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Differentiation of pluripotent stem cells to form renal organoids

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  • Publication Date:
    November 05, 2024
  • معلومة اضافية
    • Patent Number:
      12134,785
    • Appl. No:
      16/682541
    • Application Filed:
      November 13, 2019
    • نبذة مختصرة :
      A method is provided for producing renal organoids comprising nephrons, ureteric bud and vasculature and/or progenitors of these. In one embodiment, the methods includes contacting intermediate mesoderm cells with: fibroblast growth factor 9 and/or fibroblast growth factor 20 and/or fibroblast growth factor 2 and optionally, one or more selected from the group consisting of: bone morphogenic protein 7; heparin; a Wnt agonist; retinoic acid; and an RA antagonist under conditions that promote formation of vascularized renal organoids. Another embodiment includes producing mesoderm cells by sequentially contacting pluripotent stem cells with a Wnt agonist and fibroblast growth factor 9 and/or fibroblast growth factor 20 and/or fibroblast growth factor 2, followed by a relatively short re-exposure to the Wnt agonist. The renal organoids may have end uses such as for kidney repair and regeneration, bioprinting of kidneys or functional components thereof, renal cell arrays and screening compounds for nephrotoxicity.
    • Inventors:
      The University of Queensland (St. Lucia, AU)
    • Assignees:
      The University of Queensland (St. Lucia, AU)
    • Claim:
      1. A method of producing one or more renal organoids comprising: (a) differentiating intermediate mesoderm (IM) cells into nephron progenitor cells and ureteric epithelial progenitor cells by contacting the IM cells with a Wnt agonist, fibroblast growth factor 9 (FGF9), and one or more compounds selected from the group consisting of bone morphogenic protein 7 (BMP7), heparin, retinoic acid (RA), an RA analog, an RA agonist, and an RA antagonist, wherein the RA antagonist increases the relative production of nephron progenitor cells from the IM cells; and (b) culturing the nephron progenitor cells and ureteric epithelial progenitor cells under conditions that induce aggregation of the nephron progenitor cells and ureteric epithelial progenitor cells into one or more renal organoids that are at least partly vascularized and/or comprise vascular progenitors, wherein the vascularization is facilitated by conditions that promote or direct development of vascular endothelium or vascular progenitors from mesenchymal cells or tissues.
    • Claim:
      2. The method of claim 1 , wherein the RA analog or the RA agonist increases the relative production of ureteric epithelial progenitor cells from the IM cells.
    • Claim:
      3. The method of claim 1 , wherein the Wnt agonist increases the relative production of nephron progenitor cells from the IM cells.
    • Claim:
      4. The method of claim 1 , wherein the nephron progenitor cells and ureteric epithelial progenitor cells are produced synchronously or simultaneously from the IM cells.
    • Claim:
      5. The method of claim 1 , further comprising contacting posterior primitive streak cells with one or more agents that facilitate differentiation of the posterior primitive streak cells into said IM cells.
    • Claim:
      6. The method of claim 5 , further comprising contacting human pluripotent stem cells (hPSCs) with one or more agents that facilitate differentiation of the hPSCs into said posterior primitive streak cells.
    • Claim:
      7. The method of claim 1 , wherein the Wnt agonist is CHIR99021.
    • Claim:
      8. The method of claim 1 , wherein the IM cells are (i) first contacted with the Wnt agonist and (ii) then contacted with the FGF9.
    • Claim:
      9. The method of claim 8 , wherein the Wnt agonist is CHIR99021.
    • Claim:
      10. The method of claim 8 , further comprising (iii) contacting the cells resulting from step (ii) with a Wnt agonist.
    • Claim:
      11. The method of claim 10 , wherein the Wnt agonist in step (i) is CHIR99021.
    • Claim:
      12. The method of claim 10 , wherein the Wnt agonist in step (iii) is CHIR99021.
    • Claim:
      13. The method of claim 10 , wherein the Wnt agonist in step (i) is CHIR99021 and the Wnt agonist in step (iii) is CHIR99021.
    • Claim:
      14. The method of claim 13 , wherein the concentration of the Wnt agonist in step (i) is about 1-20 μM, the concentration of FGF9 is about 100-300 ng/ml, and the concentration of the Wnt agonist in step (iii) is about 1-15 μM.
