3.12195. Full text in Research Archive Show summary

  Objectives Parenteral nutrition (PN) is used for patients of varying ages with intestinal failure to supplement calories. Premature newborns with low birth weight are at a high risk for developing PN associated liver disease (PNALD) including steatosis, cholestasis, and gallbladder sludge/stones. To optimize nutrition regimens, models are required to predict PNALD. Methods We have exploited induced pluripotent stem cell derived liver organoids to provide a testing platform for PNALD. Liver organoids mimic the developing liver and contain the different hepatic cell types. The organoids have an early postnatal maturity making them a suitable model for premature newborns. To mimic PN treatment we used medium supplemented with either clinoleic (80% olive oil/20% soybean oil) or intralipid (100% soybean oil) for 7 days. Results Homogenous HNF4a staining was found in all organoids and PN treatments caused accumulation of lipids in hepatocytes. Organoids exhibited a dose dependent decrease in CYP3A4 activity and expression of hepatocyte functional genes. The lipid emulsions did not affect overall organoid viability and glucose levels had no contributory effect to the observed results. Conclusions Liver organoids could be utilized as a potential screening platform for the development of new, less hepatotoxic PN solutions. Both lipid treatments caused hepatic lipid accumulation, a significant decrease in CYP3A4 activity and a decrease in the RNA levels of both CYP3A4 and CYP1A2 in a dose dependent manner. The presence of high glucose had no additive effect, while Clinoleic at high dose, caused significant upregulation of interleukin 6 and TLR4 expression.

• Roman, Giacomo; Stavik, Benedicte; Lauritzen, Knut Husø; Sandset, Per Morten; Harrison, Sean & Sullivan, Gareth John [Show all 7 contributors for this article] (2023). “iPSC-derived liver organoids and inherited bleeding disorders: Potential and future perspectives”. Frontiers in Physiology. 14. doi: 10.3389/fphys.2023.1094249. Full text in Research Archive Show summary

  The bleeding phenotype of hereditary coagulation disorders is caused by the low or undetectable activity of the proteins involved in hemostasis, due to a broad spectrum of genetic alterations. Most of the affected coagulation factors are produced in the liver. Therefore, two-dimensional (2D) cultures of primary human hepatocytes and recombinant overexpression of the factors in non-human cell lines have been primarily used to mimic disease pathogenesis and as a model for innovative therapeutic strategies. However, neither human nor animal cells fully represent the hepatocellular biology and do not harbor the exact genetic background of the patient. As a result, the inability of the current in vitro models in recapitulating the in vivo situation has limited the studies of these inherited coagulation disorders. Induced Pluripotent Stem Cell (iPSC) technology offers a possible solution to overcome these limitations by reprogramming patient somatic cells into an embryonic-like pluripotent state, thus giving the possibility of generating an unlimited number of liver cells needed for modeling or therapeutic purposes. By combining this potential and the recent advances in the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 technology, it allows for the generation of autologous and gene corrected liver cells in the form of three-dimensional (3D) liver organoids. The organoids recapitulate cellular composition and organization of the liver, providing a more physiological model to study the biology of coagulation proteins and modeling hereditary coagulation disorders. This advanced methodology can pave the way for the development of cell-based therapeutic approaches to treat inherited coagulation disorders. In this review we will explore the use of liver organoids as a state-of-the-art methodology for modeling coagulation factors disorders and the possibilities of using organoid technology to treat the disease.

