Cardiogenic mesoderm formation regulators

11821,003

16/638918

August 13, 2018

This disclosure relates to cardiogenic mesoderm formation regulators and methods of use thereof, e.g., generating a multipotent cardiovascular progenitor cell by overexpressing Id1, Id2, Id3, Id4, Evx1, and/or Grrp1 in a stem cell.

Sanford Burnham Prebys Medical Discovery Institute (La Jolla, CA, US)

Sanford Burnham Prebys Medical Discovery Institute (La Jolla, CA, US)

1. A method of generating a multipotent cardiovascular progenitor cell, the method comprising: (i) overexpressing one or more proteins selected from the group consisting of Id1 (Inhibitor of DNA binding 1, HLH protein), Id2 (Inhibitor of DNA Binding 2, HLH Protein), Id3 (Inhibitor of DNA Binding 3, HLH Protein), Id4 (Inhibitor of DNA Binding 4, HLH Protein), Evx1 (Even-Skipped Homeobox 1), and Grrp1 (glycine/arginine rich protein 1) in a stem cell, thereby generating a multipotent cardiovascular progenitor cell; or (ii) inhibiting the expression or activity of one or both of Foxa2 (Forkhead Box A2) and Tcf3 (Transcription Factor 3) in a stem cell, thereby generating a multipotent cardiovascular progenitor cell.

2. The method of claim 1 , further comprising transfecting the stem cell with a nucleic acid comprising a sequence encoding one or more proteins selected from the group consisting of Id1, Id2, Id3, Id4, Evx1, and Grrp1.

3. The method of claim 1 , wherein the method further comprises overexpressing Mesp1 (Mesoderm posterior protein 1).

4. The method of claim 1 , wherein the method comprises inhibiting Tcf3.

5. The method of claim 1 , wherein the method comprises inhibiting Foxa2.

6. The method of claim 1 , wherein the method comprises contacting the stem cell with siTcf3.

7. The method of claim 1 , wherein the method comprises contacting the stem cell with siFoxa2.

4522811 June 1985 Eppstein et al.
6168587 January 2001 Bellhouse et al.
6194389 February 2001 Johnston et al.
6468798 October 2002 Debs et al.
6471996 October 2002 Sokoll et al.
6472375 October 2002 Hoon et al.
6849272 February 2005 Langer et al.
8058065 November 2011 Yamanaka et al.
9624471 April 2017 Ruohola-Baker et al.
20070191294 August 2007 Elmen et al.
20080249039 October 2008 Elmen et al.
20090181914 July 2009 Rosenbohm et al.
20100234451 September 2010 Work
20100249052 September 2010 Benson et al.
20100317718 December 2010 Marcusson et al.
20100330044 December 2010 Blanpain et al.
20110318775 December 2011 Mercola et al.
20130130387 May 2013 Itskovitz-Eldor et al.
20140093486 April 2014 Chiou et al.
20150099690 April 2015 Pellois
WO 2010/040112 April 2010
WO 2010/094757 August 2010
WO 2010/129746 November 2010
WO 2011/044253 April 2011

