Synthetic Cell Platforms and Methods of Use Thereof

• US 20100189760A1
• Filed: 03/21/2008
• Published: 07/29/2010
• Est. Priority Date: 03/22/2007
• Status: Active Grant

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Little, Lauren, Healy, Kevin Edward, Daugherty, Patrick Sean, Schaffer, David V.
• Granted Patent: US 8,501,905 B2
• Time in Patent Office: Days
• US Class Current: 424/423
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 17/00 Carrier-bound or immobilised peptides carrier-bound or immobilised enzymes C12N11/00; Preparation thereof
• CPC Class Codes: C12N 2500/99 Serum-free medium
• CPC Class Codes: C12N 2501/115 Basic fibroblast growth factor (bFGF, FGF-2)
• CPC Class Codes: C12N 2501/15 Transforming growth factor beta (TGF-β)
• CPC Class Codes: C12N 2533/50 Proteins
• CPC Class Codes: C12N 2533/90 Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696
• CPC Class Codes: G01N 33/5073 Stem cells
• CPC Class Codes: G01N 33/542 with steric inhibition or signal modification, e.g. fluorescent quenching

Abstract

The present invention provides synthetic cell platforms. The synthetic cell platforms can be used for culturing cells in vitro. The synthetic cell platforms can also be implanted together with bound cells into an individual. The present invention provides methods of using the platforms to provide cells or progeny of such cells for use in various applications, including clinical applications; and methods of use of the platforms to introduce cells into an individual.

Claims

1. A method of identifying a peptide that binds a cell surface molecule present on a mammalian stem cell surface, the method comprising:

a) contacting a mammalian stem cell with a population of bacterial cells, wherein each bacterial cell in the population displays on its cell surface a different synthetic peptide, said contacting resulting in a mammalian cell having bound to its surface one or more bacteria, and unbound bacteria; and

b) separating the bound bacteria from the unbound bacteria, wherein the synthetic peptide displayed by the bound bacterium is a peptide that binds a cell surface molecule present on a mammalian stem cell surface.

4. A peptide identified by the method of any one of claims 1-3.

5. A synthetic cell platform comprising:

a) a synthetic substrate; and

b) a mammalian stem cell-binding peptide of claim 4 covalently linked to the synthetic substrate.

6. An implantable cell composition comprising:

a) the synthetic platform of claim 5;

b) and a mammalian stem cell bound to the peptide.

7. A method of culturing a mammalian stem cell in vitro, the method comprising, contacting the mammalian stem cell with a synthetic platform according to claim 5, in the absence of a feeder cell layer and in the presence of a serum-free liquid medium.

8. The method of claim 7, wherein the mammalian stem cell is an adult stem cell, an embryonic stem cell, or an induced pluripotent stem cell.

9. The method of claim 7, wherein the synthetic platform promotes self renewal of the stem cell.

10. The method of claim 7, wherein the synthetic platform promotes differentiation of the stem cell.

11. A method of isolating a desired cell type or cell population, the method comprising:

a) contacting a mixed cell population comprising the desired cell type or desired cell population with a synthetic cell platform according to claim 5, forming a complex comprising the desired cell type or desired cell population bound to a peptide on the synthetic cell platform; and

b) separating unbound cells from the complex.

12. A method of enhancing survival of a cell implanted into a recipient individual, the method comprising introducing into the recipient individual an implantable composition of claim 6, wherein the platform enhances survival of the cell in the recipient individual.

13. A method of inducing in vivo differentiation of a stem cell implanted into a recipient individual, the method comprising introducing into the recipient individual an implantable composition of claim 6, wherein the mammalian cell bound to the cell platform is a stem cell, and wherein the platform induces differentiation of the stem cell in vivo.

14. A method of promoting recruitment of a cell to a site in an individual, the method comprising implanting into a site in the individual a synthetic cell platform of claim 5, wherein the covalently linked peptide promotes recruitment of the cell to the site of implantation.

15. An implantable device comprising:

a) a surface; and

b) synthetic cell platform of claim 5 coated onto the surface.

16. The device of claim 15, wherein the device is a stent.

2. A method of identifying a peptide that activates a cell signaling pathway in a mammalian stem cell, the method comprising:

a) contacting a mammalian stem cell with a population of bacterial cells, wherein each bacterial cell in the population displays on its cell surface a different synthetic peptide, wherein said mammalian stem cell is genetically modified to produce a FRET-based detectable signal upon activation of a cell signaling pathway in the mammalian stem cell; and

b) identifying a bacterial cell that, when contacted with the mammalian stem cell, activates the cell signaling pathway, wherein said identifying comprises detecting the FRET-based signal.

3. A method of identifying a peptide that activates a cell signaling pathway in a mammalian stem cell, the method comprising:

a) contacting a mammalian cell with a population of bacterial cells, wherein each bacterial cell in the population displays on its cell surface a different synthetic peptide, wherein said mammalian stem cell is genetically modified with a nucleic acid comprising a nucleotide sequence encoding a kinase sensor comprising a first member of a FRET pair, a phosphoamino acid binding domain, an amino acid sequence that is specifically recognized and phosphorylated by the kinase, and a second member of a FRET pair, and wherein, upon activation of a cell signaling pathway in the mammalian stem cell, the kinase sensor produces a detectable signal; and

b) identifying a bacterial cell that, when contacted with the mammalian stem cell, activates a cell signaling pathway, wherein said identifying comprises detecting a signal from the kinase sensor.