Ex vivo proliferation of epithelial cells

A method for proliferating epithelial cells ex vivo, comprising:

• expanding the number of cells in an originating epithelial cell population derived from differentiated tissue, embryonic stem (ES) cells, or induced pluripotent stem cells (iPSCs) under expansion culture conditions, thereby generating an expanded epithelial cell population, wherein:
• the expansion culture conditions comprise components consisting essentially of i) a transforming growth factor beta (TGF-beta) signaling inhibitor;
• ii) a Rho-associated protein kinase inhibitor, a p21-activated kinase (PAK) inhibitor, or a myosin II inhibitor;
• iii) a mitogenic growth factor;
• iv) a beta-adrenergic receptor agonist;
• v) calcium at a concentration below 100 μM;
• and vi) a serum-free base medium;
• the expansion culture conditions are serum-free and feeder-cell free culture conditions, and are defined culture conditions, xeno-free culture conditions, or defined and xeno-free culture conditions;
• the originating epithelial cell population is capable of 25 population doublings or more when cultured under the expansion culture conditions;
• and
• the originating epithelial cell population is capable of no more than 20 population doublings when cultured under control culture conditions that do not include the first agent and the second agent.

A population of ex vivo expanded epithelial cells produced by a method comprising:

• expanding the number of cells in an originating epithelial cell population derived from differentiated tissue, embryonic stem (ES) cells, or induced pluripotent stem cells (iPSCs) under expansion culture conditions, thereby generating an expanded epithelial cell population, wherein:
• the expansion culture conditions comprise a first agent consisting of an ALK5 inhibitor and a second agent chosen from a Rho-associated protein kinase inhibitor, a p21-activated kinase (PAK) inhibitor, and a myosin II inhibitor;
• the expansion culture conditions are serum-free and feeder-cell free culture conditions, and are defined culture conditions, xeno-free culture conditions, or defined and xeno-free culture conditions;
• the originating epithelial cell population is capable of 25 population doublings or more when cultured under the expansion culture conditions;
• and
• the originating epithelial cell population is capable of no more than 20 population doublings when cultured under control culture conditions that do not include the first agent and the second agent.

A method for genetically modifying cells, comprising contacting cells with one or more agents that introduce a genetic modification to the cells, wherein the cells have been expanded by a method comprising:

• expanding the number of cells in an originating epithelial cell population derived from differentiated tissue, embryonic stem (ES) cells, or induced pluripotent stem cells (iPSCs) under expansion culture conditions, thereby generating an expanded epithelial cell population, wherein:
• the expansion culture conditions comprise a first agent consisting of an ALK5 inhibitor and a second agent chosen from a Rho-associated protein kinase inhibitor, a p21-activated kinase (PAK) inhibitor, and a myosin II inhibitor;
• the expansion culture conditions are serum-free and feeder-cell free culture conditions, and are defined culture conditions, xeno-free culture conditions, or defined and xeno-free culture conditions;
• the originating epithelial cell population is capable of 25 population doublings or more when cultured under the expansion culture conditions;
• and
• the originating epithelial cell population is capable of no more than 20 population doublings when cultured under control culture conditions that do not include the first agent and the second agent.