Cunningham (Genes Dev, 31(3): 1325-1338, 2017). (Year: 2017). cited by examiner
Altschul et al., “Basic Local Alignment Search Tool,” J. Mol. Biol., 1990, 215(3): 403-410. cited by applicant
Benezra et al., “The protein Id: a negative regulator of helix-loop-helix DNA binding proteins,” Cell, 1990, 61(1): 49-59. cited by applicant
Beppu et al., “BMP type II receptor is required for gastrulation and early development of mouse embryos,” Developmental Biology, 2000, 221(1):249-258. cited by applicant
Birket et al., “Expansion and patterning of cardiovascular progenitors derived from human pluripotent stem cells,” Nature Biotechnology, 2015, 33(9):970-979. cited by applicant
Blin et al., “A Purified Population of Multipotent Cardiovascular Progenitors Derived From Primate Pluripotent Stem Cells Engrafts in Postmyocardial Infarcted Nonhuman Primates,” J Clin Invest, 2010, 120(4):1125-1139. cited by applicant
Bondue et al., “Mespl acts as a master regulator of multipotent cardiovascular progenitor specification,” Cell Stem Cell, 2008, 3(1):69-84. cited by applicant
Bruneau, “Signaling and transcriptional networks m heart development and regeneration,” Cold Spring Harb Perspect Biol, 2013, 5(3):a008292. cited by applicant
Buckingham et al., “Building the mammalian heart from hvo sources of myocardial cells,” Nature Reviews Genetics, 2005, 6(11):826-835. cited by applicant
Burridge et al., “Chemically defined generation of human cardiomyocytes,” Nature Methods,2014, 11(8):855-860. cited by applicant
Cai et al., “ISL1 identifies a cardiac progenitor population that proliferates prior to differentiation and contributes a majority of cells to the heart,” Developmental Cell, 2003, 5(6):877-889. cited by applicant
Chan et al., “Mespl patterns mesoderm into cardiac, hematopoietic, or skeletal myogenic progenitors in a context-dependent manner,” Cell Stem Cell, 2012, 12:587-601. cited by applicant
Chiapparo et al., “Mespl controls the speed, polarity, and directionality of cardiovascular progenitor migration,” J Cell Biol, 2016, 213(4):463-477. cited by applicant
Christoforou et al., “Implantation of Mouse Embryonic Stem Cell-Derived Cardiac Progenitor Cells Preserves Function of Infarcted Murine Hearts,” PLoS One, 2010, 5(7):e11536, 14 pages. cited by applicant
Colas et al., “Mix. 1/2-dependent control of FGF availability during gastrulation is essential for pronephros development in Xenopus,” Developmental Biology, 2008, 320(2):351-365. cited by applicant
Colas et al., “Whole-genome microRNA screening identifies let-7 and mir-18 as regulators of germ layer formation during early embryogenesis,” Genes & Development, 2012, 26(23):2567-2579. cited by applicant
Collop et al., “Retinoic acid signaling is essential for formation of the heart tube in Xenopus,” Developmental Biology, 2006, 291(1):96-109. cited by applicant
Costello et al., “The T-box transcription factor Eomesodermin acts upstream of Mespl to specify cardiac mesoderm during mouse gastrulation,” Nature Cell Biology, 2011, 13(9):1084-1091. cited by applicant
Czarnik, “Encoding Methods for Combinatorial Chemistry,” Curr. Opin. Chem Biol, 1997, 1(1):60-6. cited by applicant
David et al., “MesPl drives vertebrate cardiovascular differentiation through Dkk-1-mediated blockade of Wnt-signalling,” Nature Cell Biology, 2008, 10:338-345. cited by applicant
Devine et al., “Early patterning and specification of cardiac progenitors in gastrulating mesoderm,” Elife, 2014, 3:e03848, 23 pages. cited by applicant
Djiane et al., “Role of frizzled 7 in the regulation of convergent extension movements during gastrulation in Xenopus laevis,” Development, 2000, 127(14):3091-3100. cited by applicant
Ekins et al., “Microarrays: Their Origins and Applications,” Trends in Biotechnology, 1999, 17(6):217-218. cited by applicant
Ema et al., “Deletion of the selection cassette, but not cis-acting elements, in targeted Flkl-lacZ allele reveals Flkl expression in multipotent mesodermal progenitors,” Blood, 2006, 107(1):111-117. cited by applicant
Foley et al., “Multiple functions of Cerberus cooperate to induce heart downstream of Nodal,” Developmental Biology, 2007, 303(1):57-65. cited by applicant
Fraidenraich et al., “Rescue of cardiac defects in id knockout embryos by injection of embryonic stem cells,” Science, 2004, 306(5694):247-252. cited by applicant
Gadue et al., “Wnt and TGF-beta signaling are required for the induction of an in vitro model of primitive streak formation using embryonic stem cells,” Proc Natl Acad Sci US A, 2006, 103(45):16806-16811. cited by applicant
Galvin et al., “Nodal signaling regulates the bone morphogenic protein pluripotency pathway in mouse embryonic stem cells,” The Journal of Biological Chemistry, 2010, 285(26):19747-19756. cited by applicant
