# US 2014/0154802 A1 · Compositions and Methods Useful for Culturing Differentiable Cells

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- Number: US 2014/0154802 A1
- Document ID: US20140154802A1
- Status: Published application
- Kind: A1
- Application number: 14/176,894
- Filed: 2014-02-10
- Published: 2014-06-05
- Earliest claimed: 2006-02-23
- Assignees: ViaCyte, Inc.
- Applicants: Viacyte, Inc.
- Inventors: Allan J. Robins; Thomas C. Schulz
- Web page: https://patentgrep.com/patent/US20140154802A1

## CPC

- C12N 5/0606: {Pluripotent embryonic cells, e.g. embryonic stem cells [ES] (embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696)}
- G01N 33/5008: {for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics}
- G01N 33/5073: {Stem cells}
- C12N 2500/90: Serum-free medium, which may still contain naturally-sourced components
- C12N 2501/105: Insulin-like growth factors [IGF]
- C12N 2501/115: Basic fibroblast growth factor (bFGF, FGF-2)
- C12N 2501/155: Bone morphogenic proteins [BMP]; Osteogenins; Osteogenic factor; Bone inducing factor
- C12N 2501/16: Activin; Inhibin; Mullerian inhibiting substance
- C12N 2501/195: Heregulin, neu differentiation factor
- C12N 2501/235: Leukemia inhibitory factor [LIF]
- C12N 2502/13: connective tissue cells; generic mesenchyme cells, e.g. so-called "embryonic fibroblasts"
- C12N 2509/00: Methods for the dissociation of cells, e.g. specific use of enzymes
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

The present invention relates to cell culture methods and compositions that are essentially serum-free and comprise a basal salt nutrient solution and an ErbB3 ligand.

## Claims

### Claim at position 1

1-34. (canceled)

### Claim 35

Pluripotent cells in an in vitro suspension culture.

### Claim 36 (depends on 35)

The pluripotent cells of claim 35, wherein the pluripotent cells are substantially undifferentiated.

### Claim 37 (depends on 35)

The pluripotent cells of claim 35, wherein said pluripotent cells are human embryonic stem cells.

### Claim 38 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture comprises an ErbB3 ligand.

### Claim 39 (depends on 35)

The pluripotent cells of claim 35, wherein said ErbB3 ligand is selected from the group consisting of Neuregulin-1, Heregulin-β (HRG-β), Heregulin-α (HRG-α), Neu differentiation factor (NDF), acetylcholine receptor-inducing activity (ARIA), glial growth factor 2 (GGF2), motor-neuron derived factor (SMDF), Neuregulin-2, Neuregulin-2β (NRG2-β), Epiregulin, Biregulin and functional fragments thereof.

### Claim 40 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture comprises a TGF-β superfamily member.

### Claim 41 (depends on 35)

The pluripotent cells of claim 35, wherein said TGF-β family member is selected from the group consisting of Nodal, Activin A, Activin B, bone morphogenic protein-2 (BMP2), bone morphogenic protein-4 (BMP4) and functional fragments thereof.

### Claim 42 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture comprises an insulin-like growth factor.

### Claim 43 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture comprises a fibroblast growth factor.

### Claim 44 (depends on 35)

The pluripotent cells of claim 35, wherein said fibroblast growth factor is selected from the group consisting of FGF-2, FGF-7, FGF-10, FGF-22 and variants functional fragments thereof.

### Claim 45 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture comprises a ROCK inhibitor.

### Claim 46 (depends on 35)

The pluripotent cells of claim 35, wherein said Rock inhibitor is Y27632.

### Claim 47 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture does not comprise a TGF-β superfamily member.

### Claim 48 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture does not comprise a fibroblast growth factor.

### Claim 49 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture is serum free.

### Claim 50 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture is matrix free.

### Claim 51 (depends on 35)

The pluripotent cells of claim 35, wherein the cells are maintained in suspension for at least two days.

### Claim 52 (depends on 35)

The pluripotent cells of claim 35, wherein the suspension culture comprises an erbB3 ligand and is serum free.

### Claim 53

An in vitro population of pluripotent cells in suspension culture.

### Claim 54

An in vitro suspension culture comprising pluripotent cells.