• Harrison, Sean Philip; Siller, Richard; Tanaka, Yoshiaki; Dugarte, Maria Eugenia Chollet; Morena-Barrio, María Eugenia de la & Xiang, Yangfei [Show all 36 contributors for this article] (2023). Scalable production of tissue-like vascularized liver organoids from human PSCs. Experimental and Molecular Medicine. ISSN 1226-3613. 55, p. 2005–2024. doi: 10.1038/s12276-023-01074-1. Full text in Research Archive Show summary

  Abstract The lack of physiological parity between 2D cell culture and in vivo culture has led to the development of more organotypic models, such as organoids. Organoid models have been developed for a number of tissues, including the liver. Current organoid protocols are characterized by a reliance on extracellular matrices (ECMs), patterning in 2D culture, costly growth factors and a lack of cellular diversity, structure, and organization. Current hepatic organoid models are generally simplistic and composed of hepatocytes or cholangiocytes, rendering them less physiologically relevant compared to native tissue. We have developed an approach that does not require 2D patterning, is ECM independent, and employs small molecules to mimic embryonic liver development that produces large quantities of liver-like organoids. Using single-cell RNA sequencing and immunofluorescence, we demonstrate a liver-like cellular repertoire, a higher order cellular complexity, presenting with vascular luminal structures, and a population of resident macrophages: Kupffer cells. The organoids exhibit key liver functions, including drug metabolism, serum protein production, urea synthesis and coagulation factor production, with preserved post-translational modifications such as N-glycosylation and functionality. The organoids can be transplanted and maintained long term in mice producing human albumin. The organoids exhibit a complex cellular repertoire reflective of the organ and have de novo vascularization and liver-like function. These characteristics are a prerequisite for many applications from cellular therapy, tissue engineering, drug toxicity assessment, and disease modeling to basic developmental biology.

• Andersen, Elisabeth; Dugarte, Maria Eugenia Chollet; Bernardi, Francesco; Branchini, Alessio; Baroni, Marcello & Mariani, Guglielmo [Show all 12 contributors for this article] (2021). The factor vii variant p.A354v-p.p464hfs: Clinical versus intracellular and biochemical phenotypes induced by chemical chaperones. Applied Sciences. 11(13). doi: 10.3390/app11135762. Full text in Research Archive Show summary

  (1) Background: Congenital factor (F) VII deficiency is caused by mutations in the F7 gene. Patients with modest differences in FVII levels may display large differences in clinical severity. The variant p.A354V-p.P464Hfs is associated with reduced FVII antigen and activity. The aim of the study was to investigate the clinical manifestation of this variant and the underlying molecular mechanisms. (2) Methods: Analyses were conducted in 37 homozygous patients. The recombinant variant was produced in mammalian cells. (3) Results: We report a large variation in clinical phenotypes, which points out genetic and acquired components beyond F7 mutations as a source of variability. In contrast, patients displayed similarly reduced FVII plasma levels with antigen higher than its activity. Comparative analysis of the recombinant variant and of plasma samples from a subset of patients indicated the presence of an elongated variant with indistinguishable migration. Treatment of cells with the chemical chaperone 4-phenylbutyrate (4-PBA) improved the intracellular trafficking of the variant and increased its secretion to the conditioned medium up to 2-fold. However, the effect of 4-PBA on biological activity was marginal. (4) Conclusions: Chemical chaperones can be used as biochemical tools to study the intracellular fate of a trafficking-defective FVII variant.

• Andersen, Elisabeth; Dugarte, Maria Eugenia Chollet; Sletten, Marit; Stavik, Benedicte; Skarpen, Ellen & Backe, Paul Hoff [Show all 9 contributors for this article] (2021). Molecular Characterization of Two Homozygous Factor VII Variants Associated with Intracranial Bleeding. Thrombosis and Haemostasis. ISSN 0340-6245. 121(12), p. 1588–1598. doi: 10.1055/a-1450-8568. Full text in Research Archive
• Sharma, Kulbhushan; Asp, Nagham Theres; Harrison, Sean; Siller, Richard; Baumgarten, Saphira Felicitas & Gupta, Swapnil [Show all 10 contributors for this article] (2021). Autophagy modulates cell fate decisions during lineage commitment. Autophagy. ISSN 1554-8627. p. 1–17. doi: 10.1080/15548627.2021.2008691. Full text in Research Archive Show summary

  Early events during development leading to exit from a pluripotent state and commitment toward a specific germ layer still need in depth understanding. Autophagy has been shown to play a crucial role in both development and differentiation. This study employs human embryonic and induced pluripotent stem cells to understand the early events of lineage commitment with respect to the role of autophagy in this process. Our data indicate that a dip in autophagy facilitates exit from pluripotency. Upon exit, we demonstrate that the modulation of autophagy affects SOX2 levels and lineage commitment, with induction of autophagy promoting SOX2 degradation and mesendoderm formation, whereas inhibition of autophagy causes SOX2 accumulation and neuroectoderm formation. Thus, our results indicate that autophagy-mediated SOX2 turnover is a determining factor for lineage commitment. These findings will deepen our understanding of development and lead to improved methods to derive different lineages and cell types.

