Mesenchymal stromal cells and uses related thereto

Abstract

The present invention generally relates to novel preparations of mesenchymal stromal cells (MSCs) derived from hemangioblasts, methods for obtaining such MSCs, and methods of treating a pathology using such MSCs. The methods of the present invention produce substantial numbers of MSCs having a potency-retaining youthful phenotype, which are useful in the treatment of pathologies.

1 . A pharmaceutical preparation suitable for use in a mammalian patient, comprising at least 10 6  mesenchymal stromal cells and a pharmaceutically acceptable carrier, wherein the mesenchymal stromal cells have replicative capacity to undergo at least 10 population doublings in cell culture with less than 25 percent of the cells undergoing cell death, senescing or differentiating into non-MSC cells by the tenth doubling. 
     
     
         2 .- 4 . (canceled) 
     
     
         5 . The pharmaceutical preparation of  claim 1 , comprising less than one percent of a cell type other than a mesenchymal stromal cell. 
     
     
         6 - 8 . (canceled) 
     
     
         9 . The pharmaceutical preparation of  claim 1 , wherein the mesenchymal stromal cells are HLA-genotypically identical or genomically identical. 
     
     
         10 . (canceled) 
     
     
         11 . The pharmaceutical preparation of  claim 1 , wherein at least 30% of the mesenchymal stromal cells are positive for CD10. 
     
     
         12 . The pharmaceutical preparation of  claim 11 , wherein at least 60% of the mesenchymal stromal cells are positive for markers CD73, CD90, CD105, CD13, CD29, CD44, CD166 and HLA-ABC. 
     
     
         13 . The pharmaceutical preparation of  claim 1 , wherein less than 30% of the mesenchymal stromal cells are positive for markers CD31, CD34, CD45, CD133, FGFR2, CD271, Stro-1, CXCR4 and TLR3. 
     
     
         14 .- 15 . (canceled) 
     
     
         16 . The pharmaceutical preparation of  claim 1 , wherein the mesenchymal stromal cells have a statistically significant decreased content and/or enzymatic activity, relative to mesenchymal stromal cell preparations derived from bone marrow that have undergone five population doublings, of proteins involved in one or more of (i) cell cycle regulation and cellular aging, (ii) cellular energy and/or lipid metabolism, and (iii) apoptosis. 
     
     
         17 . The pharmaceutical preparation of  claim 1 , wherein the mesenchymal stromal cells have a statistically significant increased content and/or enzymatic activity of proteins involved in cytoskeleton structure and cellular dynamics relating thereto, relative to mesenchymal stromal cell preparations derived from bone marrow. 
     
     
         18 . The pharmaceutical preparation of  claim 1 , wherein the mesenchymal stromal cells do not undergo more than a 75 percent increase in cells having a forward-scattered light value, measured by flow cytometry, greater than 5,000,000 over 10 population doublings in culture. 
     
     
         19 . The pharmaceutical preparation of  claim 1 , wherein the mesenchymal stromal cells, in a resting state, express mRNA encoding Interleukin-6 at a level which is less than ten percent of the IL-6 mRNA level expressed by mesenchymal stromal cells preparations, in a resting state, derived from bone marrow or adipose tissue. 
     
     
         20 . The pharmaceutical preparation of  claim 1 , which is suitable for administration to a human patient. 
     
     
         21 .- 27 . (canceled) 
     
     
         28 . The pharmaceutical preparation of  claim 1 , wherein said mesenchymal stromal cells are human cells. 
     
     
         29 .- 31 . (canceled) 
     
     
         32 . The pharmaceutical preparation of  claim 1 , wherein said preparation comprises at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% mesenchymal stromal cells. 
     
     
         33 . The pharmaceutical preparation of  claim 1 , wherein at least 50% of said mesenchymal stromal cells are positive for (i) at least one of CD10, CD24, IL-11, AIRE-1, ANG-1, CXCL1, CD105, CD73 and CD90; (ii) at least one of CD10, CD24, IL-11, AIRE-1, ANG-1, CXCL1, CD105, CD73, CD90, CD105, CD13, CD29, CD44, CD166, CD274, and HLA-ABC; (iii) CD105, CD73 and/or CD90 or (iv) any combination thereof. 
     