Hamajima et al., “Intranasal Administration of HIV-DNA Vaccine Formulated With a Polymer, Carboxymethylcellulose, Augments Mucosal Antibody Production and Cell-Mediated Immune Response,” Clin. Immunol. Immunopathol., 1998, 88(2):205-10. cited by applicant
Hollnagel et al., “Id genes are direct targets of bone morphogenetic protein induction in embryonic stem cells,” The Journal of Biological Chemistry, 1999, 274(28):19838-19845. cited by applicant
Huang et al., “Induction of functional hepatocyte-like cells from mouse fibroblasts by defined factors,” Nature, 2011, 475(7356):386-389. cited by applicant
Katagiri et al., “Identification of a BMP-responsive element in Idl, the gene for inhibition of myogenesis,” Gene Cells, 2002, 7(9):949-960. cited by applicant
Kattman et al., “Multipotent flk-1 + cardiovascular progenitor cells give rise to the cardiomyocyte, endothelial, and vascular smooth muscle lineages,” Developmental Cell, 2006, 11(5):723-732. cited by applicant
Kee et al., “E and ID proteins branch out,” Nature Reviews Immunology, 2009, 9(3):175-184. cited by applicant
Kee et al., “To proliferate or to die: role of Id3 in cell cycle progression and survival of neural crest progenitors,” Genes & Development, 2005, 19(6):744-755. cited by applicant
Kelly et al, “Heart fields and cardiac morphogenesis,” Cold Spring Harb Perspect Med, 2014, 4(10):a015750, 10 pages. cited by applicant
Kelly et al., “The arterial pole of the mouse heart forms from Fgf10-expressing cells in pharyngeal mesoderm,” Developmental Cell, 2001, 1(3):435-440. cited by applicant
Korchynskyi et al., “Identification and functional characterization of distinct critically important bone morphogenetic protein-specific response elements in the Idl promoter,” The Journal of Biological Chemistry, 2002, 277(7):4883-4891. cited by applicant
Laflamme et al., “Cardiomyocytes derived from human embryonic stem cells in pro-survival factors enhance function of infarcted rat hearts,” Nature Biotechnology, 2007, 25(9):1015-1024. cited by applicant
Lescroart et al., “Early lineage restriction in temporally distinct populations of Mesp 1 progenitors during mammalian heart development,” Nature Cell Biology, 2014, 16:829-840. cited by applicant
Lian et al., “Directed cardiomyocyte differentiation from human pluripotent stem cells by modulating Wnt/beta-catenin signaling under fully defined conditions,” Nature Protocols, 2013, 8(1):162-175. cited by applicant
Liang et al., “Drug screening using a library of human induced pluripotent stem cell-derived cardiomyocytes reveals disease specific patterns of cardiotoxicity,” Circulation, 2013, 127:1677-91. cited by applicant
Lopez-Rovira et al., “Direct binding of Smadl and Smad4 to two distinct motifs mediates bone morphogenetic protein-specific transcriptional activation of Idl gene,” The Journal of Biological Chemistry, 2002, 277(5):3176-3185. cited by applicant
Lyden et al., “Idl and Id3 are required for neurogenesis, angiogenesis and vascularization of tumour xenografts,” Nature, 1999, 401(6754):670-677. cited by applicant
MacBeath et al., “Printing Proteins as Microarrays for High-Throughput Function Determination,” Science, 2000, 289(5485):1760-1763. cited by applicant
Marvin et al., “Inhibition of Wnt activity induces heart formation from posterior mesoderm,” Genes & Development, 2001, 15(3):316-327. cited by applicant
McKeithan et al., “An Automated Platform for Assessment of Congenital and Drug-Induced Arrhythmia with hiPSC-Derived Cardiomyocytes,” Frontiers in Physiology, 2017, 8:766, 12 pages. cited by applicant
McKeithan et al., “Serum-free generation of multipotent mesoderm (Kdr+) progenitor cells in mouse embryonic stem cells for functional genomics screening,” Current Protocols in Stem Cell Biology, 2012, Chapter 1, Unit 1F 13, 17 pages. cited by applicant
Meilhac et al., “Cardiac cell lineages that form the heart,” Cold Spring Harb Perspect Med, 2015, 5(2):a026344, 14 pages. cited by applicant
Meilhac et al., “The clonal origin of myocardial cells in different regions of the embryonic mouse heart,” Developmental Cell, 2004, 6(5):685-698. cited by applicant
Menard et al., “Transplantation of Cardiac-Committed Mouse Embryonic Stem Cells to Infarcted Sheep Myocardium: A Preclinical Study,” Lancet, 2005, 366(9490):1005-1012. cited by applicant
Menasche et al., “Towards a clinical use of human embryonic stem cell-derived cardiac progenitors: a translational experience,” Eur Heart J, 2015, 36(12):743-750. cited by applicant
Mercola et al., “Induced pluripotent stem cells in cardiovascular drug discovery,” Circulation Research, 2013, 112(3):534-548. cited by applicant
Miller et al., “Optically monitoring voltage in neurons by photo-induced electron transfer through molecular wires,” Proceedings of the National Academy of Sciences of the United States of America, 2012, 109(6):2114-2119. cited by applicant
