US 11,680,246 B2
Ex vivo proliferation of epithelial cells
Chengkang Zhang, Germantown, MD (US)
Assigned to PROPAGENIX INC., Gaithersburg, MD (US)
Filed by PROPAGENIX INC., Gaithersburg, MD (US)
Filed on Jun. 12, 2020, as Appl. No. 16/899,989.
Application 16/899,989 is a continuation of application No. 15/921,536, filed on Mar. 14, 2018, granted, now 10,711,250.
Application 15/921,536 is a continuation of application No. 15/921,546, filed on Mar. 14, 2018, granted, now 10,711,251.
Application 15/921,536 is a continuation of application No. 15/685,853, filed on Aug. 24, 2017, granted, now 9,963,680, issued on May 8, 2018.
Application 15/921,546 is a continuation of application No. 15/685,853, filed on Aug. 24, 2017, granted, now 9,963,680, issued on May 8, 2018.
Application 15/685,853 is a continuation of application No. 15/296,831, filed on Oct. 18, 2016, granted, now 9,790,471, issued on Oct. 17, 2017.
Application 15/296,831 is a continuation of application No. PCT/US2016/025396, filed on Mar. 31, 2016.
Application PCT/US2016/025396 is a continuation of application No. PCT/US2016/025396, filed on Mar. 31, 2016.
Claims priority of provisional application 62/294,896, filed on Feb. 12, 2016.
Claims priority of provisional application 62/217,406, filed on Sep. 11, 2015.
Claims priority of provisional application 62/142,851, filed on Apr. 3, 2015.
Prior Publication US 2021/0032602 A1, Feb. 4, 2021
Int. Cl. C12N 5/071 (2010.01)
CPC C12N 5/0688 (2013.01) [C12N 5/0629 (2013.01); C12N 5/0683 (2013.01); C12N 2500/90 (2013.01); C12N 2500/99 (2013.01); C12N 2501/11 (2013.01); C12N 2501/113 (2013.01); C12N 2501/117 (2013.01); C12N 2501/15 (2013.01); C12N 2501/727 (2013.01); C12N 2501/999 (2013.01); C12N 2533/54 (2013.01)] 20 Claims
 
1. A method for proliferating epithelial cells ex vivo, comprising:
expanding the number of cells in an originating epithelial cell population under serum-free and feeder-cell free expansion culture conditions, thereby generating an expanded epithelial cell population, wherein (i) the originating epithelial cell population comprises epithelial progenitor cells, wherein the cells have non-detectable expression of one or more genes chosen from LGR5, Axin2, and Prom1 during expansion, and (ii) the expansion culture conditions comprise one or more agents that inhibit transforming growth factor beta (TGF-beta) signaling in the population.
 
15. A method for proliferating epithelial cells ex vivo, comprising:
expanding the number of cells in an originating epithelial cell population under serum-free and feeder-cell free expansion culture conditions, thereby generating an expanded epithelial cell population, wherein (i) the originating epithelial cell population comprises epithelial progenitor cells, wherein the cells have non-detectable expression of one or more genes chosen from LGR5, Axin2, and Prom1 during expansion, and (ii) the expansion culture conditions comprise:
a) one or more transforming growth factor beta (TGF-beta) inhibitors;
b) one or more agents that modulate cytoskeletal structure;
c) a beta-adrenergic receptor agonist;
d) one or more mitogenic growth factors; and
e) calcium at a concentration below 100 μM.
 
18. A composition comprising
a) a cell population comprising epithelial progenitor cells, wherein the cells have non-detectable expression of one or more genes chosen from LGR5, Axin2, and Prom1;
b) one or more transforming growth factor beta (TGF-beta) inhibitors; and
c) one or more agents that modulate cytoskeletal structure.