Crystal Structures Comprising Elastin-Like Peptides

Abstract

The present invention relates to new biomimetic mineralized apatite structures. The present invention also relates to processes for the production of new biomimetic mineralized apatite structures based on natural and synthetic protein scaffolds. In particular, the invention provides synthetic crystal having a hierarchical structure formed on an elastin-like polypeptide membrane or hydrogel. The invention also provides methods of making such crystals, both in vivo and in vitro, as well as kits comprising membranes or hydrogels with cross-linking agents and/or mineralization solutions. The invention also provides the use of such structures in methods of treatment.

1 . Synthetic crystal having a hierarchical structure formed on an elastin-like polypeptide membrane or hydrogel. 
     
     
         2 . The synthetic crystal of  claim 1  having a hierarchical structure at four different length scales. 
     
     
         3 - 5 . (canceled) 
     
     
         6 . The synthetic crystal of  claim 1 , wherein the crystal is apatite. 
     
     
         7 - 8 . (canceled) 
     
     
         9 . The synthetic crystal of  claim 1 , wherein the elastin-like polypeptide membrane comprises a peptide sequence or a bioactive sequence. 
     
     
         10 . The synthetic crystal of synthetic crystal of  claim 9 , wherein the peptide sequence is selected from the group consisting of: 
       
         
           
                 
               
                   (a) 
                 
                   (SEQ ID NO: 27) 
                 
                   MGSSHHHHHHSSGLVPRGSHMESLLP-(VPGIGVPGIGVPGKGVPGIG 
                 
                     
                 
                   VPGIGVPGIGVPGIGVPGKGVPGIGVPGIGAVTGRGDSPASSVPGIGV 
                 
                     
                 
                   PGIGVPGKGVPGIGVPGIGVPGIGVPGIGVPGKGVPGIGVPGIG) 6 -V; 
                 
             
                
                
                
                
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (b) 
                 
                   (SEQ ID NO: 28) 
                 
                   MESLLP-(VPGIGVPGIGVPGKGVPGIGVPGIGEEIQIGHIPREDVDYHL 
                 
                   YPVPGIGVPGIGVPGKGVPGIGVPGIGVGVAPGVGVAPGVGVAPG) 10 -V; 
                 
             
                
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (c) 
                 
                   (SEQ ID NO: 29) 
                 
                   MESLLP-((VPGVGVPGVGVPGEGVPGVGVPGVG) 10 -(VGIPG) 60 ) 2 -V; 
                 
             
                
                
                
               
            
           
         
       
       
         
           
                 
                 
               
                     
                   (d) 
                 
                     
                   (SEQ ID NO: 30) 
                 
                     
                   MESLLP-(VPGIGVPGIGVPGKGVPGIGVPGIGVPGIGVPGIGVP 
                 
                     
                   GKGVPGIGVPGIG) 12 -V ; 
                 
             
                
                
                
                
               
            
           
         
       
       
         
           
                 
                 
               
                     
                   (e) 
                 
                     
                   (SEQ ID NO: 31) 
                 
                     
                   (VPGVGVPGVGVPGEGVPGVGVPGVG) 15 -V; 
                 
             
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (f) 
                 
                   (SEQ ID NO: 32) 
                 
                   MESLLP-[((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 -DDDEEKFLRRIGRFG- 
                 
                   ((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 ] 3 -V; 
                 
             
                
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (g) 
                 
                   (SEQ ID NO: 33) 
                 
                   MESLLP-[((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 -DDDEEKFLRRIGRFG- 
                 
                   ((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 ] 3 -(VPAVG) 20 -V; 
                 
             
                
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (h) 
                 
                   (SEQ ID NO: 34) 
                 
                   MESLLP-[((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 -DDDEEKFLRRIGRFG- 
                 
                     
                 
                   ((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 ] 3 -(VPAVG) 20 - 
                 
                     
                 
                   [((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 -DDDEEKFLRRIGRFG- 
                 
                     
                 
                   ((VPGIG) 2 VPGKG(VPGIG) 2 ) 2 ] 3 -V; 
                 
             
                
                
                
                
                
                
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (i) 
                 
                   (SEQ ID NO: 35) 
                 
                   MESLLP-(VPGVGVPGVGVPGEGVPGVGVPGVG) 10 -(VPAVG) 40 -V; 
                 
             
                
                
                
               
            
           
         
       
       
         
           
                 
               
                   (j) 
                 
                   (SEQ ID NO: 36) 
                 
                   MESLLP-(VPGVGVPGVGVPGEGVPGVGVPGVG) 10 -(VPAVG) 60 -V; 
                 
             
                
                
                
               
            
           
         
         (k) MESLLP-(VPGVGVPGVGVPGEGVPGVGVPGVG) 20 -(VPAVG) 40 -V (SEQ ID NO:37); and 
       
       
         
           
                 
                 
               
                     
                   (l) 
                 
                     
                   (SEQ ID NO: 38) 
                 
                     
                   MESLLP-(VPGVGVPGVGVPGEGVPGVGVPGVG) 10 -(VPAVG) 40 - 
                 
                     
                   (VPGVGVPGVGVPGEGVPGVGVPGVG) 10 -V. 
                 