• Harrison, Sean; Siller, Richard; Tanaka, Yoshiaki; Xiang, Yangfei; Patterson, Benjamin & Kempf, Henning [Show all 29 contributors for this article] (2020). Scalable production of tissue-like vascularised liver organoids from human PSCs. BioRxiv. ISSN 0362-4331. doi: 10.1101/2020.12.02.406835. Full text in Research Archive
• Andersen, Elisabeth; Dugarte, Maria Eugenia Chollet; Baroni, Marcello; Pinotti, Mirko; Bernardi, Francesco & Skarpen, Ellen [Show all 8 contributors for this article] (2019). The effect of the chemical chaperone 4-phenylbutyrate on secretion and activity of the p.Q160R missense variant of coagulation factor FVII. Cell & Bioscience. 9:69, p. 1–10. doi: 10.1186/s13578-019-0333-8. Full text in Research Archive Show summary

  Background Congenital coagulation factor (F) VII deficiency is a rare bleeding disorder caused by mutations in the F7 gene. The missense factor FVII variant p.Q160R is the disease-causing mutation in all Norwegian FVII deficient patients and results in reduced biological activity and antigen levels of FVII in patient plasma. Previous in vitro studies on this variant demonstrated impaired intracellular trafficking and reduced secretion, possibly due to protein misfolding. The aim of the study was therefore to assess the impact of chemical chaperones on cellular processing and secretion of this variant using a cell model based on overexpression of the recombinant protein. Results Through screening of compounds, we identified 4-phenylbutyrate (4-PBA) to increase the secretion of recombinant (r) FVII-160R by ~ 2.5-fold. Additionally, treatment with 4-PBA resulted in a modest increase in specific biological activity. Intracellular localization studies revealed that upon treatment with 4-PBA, rFVII-160R was secreted through Golgi and Golgi reassembly-stacking protein (GRASP)-structures. Conclusions The present study demonstrates that the chemical chaperone 4-PBA, restores intracellular trafficking and increases the secretion of a missense FVII variant with functional properties in the extrinsic coagulation pathway.

• Andersen, Elisabeth; Dugarte, Maria Eugenia Chollet; Myklebust, Christiane Filion; Pinotti, Mirko; Bernardi, Francesco & Chuansumrit, Ampaiwan [Show all 9 contributors for this article] (2018). Activation of Endoplasmic Reticulum Stress and Unfolded Protein Response in Congenital Factor VII Deficiency. Thrombosis and Haemostasis. ISSN 0340-6245. 118(4), p. 664–675. doi: 10.1055/s-0038-1637690. Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; Andersen, Elisabeth; Skarpen, Ellen; Myklebust, Christiane Filion; Koehler, Christian & Morth, Jens Preben [Show all 12 contributors for this article] (2018). Factor VII deficiency: Unveiling the cellular and molecular mechanisms underlying Three model alterations of the enzyme catalytic domain. Biochimica et Biophysica Acta - Molecular Basis of Disease. ISSN 0925-4439. 1864(3), p. 660–667. doi: 10.1016/j.bbadis.2017.12.016. Full text in Research Archive
• Chollett, Maria Eugenia; Skarpen, Ellen; Iversen, Nina; Sandset, Per Morten & Skretting, Grethe (2015). The chemical chaperone sodium 4-phenylbutyrate improves the secretion of the protein CA267T mutant in CHO-K1 cells through the GRASP55 pathway. Cell & Bioscience. doi: 10.1186/s13578-015-0048-4. Full text in Research Archive Show summary

