Human Extensively Self-Renewing Erythroblasts (ESRE)

Abstract

The present invention provides a human cell population that can self-renew extensively and yet retain the capacity to differentiate into red blood cells (RBCs). These cells are referred to as extensively self-renewing erythroblasts (ESREs). The cells of the invention serve among other things as a renewable source of transfusable RBCs.

1 - 5 . (canceled) 
     
     
         6 . A composition comprising:
 an isolated population of human ESRE and   an expansion medium comprising Epo, SCF, dexamethasone, and a lipid mixture.   
     
     
         7 . The composition of  claim 6 , wherein the expansion medium further comprises one or more of a Bmi-1 protein, a Bmi-1 peptide, and a Bmi-1 regulating agent. 
     
     
         8 - 17 . (canceled) 
     
     
         18 . A method of delivering a gene to a mammal, the method comprising administering a gene delivery vehicle to the mammal wherein the vehicle comprises an extensively self-renewing erythroblast (ESRE) that is genetically modified to express one or more therapeutic proteins. 
     
     
         19 . A method of generating a human red blood cell (RBC), the method comprising culturing an extensively self-renewing erythroblast (ESRE) in a differentiation medium comprising EPO and insulin. 
     
     
         20 - 33 . (canceled) 
     
     
         34 . The composition of  claim 6 , wherein the population of ESRE proliferates in culture and maintains the ability to terminally mature, wherein the ESRE is not immortalized. 
     
     
         35 . The composition of  claim 6 , wherein the population of ESRE is capable of undergoing at least 30 cell divisions. 
     
     
         36 . The composition of  claim 6 , wherein the population of ESRE is derived from a population of definitive erythroid progenitors further wherein the population of ESRE exhibits a higher expression level of Bmi-1 compared to erythroblasts undergoing restrictive self-renewal and primary proerythroblasts. 
     
     
         37 . The composition of  claim 6 , wherein the ESRE is genetically modified to express one or more therapeutic proteins. 
     
     
         38 . The composition of  claim 6 , wherein the ESRE is genetically modified to express a factor selected from the group consisting of factor IX, factor VIIIc, von Willebrand's factor, tissue plasminogen activator, protein C, protein S, antithrombin III, and any combination thereof. 
     
     
         39 . The composition of  claim 6 , wherein the ESRE are generated by culturing human cells selected from the group consisting of human stem cells and human progenitor cells, to generate a population of definitive erythroid progenitor cells; and expanding the population of definitive erythroid progenitor cells in an expansion medium comprising Epo, SCF, dexamethasone, and a lipid mixture. 
     
     
         40 . The composition of  claim 39 , wherein the human cells are selected from the group consisting of embryonic stem cells, induced-Pluripotent Stem (iPS) cells, adult stem cells, cord cells, bone marrow cells, and a combination thereof. 
     
     
         41 . The composition of  claim 39 , wherein the population of definitive erythroid progenitor cells is derived from embryoid bodies (EB). 
     
     
         42 . The composition of  claim 39 , wherein the cells are cultured for at least about 20 days. 
     
     
         43 . The composition of  claim 39 , wherein the definitive erythroid progenitor cells are contacted with one or more of the group consisting of Bmi-1 protein, Bmi-1 peptide, and a Bmi-1 regulating agent. 
     
     
         44 . The composition of  claim 43 , wherein the Bmi-1 regulating agent is a hedgehog ligand. 
     
     
         45 . The composition of  claim 6 , wherein the expansion media is animal component-free medium supplemented with about 2 U/mL Epo, about 100 ng/mL human recombinant SCF, about 10 −6 M dexamethasone, about 40 ng/mL human recombinant insulin-like growth factor-1, and about 0.4% lipid mixture. 
     
     
         46 . The method of  claim 18 , wherein the ESRE is genetically modified to express a factor selected from the group consisting of factor IX, factor VIIIc, von Willebrand's factor, tissue plasminogen activator, protein C, protein S, antithrombin III, and any combination thereof.