             
                
                
                
                
               
            
           
         
       
     
     
         11 . The synthetic crystal of  claim 9 , wherein the bioactive sequence is selected from the group consisting of MGSSHHHHHHSSGLVPRGSHMESLLP-(((VPGIG) 2 (VPGKG)(VPGIG) 2 ) 2 AVTGRGDSPA SS((VPGIG) 2 (VPGKG)(VPGIG) 2 ) 2 ) 6 (SEQ ID NO:39), RGDS (SEQ ID NO:1); and MESLLP-(((VPGIG) 2 (VPGKG)(VPGIG) 2 ) 2 -DDDEEKFLRRIGRFG((VPGIG) 2 (VPGKG) (VPGIG) 2 ) 2 ) 3  (SEQ ID NO:32). 
     
     
         12 . The synthetic crystal of  claim 9 , wherein the bioactive sequence is selected from the group consisting of Gly-Xaa-Xaa-Xaa-Xaa (SEQ ID NO:3), Xaa-Gly-Xaa-Xaa-Xaa (SEQ ID NO:4), Xaa-Xaa-Gly-Xaa-Xaa (SEQ ID NO:5), Xaa-Xaa-Xaa-Gly-Xaa (SEQ ID NO:6) and Xaa-Xaa-Xaa-Xaa-Gly (SEQ ID NO:7), wherein Xaa is any amino acid. 
     
     
         13 . The synthetic crystal of  claim 9 , wherein the bioactive sequence is selected from the group consisting of (Gly-Xaa-Xaa-Xaa-Xaa)y (SEQ ID NO:13), (Xaa-Gly-Xaa-Xaa-Xaa)y (SEQ ID NO:14), (Xaa-Xaa-Gly-Xaa-Xaa)y (SEQ ID NO:15), (Xaa-Xaa-Xaa-Gly-Xaa)y (SEQ ID NO:16) and (Xaa-Xaa-Xaa-Xaa-Gly)y (SEQ ID NO:17), wherein Xaa is any amino acid and wherein y is the number of repeats. 
     
     
         14 . The synthetic crystal of  claim 9 , wherein the bioactive sequence is selected from the group consisting of (VPGXG; SEQ ID NO:19), (PGIPG; SEQ ID NO:21), (PVGSG; SEQ ID NO:23) and (VGFPG; SEQ ID NO:25) wherein X is any amino acid apart from proline. 
     
     
         15 . The synthetic crystal of  claim 9 , wherein the bioactive sequence is selected from the group consisting of (VPGXG)y (SEQ ID NO:20), (PGIPG)y (SEQ ID NO:22), (PVGSG)y (SEQ ID NO:24) and (VGFPG)y (SEQ ID NO:26), wherein X is any amino acid apart from proline and wherein y is the number of repeats. 
     
     
         16 - 42 . (canceled) 
     
     
         43 . A process for producing hierarchically ordered crystal structures comprising the step
 contacting an elastin-like polypeptide membrane or hydrogel with a solution of calcium and phosphate ions.   
     
     
         44 . The process of  claim 43 , wherein the solution further comprises fluoride ions. 
     
     
         45 . The process of  claim 43 , wherein the ELP membrane or hydrogel is cross-linked. 
     
     
         46 - 91 . (canceled) 
     
     
         92 . A method of treatment of or prevention of demineralisation of teeth, dental disease, demineralisation of bone, low bone density, bone disease, bone defect, or tooth hypersensitivity in a subject, comprising administration to the subject a synthetic crystal as defined in  claim 1 . 
     
     
         93 . The method of treatment or of prevention of  claim 92  wherein:
 the dental disease is dental caries, dental erosion, alveolar bone erosion, periodontitis, peri-implantitis or dental pulp disease; 
 the bone disease is osteoporosis, osteoarthritis, osteosclerosis, osteogenesis imperfecta, Paget's disease of bone, metabolic bone disease, osteomalacia, osteopenia, arthritis or sarcoma; 
 the bone defect is a bone fracture, bone fracture associated with trauma, bone injury, bone cavity, bone lesion, or damage to the bone-cartilage interface, bone-ligament interface, or a bone-tendon interface. 
 
     
     
         94 - 99 . (canceled) 
     
     
         100 . A method comprising applying the synthetic crystal of  claim 1  to a tooth surface or a bone surface. 
     
     
         101 . A dental enamel or bone graft comprising the synthetic crystal of  claim 1 . 
     
     
         102 - 103 . (canceled) 
     
     
         104 . A medical device, medical implant, synthetic graft, coating, prosthesis, orthosis, paste, malleable putty or film comprising the synthetic crystal of  claim 1 . 
     
     
         105 . A kit comprising an ELP membrane or hydrogel, and further comprising a mineralising solution and/or a cross-linker. 
     
     
         106 - 109 . (canceled) 
     
     
         110 . A method comprising contacting an elastin-like polypeptide membrane or hydrogel with a solution of calcium and phosphate ions in vivo. 
     
     
         111 - 127 . (canceled)