Chemical Genetic Approach to Production of hESC-derived Cardiomyocytes
Chemical Genetic Approach to Production of hESC-derived Cardiomyocytes
Stem Cell Use:
Embryonic Stem Cell
Statement of Benefit to California:
Year 1The goal of this project is to identify small molecules that stimulate cardiomyocyte differentiation from stem cells. The strategy is to use embryonic stem ESC)-derived progenitors to screen for compounds and then optimize their chemical properties to generate molecules that can be used as reagents and potentially as lead compounds to develop drugs to stimulate regeneration in patient hearts. During year 2, progress is reported in: 1) optimizing the biological and pharmaceutical properties of 4 chemically diverse compounds discovered in year 1; 2) patent application filed on these compounds; 3) identification of targets and biological mechanism of action of 2 of the 4 compounds; 4) 1 compound has been validated in hESCs; 5) pilot screening completed of a new stem cell screen to discover molecules that act on late stage progenitors similar to cells thought to exist in the adult heart; 5) new assays developed and screened for discovering modulators of the Wnt pathway that enhance cardiomyocyte production. Thus, there are a total of 8 chemically distinct compounds under study and additional assays have been developed that should bring additional compounds into the pipeline during year 3.
Year 2This progress report covers FY3 of the project to identify and characterize novel small molecule probes of cardiomyocyte differentiation from stem cells. During FY3, we characterized 11 novel chemical entities that promote cardiomyocyte differentiation. The small, drug-like molecules affect distinct steps in cardiomyocyte differentiation – 5 compounds promote formation of uncommitted cardiac progenitors, 2 stimulated committed cardiac precursors, while 2 compounds act later to stimulate differentiation into cardiomyocytes. Thus, these compounds are novel probes of stem cell differentiation. Some of the compounds are characterized to act upon particular cellular target proteins while the targets of other compounds are unknown. Of the latter class, candidate targets have been characterized by biochemical studies; one of which has been confirmed by RNA interference, yielding a new pathway in cardiac cell formation from stem cells. Three of the chemical series have been described in a patent application. Additional primary hits are being characterized. For FY4, we will continue characterization of a novel compounds. Particular focus will be on 4 chemical entities that promote later stages of human stem cell cardiomyocyte differentiation and on characterizing and discovering additional candidates that act on late-stage differentiation. In addition, we will develop a new pathway screen for a cellular target involved in specifying cardiomyocyte progenitors that have recently been shown to form new myocytes in vivo. Our new compounds are valuable probes of the underlying mechanism(s) responsible for making cardiac cells from stem cells. Moreover, recent data has shown that endogenous stem cells that reside in the adult heart resemble progenitors in the hESC cultures, so certain of our compounds can be considered as targeting cellular proteins and signaling pathways that might be beneficial to stimulate endogenous regeneration. Towards this goal, we will optimize the drug-like properties of the compounds in anticipation of in vivo testing for regenerative potential.
Year 3This research led to the discovery of small molecules that promote the formation of heart muscle cells from human pluripotent stem cells. The project used high throughput screening technology and medicinal chemistry, similar to that used in pharmaceutical companies, to discover and optimize the molecules. The cellular processes targeted by the compounds were also investigated, and in several cases this research uncovered novel roles for key cellular proteins and signaling pathways, such as Wnt and TGFb signaling, in stem cell differentiation. The compounds will be useful as reagents for cardiomyocyte preparation from stem cells, and patent applications have been filed.
- J Med Chem (2012) Synthesis and SAR of b-Annulated 1,4-Dihydropyridines Define Cardiomyogenic Compounds as Novel Inhibitors of TGFbeta Signaling. (PubMed: 23130626)
- Genes Dev (2012) Whole-genome microRNA screening identifies let-7 and mir-18 as regulators of germ layer formation during early embryogenesis. (PubMed: 23152446)
- Curr Protoc Stem Cell Biol (2012) Serum-free generation of multipotent mesoderm (kdr(+)) progenitor cells in mouse embryonic stem cells for functional genomics screening. (PubMed: 23154934)
- Cell Stem Cell (2012) Small Molecule-Mediated TGF-beta Type II Receptor Degradation Promotes Cardiomyogenesis in Embryonic Stem Cells. (PubMed: 22862949)
- J Med Chem (2012) Wnt Inhibition Correlates with Human Embryonic Stem Cell Cardiomyogenesis: A Structure-Activity Relationship Study Based on Inhibitors for the Wnt Response. (PubMed: 22191557)
- J Med Chem (2012) Wnt Inhibition Correlates with Human Embryonic Stem Cell Cardiomyogenesis: A Structure-Activity Relationship Study Based on Inhibitors for the Wnt Response.
- J Cardiovasc Transl Res (2011) A chemical biology approach to myocardial regeneration. (PubMed: 21424858)
- Mol Cell (2011) Fine-tuning of Drp1/Fis1 availability by AKAP121/Siah2 regulates mitochondrial adaptation to hypoxia. (PubMed: 22099302)
- Circ Res (2011) Small-Molecule Inhibitors of the Wnt Pathway Potently Promote Cardiomyocytes From Human Embryonic Stem Cell-Derived Mesoderm. (PubMed: 21737789)
- Mol Cell (2011) Fine-tuning of Drp1/Fis1 availability by AKAP121/Siah2 regulates mitochondrial adaptation to hypoxia.
- Ann N Y Acad Sci (2010) Cardiac myocyte force development during differentiation and maturation. (PubMed: 20201894)
- Proc Natl Acad Sci U S A (2010) Alternative splicing regulates mouse embryonic stem cell pluripotency and differentiation. (PubMed: 20498046)
- Stem Cells Dev (2010) Non-cardiomyocytes influence the electrophysiological maturation of human embryonic stem cell-derived cardiomyocytes during differentiation. (PubMed: 20001453)
- Circulation (2009) Electrophysiological challenges of cell-based myocardial repair. (PubMed: 20008740)
- Pediatr Cardiol (2009) Natural and synthetic regulators of embryonic stem cell cardiogenesis. (PubMed: 19319460)
- PLoS One (2009) Lentiviral vectors and protocols for creation of stable hESC lines for fluorescent tracking and drug resistance selection of cardiomyocytes. (PubMed: 19352491)
- PLoS Comput Biol (2009) Alternative splicing in the differentiation of human embryonic stem cells into cardiac precursors. (PubMed: 19893621)
- PLoS One (2008) Contrasting expression of keratins in mouse and human embryonic stem cells. (PubMed: 18941637)
- Dev Biol (2008) A novel activity of the Dickkopf-1 amino terminal domain promotes axial and heart development independently of canonical Wnt inhibition. (PubMed: 18840423)
- Assay Drug Dev Technol (2008) A comparative analysis of standard microtiter plate reading versus imaging in cellular assays. (PubMed: 18795873)