Factors and cells that provide for induction of bone, bone marrow, and cartilage

Abstract

Methods, compositions and kits for producing functional chondrocytes, skeletal cells, bone marrow stromal cells, and progenitor cells thereof are provided. These methods, compositions and kits find use in producing chondrocytes, osteoblasts, stromal cells, and progenitor cells thereof in vivo, or in vitro for transplantation, for experimental evaluation, as a source of lineage- and cell-specific products, and the like, for example for use in treating human disorders of the cartilage, bone and hematopoietic system. In some embodiments, specific combinations of protein factors are identified for reprogramming non-skeletal cells into bones, hematopoietic stroma, and chondrocytes, which may be provided in vitro or in vivo.

1 . A method for regenerating articular cartilage in a human individual, the method comprising:
 administering to an individual in a matrix, at a site where regeneration of articular cartilage is desired, a combination of factors, comprising: human BMP2 protein in an effective dose; and a VEGF inhibitor in a dose effective to induce chondrogenesis;   wherein articular cartilage is regenerated at the site.   
     
     
         2 - 27 . (canceled) 
     
     
         28 . The method of  claim 1 , wherein the matrix is a scaffold, paste or implant, and wherein articular cartilage is formed at the site of the matrix. 
     
     
         29 . The method of  claim 1 , wherein the VEGF inhibitor is an antibody that specifically binds to human VEGF. 
     
     
         30 . The method of  claim 1 , wherein the VEGF inhibitor is a soluble VEGF receptor. 
     
     
         31 . The method of  claim 1 , wherein the VEGF inhibitor is selected from: ABT-869; AEE-788; AG-13736; AG-028262; Angiostatin; bevacizumab; AVE-8062; AZD-2171; sorafenib; BMS-387032; CEP-7055; CHIR-258; CP-547632; 786034; GW-654652; IMC-1C11; KRN-951; PKC-412; benzoylstaurosporine; CGP-41251; midostaurin; STI-412; PTK-787; vatalanib; sunitinib; semaxanib; SU-666; VEGF Trap; Thalidomide; XL-647; XL-999; XL-880; ZD-6474; and ZK-304709. 
     
     
         32 . The method of  claim 1 , wherein the matrix is a biodegradable matrix comprised comprising one or more of matrigel, polylactic acid, polyglycolic acid, poly(lactic-co-glycolic acid); collagen, and alginate. 
     
     
         33 . The method of  claim 1  wherein following the administering steps, an articular cartilage mass of the treated individual is increased at least 50% relative to an untreated control. 
     
     
         34 . A method for regenerating cartilage or bone, the method comprising:
 administering to an individual an effective dose of the human skeletal stem cell;   wherein the cell is selected from human tissue for the phenotype of negative for expression of CD45, CD235, Tie2, and CD31; and positive for expression of podoplanin (PDPN); at a site where regeneration of bone or cartilage is desired.   
     
     
         35 . The method of  claim 34 , wherein the cells are further selected for expression of CD73 and CD164. 
     
     
         36 . The method of  claim 35 , wherein the cells are [PDPN + CD146 − CD73 − CD164 − ] and are implanted to regenerate cartilage. 
     
     
         37 . The method of  claim 35 , wherein the cells are [PDPN + CD146 − CD73 − CD164 + ] and are implanted to regenerate bone. 
     
     
         38 . The method of  claim 35 , wherein the cells are [PDPN + CD146 − CD73 + CD164 + ] and are implanted for endochondral regeneration. 
     
     
         39 . The method of  claim 36 , wherein an effective dose of a VEGF inhibitor is administered to induce a chondrogenic fate in the cells. 
     
     
         40 . The method of  claim 37 , wherein an effective dose of a Wnt protein is administered to induce an osteogenic fate in the cells. 
     
     
         41 . The method of  claim 34 , wherein the cells are provided in a scaffold, paste or implant, and wherein cartilage or bone is formed at the site of the implant. 
     
     
         42 . A method of isolating a human skeletal stem cell population, the method comprising:
 selecting a cell from a bone tissue for the phenotype of negative for expression of CD45, CD235, Tie2, and CD31; and positive for expression of podoplanin (PDPN).   
     
     
         43 . The method of  claim 43 , wherein the cells are further selected for expression of CD73 and CD164. 
     
     
         44 . A method for regenerating articular cartilage in a human individual, the method comprising:
 administering to the individual at a site where regeneration of cartilage is desired, a composition consisting of human BMP2, wherein human BMP2 is provided at a concentration from 1 mg/ml to 1 mg/ml; an effective dose of the isolated human mesenchymal stem cells (MSC) that express BMP receptor (BMPR1) to reprogram the MSC to human skeletal stem cells (hSSC), and the VEGF inhibitor is administered in a dose effective to induce a chondrogenic fate in the hSSC;   wherein following the administering steps, wherein articular cartilage is regenerated at the site.   
     
     
         45 . The method of  claim 44 , wherein the combination of cells, BMP2 and VEGF inhibitor are provided in a matrix. 
     
     
         46 . The method of  claim 44 , wherein the VEGF inhibitor is an antibody that specifically binds to human VEGF or a soluble VEGF receptor. 
     
     
         47 . The method of  claim 44 , wherein the VEGF inhibitor is selected from: ABT-869; AEE-788; AG-13736; AG-028262; Angiostatin; bevacizumab; AVE-8062; AZD-2171; sorafenib; BMS-387032; CEP-7055; CHIR-258; CP-547632; 786034; GW-654652; IMC-1C11; KRN-951; PKC-412; benzoylstaurosporine; CGP-41251; midostaurin; STI-412; PTK-787; vatalanib; sunitinib; semaxanib; SU-666; VEGF Trap; Thalidomide; XL-647; XL-999; XL-880; ZD-6474; and ZK-304709. 
     
     
         48 . The method of  claim 44  wherein following the administering steps, an articular cartilage mass of the treated individual is increased at least 50% relative to an untreated control.