    • Claim:
      15. The method of claim 14 , wherein the concentration of the Wnt agonist in step (i) is 5-15 μM.
    • Claim:
      16. The method of claim 1 , wherein the one or more compounds is heparin.
    • Claim:
      17. The method of claim 16 , wherein the IM cells are (i) first contacted with the Wnt agonist and (ii) then contacted with the FGF9 and the heparin.
    • Claim:
      18. The method of claim 17 , wherein the Wnt agonist is CHIR99021.
    • Claim:
      19. The method of claim 17 , further comprising (iii) contacting the cells resulting from step (ii) with a Wnt agonist.
    • Claim:
      20. The method of claim 19 , wherein the Wnt agonist in step (i) is CHIR99021.
    • Claim:
      21. The method of claim 19 , wherein the Wnt agonist in step (iii) is CHIR99021.
    • Claim:
      22. The method of claim 19 , wherein the Wnt agonist in step (i) is CHIR99021 and the Wnt agonist in step (iii) is CHIR99021.
    • Claim:
      23. The method of claim 22 , wherein the concentration of the Wnt agonist in step (i) is about 1-20 μM, the concentration of FGF9 is about 100-300 ng/ml, and the concentration of the Wnt agonist in step (iii) is about 1-15 μM.
    • Claim:
      24. The method of claim 23 , wherein the concentration of the Wnt agonist in step (i) is 5-15 μM.
    • Patent References Cited:
      20050244962 November 2005 Thomson et al.
      20070128174 June 2007 Kleinsek et al.
      20120116568 May 2012 Murphy et al.
      20130122536 May 2013 Osafune et al.
      20130122589 May 2013 Kimber et al.
      20130164339 June 2013 Murphy et al.
      20130190210 July 2013 Murphy et al.
      20140012407 January 2014 Murphy et al.
      20140363888 December 2014 Osafune et al.
      20160237409 August 2016 Little et al.
      101072868 November 2007
      2009508650 March 2009
      2014531204 November 2014
      2010007031 January 2010
      2012013969 February 2012
      2012168167 December 2012
      2013040087 March 2013
      2013094771 June 2013
      2014110590 July 2014
      2014197934 December 2014
      2015069619 May 2015
      2016094948 June 2016












































    • Other References:
      Barak et al., 2012, Developmental Cell, vol. 22, pp. 1191-1207 (Year: 2012). cited by examiner
      Xinaris et al. (2012, J. Americ. Soc. Nephr., vol. 23, pp. 1857-1868) (Year: 2012). cited by examiner
      Lam et al. (Jul. 2014, Semin. Nephrol., vol. 34(4), pp. 445-461). (Year: 2014). cited by examiner
      Schumacher et al. (2021, Regenerative Med., vol. 6:45, pp. 1-11) (Year: 2021). cited by examiner
      Takasato et al. (ePub Dec. 15, 2013, Nature Cell Biology, vol. 16(1), pp. 118-126 + Supplementary Data). (Year: 2013). cited by examiner
      Abu-Abed et al., “The retinoic acid-metabolizing enzyme, CYP26AI, is essential for normal hindbrain patterning, vertebral identity, and development of posterior structures,” Genes & Development, Jan. 2001, vol. 15, pp. 226-240. cited by applicant
      Briggs et al., “Integration-Free Induced Pluripotent Stem Cells Model Genetic and Neural Developmental Features of Down Syndrome Etiology,” Stem Cells, Mar. 2013, vol. 31, pp. 467-478. cited by applicant
      Brown et al., “Role for compartmentalization in nephron progenitor differentiation”< PNAS, Mar. 2013, vol. 110, No. 12, pp. 4640-4645. cited by applicant
      Brunskill et al., “Atlas of Gene Expression in the Developing Kidney at Microanatomic Resolution,” Developmental Cell, Nov. 2008, vol. 15, pp. 781-791. cited by applicant
      Brunskill et al., “Defining the Molecular Character of the Developing and Adult Kidney Podocyte,” PLoS ONE, Sep. 2011, vol. 6, No. 9, p. e24640, 12 pages. cited by applicant
