Patent ID: 9745551
Date: 2017-08-29
CPC Classifications: A61K,C12N

Claim:
1. A method for producing human embryonic stem cell-derived mesenchymal stem cells (hES-MSCs) comprising: a) culturing human embryonic stem cells in a serum free medium comprising at least one GSK3 inhibitor at a concentration ranging from 0.05 μM to 0.2 μM, wherein the human embryonic stem cells are cultured in the absence of feeder cells; b) culturing the cells resulting from step a) in a serum-free medium comprising vascular endothelial growth factor (VEGF) and bone morphogenic protein 4 (BMP4) in an amount sufficient to induce formation of embryoid bodies comprising human hemangio-colony forming cells; c) adding at least one growth factor to the culture resulting from step b), wherein the growth factor is in an amount sufficient to expand human hemangio-colony forming cells; d) disaggregating the hemangio-colony forming cells resulting from step c) into single cells; and e) culturing the single hemangio-colony forming cells resulting from step d) in mesenchymal stem cell medium containing serum, knockout serum replacement (KOSR) or in a serum-free medium to induce differentiation of the single cells into human mesenchymal stem cells; wherein at least 90% of the hES-MSCs express CD73; and said hES-MSCs: (i) comprise greater than 95% of cells expressing CD73, CD90, CD105, CD146, CD166, and CD44; (ii) comprise greater than 80% of cells expressing CD13, CD29, CD54, and CD49E; (iii) comprise less than 5% of cells expressing CD45, CD34, CD31 and SSEA4; (iv) express IL-10 and TGFβ; (v) comprise less than 2% of cells expressing IL-6, IL-12 and TNFα; and (vi) comprise less than 0.001% of cells co-expressing OCT4, NANOG, TRA-1-60 and SSEA4.