IP Library › Granted Patent US 12,606,798
Granted Patent B2
US 12,606,798 · App. 17/715,889 · Granted Apr 21, 2026

Methods and materials for hematoendothelial differentiation of human pluripotent stem cells under defined conditions

Inventors: Igor Slukvin (Verona, WI); Gene Ichiro Uenishi (Madison, WI)
Assignee: Wisconsin Alumni Research Foundation
C12N5/0647C12N5/0056C12N5/0636C12N5/069C12N5/0695C12N2500/02C12N2501/115C12N2501/155C12N2501/16C12N2501/165C12N2502/1394C12N2506/02C12N2506/45C12N2533/54
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Quick Facts
Patent No.
US 12,606,798
App. No.
17/715,889
Granted
Apr 21, 2026
Kind
B2
Abstract

Methods, kits and compositions for differentiating pluripotent stem cells into cells of endothelial and hematopoietic lineages are disclosed.

Claims (16)

1 . A xenogen-free, albumin-free culture system capable of differentiating human pluripotent stem cells into mesoderm, endothelial and hematopoietic progenitor cells comprising:

a) a solid substrate comprising a layer of Tenascin C or collagen; and

b) xenogen-free and albumin-free culture medium comprising:

about 50 to about 250 ng/ml BMP4;

about 10 to about 15 ng/ml Activin A;

about 10 to about 50 ng/ml FGF2; and

about 1 to about 2 mM LiCl,

wherein the culture system, when seeded with human pluripotent stem cells, produces EMH lin − KDR + APLNR + PDGFRalpha + primitive mesoderm cells.

2 . The xenogen-free, albumin-free culture system of claim 1 , wherein the medium further comprises a base medium, L-ascorbic acid 2-phopshate Mg 2+ salt monothioglycerol, sodium selenite, polyvinyl alcohol, non-essential amino acids (NEAA), chemically defined lipid concentrate, holo-transferrin, and insulin.

3 . The xenogen-free, albumin-free culture system of claim 2 , wherein the medium comprises 64 mg/L L-ascorbic acid 2-Phosphate Mg 2+ salt, 40 μl/L monothioglycerol, 8.4 μg/L additional sodium selenite, 10 mg/L polyvinyl alcohol, 1× non-essential amino acids, 0.1×chemically-defined lipid concentrate, 10.6 mg/L holo-transferrin, and 20 mg/L insulin.

4 . The xenogen-free culture system of claim 1 , wherein the medium of step b further comprises: about 50 to about 100 ng/ml stem cell factor (SCF), about 50 to about 100 ng/ml thrombopoietin (TPO), about 50 to about 100 ng/ml IL-6, and about 5 to about 15 ng/ml IL-3.

5 . The xenogen-free, albumin-free culture system of claim 1 , wherein the human pluripotent stem cells can be seeded at a concentration of about 0.25 μg/cm 2 to about 1 μg/cm 2 .

6 . The xenogen-free, albumin-free culture system of claim 1 , wherein the solid substrate comprises a layer of Tenascin C.

7 . The xenogen-free, albumin-free culture system of claim 6 , wherein the layer of Tenascin C is at a concentration of about 0.25 μg/cm 2 to about 1 μg/cm 2 .

8 . The xenogen-free culture system of claim 1 , wherein the solid substrate of (a) further comprises human pluripotent stem cells seeded thereupon.

9 . The xenogen-free, albumin-free culture system of claim 1 , wherein the solid substrate comprises a layer of collagen.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Apr 19, 2022
From: SLUKVIN, IGOR; UENISHI, GENE
To: WISCONSIN ALUMNI RESEARCH FOUNDATION
Reel/Frame 059634/0351 →
Continuity (4)
Continuation 15917504 · Mar 9, 2018
Continuation 14206778 · Mar 12, 2014
Provisional Application 61779564 · Mar 13, 2013
Related Publication 20220228118A1 · Jul 21, 2022
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