      Cebrian et al., “The Number of Fetal Nephron Progenitor Cells Limits Ureteric Branching and Adult Nephron Endowment,” Cell Reports, Apr. 2014, vol. 7, pp. 127-137. cited by applicant
      Cheng et al., “Tissue Distribution, Ontogeny, and Hormonal Regulation of Xenobiotic Transporters in Mouse Kidneys,” Drug Metabolism and Disposition, Nov. 2009, vol. 37, No. 11, pp. 2178-2185. cited by applicant
      Cummings et al., “Cisplatin-Induced Renal Cell Apoptosis: Caspase 3-Dependent and -Independent Pathways,” The Journal of Pharmacology and Experimental Therapeutics, Jul. 2002, vol. 302, No. 1, pp. 8-17. cited by applicant
      Dobin et al., “STAR: ultrafast universal RNA-seq aligner,” Bioinformatics, Oct. 2012, vol. 29, No. 1, pp. 15-21. cited by applicant
      Duester, Gregg, “Retinoic Acid Synthesis and Signaling During Early Organogenesis,” Cell, Sep. 2008, vol. 134, No. 6, pp. 921-931. cited by applicant
      Floege et al., “Localization of PDGF a-receptor in the developing and mature human kidney,” Kidney International, Apr. 1997, vol. 51, No. 4, pp. 1140-1150. cited by applicant
      International Search Report for International Application No. PCT/AU2015/050798, mailed on Apr. 27, 2016, 7 pages. cited by applicant
      James et al., “Patterning of the Avian Intermediate Mesoderm by Lateral Plate and Axial Tissues,” Dev. Biol, Jan. 2003, vol. 253, pp. 109-124. cited by applicant
      Kang et al., “Differentiation of human pluripotent stem cells into nephron progenitor cells in a serum and feeder free system,” PLoS One, Apr. 2014, vol. 9, p. e94888, 11 pages. cited by applicant
      Kobayashi et al., “Identification of a Multipotent Self-Renewing Stromal Progenitor Population during Mammalian Kidney Organogenesis,” Stem Cell Reports, Oct. 2014, vol. 3, pp. 650-662. cited by applicant
      Lam et al., “Rapid and Efficient Differentiation of Human Pluripotent Stem Cells into Intermediate Mesoderm that Forms Tubules Expressing Kidney Proximal Tubular Markers,” J. Am. Soc. Nephrol., Jun. 2014, vol. 25, No. 6, pp. 1211-1225. cited by applicant
      Loughna et al., “Effects of oxygen on vascular patterning in Tie I/Lacz Metanephric kidneys in vitro,” Biochem. Biophys. Res. Commun., Jun. 1998, vol. 247, No. 2, pp. 361-366. cited by applicant
      Love et al., “Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2,” Genome Biol, Dec. 2014, vol. 15, p. 550, 21 pages. cited by applicant
      Mae et al., “Monitoring and robust induction of nephrogenic intermediate mesoderm from human pluripotent stem cells,” Nat. Commun., Jan. 2013, vol. 4, pp. 1-11. cited by applicant
      Mese et al., “The role of caspase family protease, caspase-3 on cisplatin-induced apoptosis in cisplatin 30 resistant A43 I cell line, ” Cancer Chemother. Pharmacol., Sep. 2000, vol. 46, pp. 241-245. cited by applicant
      Mugford et al., “Osrl expression demarcates a multi-potent population of intermediate mesoderm that undergoes progressive restriction to an Osrl-dependent nephron progenitor compartment within the mammalian kidney,” Dev. Biol., Dec. 2008, vol. 324, pp. 88-98. cited by applicant
      Murphy et al., “3D bioprinting of tissues and organs,” Nat. Biotechnol., Aug. 2014, vol. 32, pp. 773-785. cited by applicant
      Naujok et al., “The Generation of Definitive Endoderm from Human Embryonic Stem Cells is Initially Independent from Activin A but Requires Canonical Wnt-Signaling,” Stem Cell Rev. Rep., Jun. 2014, vol. 10, pp. 480-493. cited by applicant
