US 12,378,341 B2
Tuneable cell substrates
Chibeza Chintu Agley, Cambridgeshire (GB); Kevin Chalut, Cambridgeshire (GB); and Jose Carlos Rebelo da Silva, Cambridgeshire (GB)
Assigned to Cambridge Enterprise Limited, Cambridgeshire (GB)
Appl. No. 16/772,584
Filed by Cambridge Enterprise Limited, Cambridgeshire (GB)
PCT Filed Dec. 14, 2018, PCT No. PCT/EP2018/085079
§ 371(c)(1), (2) Date Jun. 12, 2020,
PCT Pub. No. WO2019/115814, PCT Pub. Date Jun. 20, 2019.
Claims priority of application No. 1721046 (GB), filed on Dec. 15, 2017.
Prior Publication US 2020/0399410 A1, Dec. 24, 2020
Int. Cl. C08F 220/56 (2006.01); B82Y 5/00 (2011.01); C08F 222/38 (2006.01); C12M 3/02 (2006.01); C12N 5/00 (2006.01); C12N 5/074 (2010.01); C12N 5/077 (2010.01)
CPC C08F 220/56 (2013.01) [C08F 222/385 (2013.01); C12M 3/02 (2013.01); C12N 5/0068 (2013.01); C12N 5/0607 (2013.01); C12N 5/0654 (2013.01); B82Y 5/00 (2013.01); C12N 2501/155 (2013.01); C12N 2506/45 (2013.01); C12N 2533/30 (2013.01)] 18 Claims
 
1. A substrate for culture of mammalian stem and progenitor cells culture comprising;
(i) a co-polymer hydrogel formed from a co-polymer, wherein the co-polymer comprises cross-linked chains of monomeric units of acrylamide, a N,N′ methylenebisacrylamide (bisacrylamide) cross-linking agent, and a coupling compound in the hydrogel; and
(ii) a cell adhesion molecule, said cell adhesion molecule being covalently coupled to coupling compound monomeric units of the co-polymer,
wherein the coupling compound is 6-acrylamidohexanoic acid (6AHA) and
the co-polymer hydrogel comprises 1-10% (w/v in g/100 ml) acrylamide monomeric units and 10 to 100 mM 6AHA,
wherein the substrate constitutes a stable tunable synthetic stem cell niche for maintenance of stable mammalian stem and progenitor cell cultures;
wherein concentration of the monomeric units of acrylamide in the co-polymer hydrogel controls stiffness of the substrate; and
wherein stiffness of the hydrogel corresponds to stiffness of a biological tissue;
wherein the cell adhesion molecule forms a molecular anchoring site for the cultured mammalian stem or progenitor cells;
wherein concentration of the coupling compound in the copolymer hydrogel controls cell adhesion molecule density of the substrate; and
wherein density of the cell adhesion molecule reflects density of the cell adhesion molecule in a biological tissue;
wherein stiffness of the substrate and density of the cell adhesion molecules are independently tunable;
wherein the substrate with a defined stiffness enables selection of cells from specific tissues; and
wherein the substrate with a high density of cell adhesion molecules enables selection of mammalian stem or progenitor cells that originate from a high adhesion environment; and the substrate with a low density of cell adhesion molecules enables selection of mammalian stem or progenitor cells that originate from a low adhesion environment.