  Some inherited coagulation factor deficiencies are caused by intracellular retention or degradation of misfolded proteins, and chemical chaperones have been shown to reverse protein misfolding. The purpose of the present study was to investigate whether chemical chaperones may improve secretion of the protein CA267T (PCA267T) mutant in a cellular model. Using stably transfected Chinese hamster ovary cells (CHO-K1) expressing PCA267T we demonstrate that sodium 4-phenylbutyrate (PBA) increased the secretion of PCA267T by approximately 4-fold in comparison with untreated cells, and that this secretion seemed to follow an unconventional pathway via the Golgi reassembly stacking protein (GRASP55).

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• Dugarte, Maria Eugenia Chollet; Bhattacharya, Anindita; Roman, Giacomo; Harrison, Sean Philip; Baumgarten, Saphira Felicitas & Andersen, Elisabeth [Show all 12 contributors for this article] (2025). Autologous gene-corrected hepatic organoids for the treatment of FVII deficiency. Research and Practice in Thrombosis and Haemostasis (RPTH). 9(S2), p. 265–266. doi: 10.1016/j.rpth.2025.102925. Full text in Research Archive
• Stavik, Benedicte; Bhattacharya, Anindita; Lauritzen, Knut Husø; Baumgarten, Saphira Felicitas; barrio, maria eugenia & corral, javier [Show all 10 contributors for this article] (2025). Human serum enhances the expression and production of antithrombin by hepatic organoids. Research and Practice in Thrombosis and Haemostasis (RPTH). 9(S2), p. 405–406. doi: 10.1016/j.rpth.2025.102923. Full text in Research Archive
• Roman, Giacomo; Lauritzen, Knut Husø; Harrison, Sean Philip; Bhattacharya, Anindita; Mowinckel, Marie-Christine & Andresen, Marianne Seierstad [Show all 14 contributors for this article] (2025). Gene editing in patient-derived iPSCs restores coagulation FVII deficiency in 3D liver organoids. Research and Practice in Thrombosis and Haemostasis (RPTH). 9(S2), p. 950–951. doi: 10.1016/j.rpth.2025.102923. Full text in Research Archive
• Lauritzen, Knut Husø; Bhattacharya, Anindita; Dugarte, Maria Eugenia Chollet; Sandset, Per Morten & Stavik, Benedicte (2024). Mitochondrial maturation in hepatic organoids. Full text in Research Archive
• Bhattacharya, Anindita; Roman, Giacomo; Harrison, Sean Philip; Sullivan, Gareth John; Mowinckel, Marie-Christine & Sandset, Per Morten [Show all 10 contributors for this article] (2024). Developing cell-based genome editing approach for antithrombin deficiency: a therapeutic prospect. Research and Practice in Thrombosis and Haemostasis (RPTH). doi: 10.1016/j.rpth.2024.102504. Full text in Research Archive
• Andresen, Marianne Seierstad; Mowinckel, Marie-Christine; Skarpen, Ellen; Andersen, Elisabeth; Sandset, Per Morten & Stavik, Benedicte [Show all 7 contributors for this article] (2024). Verification of two Pharmacological Enhancers of Factor VII Activity in the FVII p.Q160R variant. Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; Bhattacharya, Anindita; Roman, Giacomo; Harrison, Sean; Baumgarten, Saphira Felicitas & Andersen, Elisabeth [Show all 12 contributors for this article] (2024). Autologous gene-corrected stem cell-derived hepatic organoids for the treatment of FVII deficiency. HemaSphere. doi: 10.1002/hem3.104. Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; Andersen, Elisabeth; Harrison, Sean Philip; Morena-Barrio, Maria Eugenia de la; Baumgarten, Saphira Felicitas & Bhattacharya, Anindita [Show all 14 contributors for this article] (2023). OC 07.5 Modeling and Correcting FVII Deficiency using Human Stem Cell-Derived Hepatic Organoids. Research and Practice in Thrombosis and Haemostasis (RPTH). 7(Oral communications ISTH 2023). doi: 10.1016/j.rpth.2023.100470. Full text in Research Archive