     
         34 .- 39 . (canceled) 
     
     
         40 . The pharmaceutical preparation of  claim 1 , wherein said mesenchymal stromal cells are not derived from bone marrow and the potency of the preparation in an immune regulatory assay is greater than the potency of a preparation of bone marrow derived mesenchymal stromal cells. 
     
     
         41 . The pharmaceutical preparation of  claim 40 , wherein potency is assayed by an immune regulatory assay that determines the EC50 dose. 
     
     
         42 . The pharmaceutical preparation of  claim 1 , wherein the preparation retains between about 50 and 100% of its proliferative capacity after ten population doublings. 
     
     
         43 .- 103 . (canceled) 
     
     
         104 . A kit comprising the mesenchymal stromal cells or the preparation of mesenchymal stromal cells of  claim 1 , wherein said cells or preparation of cells are frozen or cryopreserved. 
     
     
         105 . (canceled) 
     
     
         106 . A method for treating a disease or disorder, comprising administering an effective amount of the mesenchymal stromal cells or the preparation of mesenchymal stromal cells according to  claim 1  to a subject in need thereof. 
     
     
         107 .- 108 . (canceled) 
     
     
         109 . The method of  claim 106 , wherein the disease or disorder is selected from multiple sclerosis, systemic sclerosis, hematological malignancies, myocardial infarction, organ transplantation rejection, chronic allograft nephropathy, cirrhosis, liver failure, heart failure, GvHD, tibial fracture, left ventricular dysfunction, leukemia, myelodysplastic syndrome, Crohn's disease, diabetes, chronic obstructive pulmonary disease, osteogenesis imperfecta, homozygous familial hypocholesterolemia, treatment following meniscectomy, adult periodontitis, vasculogenesis in patients with severe myocardial ischemia, spinal cord injury, osteodysplasia, critical limb ischemia, diabetic foot disease, primary Sjogren's syndrome, osteoarthritis, cartilage defects, multisystem atrophy, amyotropic lateral sclerosis, cardiac surgery, refractory systemic lupus erythematosis, living kidney allografts, nonmalignant red blood cell disorders, thermal burn, Parkinson's disease, microfractures, epidermolysis bullosa, severe coronary ischemia, idiopathic dilated cardiomyopathy, osteonecrosis femoral head, lupus nephritis, bone void defects, ischemic cerebral stroke, after stroke, acute radiation syndrome, pulmonary disease, arthritis, bone regeneration, or combinations thereof. 
     
     
         110 . The method of  claim 106 ,
 wherein the disease or disorder is uveitis.   
     
     
         111 .- 125 . (canceled) 
     
     
         126 . The method of  claim 106 , wherein the mesenchymal stromal cells are mitotically inactivated. 
     
     
         127 . The composition of  claim 1 , wherein said mesenchymal stromal cells are able to undergo
 (a) at least 10 population doublings and wherein the 10 population doublings occur within about 27 days, less than about 26 days, than 25 days, than about 24 days, less than about 23 days, less than about 22 days, or lower;   (b) at least 15 population doublings;   (c) at least 20, 25, 30, 35, 40, 45, 50 or more population doublings; or   (d) at least 15 population doublings, at least 20 population doublings, or at least 25 population doublings in culture.   
     
     
         128 . The composition of  claim 1 , wherein
 (a) at least 30% of the mesenchymal stromal cells are positive for CD10;   (b) at least 60% of the mesenchymal stromal cells are positive for markers CD73, CD90, CD105, CD13, CD29, CD44, CD166 and HLA-ABC; or   (c) less than 30% of the mesenchymal stromal cells are positive for markers CD31, CD34, CD45, CD133, FGFR2, CD271, Stro-1, CXCR4 and TLR3.