Generation Of A Mesenchymal Stromal Cell Bank From The Pooled Mononuclear Cells Of Multiple Bone Marrow Donors

Abstract

The present invention pertains to an improved mesenchymal stromal cell (MSC) preparation and a method for producing the same. The invention provides a new strategy to isolate MSC from bone marrow mononuclear cells (BM-MNCs) by pooling BM-MNCs of multiple unrelated (third-party) bone marrow donors. The MSC preparation manufactured in accordance with the methodology of the invention is characterized by a stable proliferative capability and an increased immunosuppressive potential when compared to individual donor MSC preparations or a pool of individual MSCs generated from multiple donors. The MSCs prepared according to the invention are particularly useful for medical applications such as the treatment of graft-versus-host disease (GvHD) in recipients with hematopoietic stem cell transplants, patients with autoimmune disorders and as a cell-based therapy in regenerative medicine.

1 .- 22 . (canceled) 
     
     
         23 . A mesenchymal stromal cells (MSC) preparation obtainable by a method comprising the steps of:
 a) pooling individual bone marrow samples obtained from at least two genetically distinct donors to obtain a sample cell-pool; and   b) isolating mesenchymal stromal cells from said sample cell-pool.   
     
     
         24 . The MSC preparation according to  claim 23 , wherein the method further includes the step of culturing said sample cell-pool prior to isolating mesenchymal stromal cells from said sample cell-pool. 
     
     
         25 . The MSC preparation of  claim 23 , wherein said bone marrow sample is a mammalian bone marrow sample and said mesenchymal stromal cell is a mammalian mesenchymal stromal cell. 
     
     
         26 . The MSC preparation according to  claim 23 , wherein said bone marrow samples are bone marrow mononuclear cell samples. 
     
     
         27 . The MSC preparation according to  claim 23 , wherein said bone marrow samples are obtained from at least three, at least four, at least five, at least six, at least seven, or at least eight genetically distinct donors. 
     
     
         28 . The MSC preparation according to  claim 23 , wherein the method further includes the step of storing said isolated mesenchymal stromal cells. 
     
     
         29 . The MSC preparation of  claim 23 , further including the steps of:
 a) determining the hTERT negative and polygenic status of the isolated mesenchymal stromal cells; and   b) enriching the isolated mesenchymal stromal cells to obtain hTERT negative and polygenic isolated mesenchymal stromal cells.   
     
     
         30 . The MSC preparation of claim  7 , wherein the method further includes the step of enriching the hTERT and polygenic isolated, enriched mesenchymal stromal cells to:
 a) At least 95% CD73+ cells, and/or   b) At least 95% CD90+ cells, and/or   c) At least 95% CD 105+ cells, and/or   d) At least 95% HLA-class 1+ cells, and/or   e) Less than 1% CD45+ cells, and/or   f) Less than 1% CD 14+ cells, and/or   g) Less than 1% CD34+ cells, and/or   h) Less than 5% HLA-DR+ cells.   
     
     
         31 . The MSC preparation of  claim 30 , wherein the hTERT and polygenic isolated mesenchymal stromal cells are (a), (b) and (c), and/or (e), (f) and (g). 
     
     
         32 . An isolated cellular composition comprising bone marrow cells from at least two genetically distinct bone marrow donors, made by a process comprising the steps of:
 a) pooling at least two monogenic and genetically distinct bone marrow samples to obtain a sample cell-pool; and   b) isolating a stromal cell fraction from the sample cell-pool to thereby obtain the isolated cellular composition.   
     
     
         33 . The isolated cellular composition of claim  11 , further including the step of isolating mesenchymal stromal cells from the isolated cellular composition to thereby obtain isolated mesenchymal stromal cells. 
     
     
         34 . A method for the treatment of a disease in a subject in need of the treatment, wherein the treatment comprises the administration of a therapeutically effective amount of an MSC-preparation obtainable by a method comprising the steps of:
 a) pooling individual bone marrow samples obtained from at least two genetically distinct donors to obtain a sample cell-pool; and   b) isolating mesenchymal stromal cells from said sample cell-pool.   
     
     
         35 . The method of  claim 34 , wherein the disease is an autoimmune diseases selected from the group consisting of multiple sclerosis, Type 1 diabetes, rheumatoid arthritis, uveitis, autoimmune thyroid disease, inflammatory bowel disease (IBD), scleroderma, Graves' Disease, lupus, Crohn's disease, autoimmune lymphoproliferative disease (ALPS), demyelinating disease, autoimmune encephalomyelitis, autoimmune gastritis (AIG), autoimmune glomerular diseases, and graft-versus-host disease (GvHD). 
     
     
         36 . The MSC preparation of  claim 25 , wherein the mammalian bone marrow sample is a human bone marrow sample and the mammalian mesenchymal stromal cell is a human mesenchymal stromal cell. 
     
     
         37 . The MSC preparation of  claim 23 , wherein the method further includes the step of culturing the isolated mesenchymal stromal cells.