• Roman, Giacomo; Lauritzen, Knut Husø; Harrison, Sean Philip; Sullivan, Gareth John; Iversen, Nina & Henriksson, Carola Elisabeth [Show all 10 contributors for this article] (2023). OC 07.1 Autologous CRISPR/Cas9 Correction of a Missense Variant from a Patient with Severe, Inherited FVII Deficiency. Research and Practice in Thrombosis and Haemostasis (RPTH). 7(Oral communications ISTH 2023). doi: 10.1016/j.rpth.2023.100475. Full text in Research Archive
• Roman, Giacomo; Lauritzen, Knut Husø; Harrison, Sean; Sullivan, Gareth John; Iversen, Nina & Henriksson, Carola Elisabeth [Show all 10 contributors for this article] (2023). Autologous CRISPR/Cas9 correction of a missense variant from a patient with severe, inherited FVII deficiency. Full text in Research Archive
• Roman, Giacomo; Lauritzen, Knut Husø; Harrison, Sean Philip; Sullivan, Gareth John; Iversen, Nina & Henriksson, Carola Elisabeth [Show all 10 contributors for this article] (2023). OC 07.1 Autologous CRISPR/Cas9 Correction of a Missense Variant from a Patient with Severe, Inherited FVII Deficiency. Research and Practice in Thrombosis and Haemostasis (RPTH). doi: 10.1016/j.rpth.2023.100475. Full text in Research Archive
• Andresen, Marianne Seierstad; Mowinckel, Marie-Christine; Stavik, Benedicte; Sandset, Per Morten & Dugarte, Maria Eugenia Chollet (2022). Evaluation of Pharmacological Enhancers of Mutated Factor VII Activity ex vivo. Research and Practice in Thrombosis and Haemostasis (RPTH). Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; Harrison, Sean; Baumgarten, Saphira Felicitas; Morena-Barrio, Maria Eugenia de la; Bravo-Pérez, Carlos & Mowinckel, Marie-Christine [Show all 12 contributors for this article] (2022). Human iPSC-derived hepatic organoids – a model for blood coagulation research and therapy. Human Gene Therapy. ISSN 1043-0342. doi: 10.1089/hum.2022.29225.abstracts. Full text in Research Archive
• andersen, elisabeth; Mowinckel, Marie-Christine; Stavik, Benedicte; Sandset, Per Morten & Dugarte, Maria Eugenia Chollet (2021). High-throughput Screening for Pharmacological Enhancers of Mutated Factor VII Activity. Research and Practice in Thrombosis and Haemostasis (RPTH). 5. Full text in Research Archive
• andersen, elisabeth; Dugarte, Maria Eugenia Chollet; Sletten, Marit; Stavik, Benedicte; Myklebust, Christiane Fillion & Skarpen, Ellen [Show all 11 contributors for this article] (2020). Molecular Characterization of Two Homozygous Factor VII Variants Associated with Intracranial Bleeding. Blood. ISSN 0006-4971. 136. doi: 10.1182/blood-2020-139219. Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; andersen, elisabeth; Morena-Barrio, Maria Eugenia de la; Bravo-Perez, Carlos; Mowinckel, Marie-Christine & Myklebust, Christiane Fillion [Show all 12 contributors for this article] (2020). Characterizing Coagulation FVII from iPSC-Hepatocytes-like Cells: Setting the Basis for Cell Therapy Development. Blood. ISSN 0006-4971. 136. doi: 10.1182/blood-2020-139211. Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; Andersen, Elisabeth; Skarpen, Ellen; Mowinckel, Marie-Christine; Stavik, Benedicte & Sullivan, Gareth [Show all 7 contributors for this article] (2019). FVII: A human pluripotent stem cell derived hepatocyte model with potential therapeutic applications. Research and Practice in Thrombosis and Haemostasis (RPTH). 3. Full text in Research Archive

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