      Orlova et al., “Generation, expansion and functional analysis of endothelial cells and pericytes derived from human pluripotent stem cells,” Nat Protoc., May 2014, vol. 9, No. 6, pp. 1514-1531. cited by applicant
      Park et al., “Six2 and Wnt regulate self-renewal and commitment of nephron progenitors through shared gene regulatory networks,” Dev. Cell, Sep. 2012, vol. 23, pp. 637-651. cited by applicant
      Roost et al., “KeyGenes, a Tool to Probe Tissue Differentiation Using a Human Fetal Transcriptional Atlas,” Stem Cell Reports, Jun. 2015, vol. 4, No. 6, pp. 1112-1124. cited by applicant
      Sakai et al., The retinoic acid-inactivating enzyme CYP26 is essential for establishing an uneven distribution of retinoic acid along the anterior-posterior axis within the mouse embryo, Genes & Development, Jan. 2001, vol. 15, pp. 213-225. cited by applicant
      Short et al., “Global quantification of tissue dynamics in the developing mouse kidney,” Dev. Cell, Apr. 2014, vol. 29, pp. 188-202. cited by applicant
      Sims-Lucas et al., “Endothelial Progenitors Exist within the Kidney and Lung Mesenchyme,” PLoS One, Jun. 2013, vol. 8, No. 6, p. e65993, 8 pages. cited by applicant
      Sweetman et al., “The migration of paraxial and lateral plate mesoderm cells emerging from the late primitive streak is controlled by different Wnt signals,” BMC Dev. Biol., Jun. 2008, vol. 8, pp. 1-15. cited by applicant
      Taguchi, A., et al., “Redefining the In Vivo Origin of Metanephric Nephron Progenitors Enables Generation of Complex Kidney Structures from Pluripotent Stem Cells,” Cell Stem Cell, Jan. 2014, vol. 14, pp. 53-67. cited by applicant
      Takasato et al., ““Directing human embryonic stem cell differentiation towards a renal lineage generates a self- organizing kidney,”” Nature Cell Biology, Dec. 15, 2013, vol. 16, No. 1, pp. 118-127. cited by applicant
      Takasato et al., “Kidney organoids from human iPS cells contain multiple lineages and model human hephrogenesis,” Nature, Oct. 2015, vol. 526, pp. 564-568. cited by applicant
      Takasato et al., “The origin of the mammalian kidney: implications for recreating the kidney in vitro,” Development, Jun. 2015, vol. 142, pp. 1937-1947. cited by applicant
      Thiagarajan et al., “Identification of anchor genes during kidney development defines ontological relationships, molecular subcompartments and regulatory pathways,” PLoS One, Feb. 2011, vol. 6, No. 2, p. e17286, 15 pages. cited by applicant
      Xia et al., “Directed differentiation of human pluripotent cells to ureteric bud kidney progenitor-like cells,” Nat. Cell Biol., Nov. 2013, vol. 15, No. 12, pp. 1507-1515. cited by applicant
      Xinaris et al., “in Vivo Maturation of Functional Renal Organoids Formed from Embryonic Cell Suspensions,” JASN, Nov. 2012, vol. 23, pp. 1857-1868. cited by applicant
      Xu et al., “Eya1 interacts with Six2 and Myc to regulate expansion of the nephron progenitor pool during nephrogenesis, ” Dev. Cell, Nov. 2014, vol. 31, No. 4, pp. 434-447. cited by applicant
      Araoka et al., “Efficient and Rapid Induction of Human iPSCs/ESCs into Nephrogenic Intermediate Mesoderm Using Small Molecule-Based Differentiation Methods,” PLOS One, Jan. 2014, vol. 9, No. 1, pp. e84881, 14 pages. cited by applicant
      Ou et al., “Fibroblast growth factor and organ development”, Journal of Clinical Rehabilitative Tissue Engineering Research, Apr. 2011, vol. 15, No. 15, pp. 2800-2804. cited by applicant
    • Assistant Examiner:
      Montanari, David A
    • Primary Examiner:
      Singh, Anoop K
    • Attorney, Agent or Firm:
      Lum, Leon Y.
    • الرقم المعرف:
      edspgr.12134785