Moretti et al., “Pluripotent stem cell models of human heart disease,” Cold Spring Harb Perspect Med, 2013, 3(11):a014027, 20 pages. cited by applicant
Nieuwkoop et al., “The “organization centre”. 3. Segregation and pattern formation in morphogenetic fields,” Acta Biotheor, 1967, 17(4):178-194. cited by applicant
Niola et al., “Id proteins synchronize sternness and anchorage to the niche of neural stem cells,” Nature Cell Biology, 2012, 14(5):477-487. cited by applicant
Niola et al., “Mesenchymal high-grade glioma is maintained by the ID-RAPI axis,” The Journal of Clinical Investigation, 2013, 123(1):405-417. cited by applicant
Olson, “Gene regulatory networks in the evolution and development of the heart,” Science, 2006, 313(5795):1922-1927. cited by applicant
Paige et al., “A temporal 5 chromatin signature in human embryonic stem cells identifies regulators of cardiac development,” Cell, 2012, 151(1):221-232. cited by applicant
Pandur et al., “Wnt-11 activation of a non-canonical Wnt signalling pathway is required for cardiogenesis,” Nature, 2002, 418(6898):636-641. cited by applicant
PCT International Search Report and Written Opinion in International Appln. No. PCT/US2018/046536, dated Nov. 12, 2018, 1 pages. cited by applicant
Peng, “Xenopus laevis: Practical uses in cell and molecular biology. Solutions and Protocols,” Methods Cell Biol, 1991, 36:657-662. cited by applicant
Raffin et al., “Subdivision of the cardiac Nkx2.5 expression domain into myogenic and nonmyogenic compartments,” Developmental Biology, 2000, 218(2):326-340. cited by applicant
Roschger et al., “The Id-protein family in developmental and cancer-associated pathways,” Cell Commun Signal, 2017, 15(1):7, 26 pages. cited by applicant
Saga et al., “Mespl expression is the earliest sign of cardiovascular development,” Trends in Cardiovascular Medicine, 2000, 10(8):345-352. cited by applicant
Saga et al., “MesPl: a novel basic helix-loop-helix protein expressed in the nascent mesodermal cells during mouse gastrulation,” Development, 1996, 122:2769-2778. cited by applicant
Schneider et al., “Wnt antagonism initiates cardiogenesis in Xenopus laevis,” Genes & Development, 2001, 15(3):304-315. cited by applicant
Schultheiss et al., “A role for bone morphogenetic proteins in the induction of cardiac myogenesis,” Genes & Development, 1997, 11(4):451-462. cited by applicant
Sekiya et al., “Direct conversion of mouse fibroblasts to hepatocyte-like cells by defined factors,” Nature, 2011, 475(7356):390-393. cited by applicant
Stainier, “A glimpse into the molecular entrails of endoderm formation,” Genes & Development, 2002, 16(8):893-907. cited by applicant
Tomescot et al., “Differentiation in Vivo of Cardiac Committed Human Embryonic Stem Cells in Postmyocardial Infarcted Rats,” Stem Cells, 2007, 25(9):2200-2205. cited by applicant
Viotti et al., “SOX17 Links Gut Endoderm Morphogenesis and Germ Layer Segregation,” Nature Cell Biology, 2014, 16(12):1146-1156. cited by applicant
Wilkinson et al., “Detection of messenger RNA by in situ hybridization to tissue sections and whole mounts,” Methods in Enzymology, 1993, 225:361-373. cited by applicant
Yang et al., “Human cardiovascular progenitor cells develop from a KDR+ embryonic-stem-cell-derived population,” Nature, 2008, 453(7194):524-528. cited by applicant
Yang et al., “Id proteins in the vasculature: from molecular biology to cardiopulmonary medicine,” Cardiovascular Research, 2014, 104(3):388-398. cited by applicant
Yoon et al., “HEB and E2A function as SMAD/FOXH1 cofactors,” Genes Dev, 2011, 25(15):1654-1661. cited by applicant
Yoshida et al., “Cell lineage in mammalian craniofacial mesenchyme,” Mech Dev, 2008, 125(9-10):797-808. cited by applicant
Yu et al., “Generation of First Heart Field-like Cardiac Progenitors and Ventricular-like Cardiomyocytes from Human Pluripotent Stem Cells,” J Vis Exp., 2018, 136:57688, 9 pages. cited by applicant
Zhang et al., “PowerBLAST: A New Network BLAST Application for Interactive or Automated Sequence Analysis and Annotation,” Genome Res., 1997, 7(6):649-656. cited by applicant
Zhao et al., “Loss of both GAT A4 and GAT A6 blocks cardiac myocyte differentiation and results in acardiain mice,” Developmental Biology, 2008, 317(2):614-619. cited by applicant
Thomas Cunningham et al., “Id genes are essential for early heart formation”, Genes and Development, vol. 31, No. 13, Jul. 1, 2017, pp. 1325-1338. cited by applicant
International Application No. PCT/US2018/046536, International Preliminary Report on Patentability and Written Opinion of the International Searching Authority, dated Feb. 27, 2020, 9 pages. cited by applicant

edspgr.11821003