US2004033956A1PendingUtilityA1

Method of promoting production of living tissue equivalents

55
Assignee: UNIV SOUTHERN CALIFORNIAPriority: Mar 11, 1998Filed: Jun 18, 2002Published: Feb 19, 2004
Est. expiryMar 11, 2018(expired)· nominal 20-yr term from priority
Inventors:Kathleen E. RodgersGere S. Dizerega
C12N 2533/54C12N 2533/70C12N 2501/32C12N 5/0629
55
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Claims

Abstract

The present invention provides methods, improved cell culture medium and kits for accelerating the generation of tissue equivalents, and for improving the quality of tissue equivalents, by growth in the presence of angiotensinogen, AI, AI analogues, AI fragments and analogues thereof, AII, AII analogues, AII fragments and analogues thereof and/or AII AT 2 type 2 receptor agonists.

Claims

exact text as granted — not AI-modified
We claim:  
     
         1 . An improved method for producing a tissue equivalent, the improvement comprising contacting the tissue equivalent with an amount effective to accelerate generation of tissue equivalents of at least one active agent comprising a sequence consisting of at least three contiguous amino acids of groups R 1 -R 8  in the sequence of general formula I  
       R 1 -R 2 -R 3 -R 4 -R 5 -R 6 -R 7 -R 8    in which R 1  and R 2  together form a group of formula    X-R A -R B -,    wherein X is H or a one to three peptide group    R A  is selected from Asp, Glu, Asn, Acpc, Ala, Me 2 Gly, Pro, Bet, Glu(NH 2 ), Gly, Asp(NH 2 ) and Sue;    R B  is selected from Arg, Lys, Ala, Om, Ser(Ac), Sar, D-Arg and D-Lys;    R 3  is selected from the group consisting of Val, Ala, Leu, norLeu, Ile, Gly, Pro, Aib, Acpc, Lys and Tyr;    R 4  is selected from the group consisting of Tyr, Tyr(PO 3 ) 2 , Thr, Ser, Ala, homoSer and azaTyr;    R 5  is selected from the group consisting of Ile, Ala, Leu, norLeu, Val and Gly;    R 6  is His, Arg or 6-NH 2 -Phe;    R 7  is Pro or Ala; and    R 8  is selected from the group consisting of Phe, Phe(Br), Ile and Tyr, excluding sequences including R 4  as a terminal Tyr group.    
     
     
         2 . The method of  claim 1  wherein the active agent is selected from the group consisting of angiotensinogen, SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO: 32, SEQ ID NO:33, SEQ ID NO: 34; SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, and SEQ ID NO:42.  
     
     
         3 . The method of  claim 1  wherein the active agent is SEQ ID NO:1, SEQ ID NO:4, SEQ ID NO:18, SEQ ID NO:31 SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, or SEQ ID NO:42.  
     
     
         4 . The method of  claim 1  wherein the concentration of active agent is between about 0.1 ng/kg and about 1.0 mg/kg.  
     
     
         5 . The method of  claim 1  where the tissue equivalent is selected from the group consisting of a skin, dermis, bone, bone marrow, pancreas, heart valve, vascular graft, cartilage, ligament, liver, and kidney tissue equivalent.  
     
     
         6 . An improved chemically defined medium for the culture of tissue equivalents, wherein the improvement comprises contacting the tissue equivalent with an amount effective to accelerate generation of tissue equivalents of at least one active agent comprising a sequence consisting of at least three contiguous amino acids of groups R 1 -R 8  in the sequence of general formula I  
       R 1 -R 2 -R 3 -R 4 -R 5 -R 6 -R 7 -R 8    in which R 1  and R 2  together form a group of formula    X-R A -R B -,    wherein X is H or a one to three peptide group    R A  is selected from Asp, Glu, Asn, Acpc, Ala, Me 2 Gly, Pro, Bet, Glu(NH 2 ), Gly, Asp(NH2) and Suc;    R B  is selected from Arg, Lys, Ala, Om, Ser(Ac), Sar, D-Arg and D-Lys;    R 3  is selected from the group consisting of Val, Ala, Leu, norLeu, Ile, Gly, Pro, Aib, Acpc, Lys and Tyr;    R 4  is selected from the group consisting of Tyr, Tyr(PO 3 ) 2 , Thr, Ser, Ala, homoSer and azaTyr;    R 5  is selected from the group consisting of Ile, Ala, Leu, norLeu, Val and Gly;    R 6  is His, Arg or 6-NH 2 -Phe;    R 7  is Pro or Ala; and    R 8  is selected from the group consisting of Phe, Phe(Br), Ile and Tyr, excluding sequences including R 4  as a terminal Tyr group.    
     
     
         7 . The improved chemically defined medium of  claim 6  wherein the active agent is selected from the group consisting of angiotensinogen, SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO: 32, SEQ ID NO:33, SEQ ID NO: 34; SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, and SEQ ID NO:42.  
     
     
         8 . The improved chemically defined medium of  claim 6  wherein the active agent is SEQ ID NO:1, SEQ ID NO:4, SEQ ID NO:18, SEQ ID NO:31,SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, or SEQ ID NO:42.  
     
     
         9 . The improved chemically defined medium of  claim 6  wherein the concentration of active agent is between about 0.1 ng/ml and about 1.0 mg/ml.  
     
     
         10 . The improved chemically defined medium of  claim 6  where the tissue equivalent is selected from the group consisting of a skin, dermis, bone, bone marrow, pancreas, heart valve, vascular graft, cartilage, ligament, liver, and kidney tissue equivalent.  
     
     
         11 . An improved kit for the culture of tissue equivalents, wherein the improvement comprises providing 
 a). an amount effective to accelerate generation of tissue equivalents of at least one active agent comprising a sequence consisting of at least three contiguous amino acids of groups R 1 -R 8  in the sequence of general formula I    R 1 -R 2 -R 3 -R 4 -R 5 -R 6 -R 7 -R 8      in which R 1  and R 2  together form a group of formula    X-R A -R B -,    wherein X is H or a one to three peptide group    R A  is selected from Asp, Glu, Asn, Acpc, Ala, Me 2 Gly, Pro, Bet, Glu(NH 2 ), Gly, Asp(NH 2 ) and Suc;    R B  is selected from Arg, Lys, Ala, Orn, Ser(Ac), Sar, D-Arg and D-Lys;    R 3  is selected from the group consisting of Val, Ala, Leu, norLeu, Ile, Gly, Pro, Aib, Acpc, Lys and Tyr;    R 4  is selected from the group consisting of Tyr, Tyr(PO 3 ) 2 , Thr, Ser, Ala, homoSer and azaTyr;    R 5  is selected from the group consisting of Ile, Ala, Leu, norLeu, Val and Gly;    R 6  is His, Arg or 6-NH 2 -Phe;    R 7  is Pro or Ala; and    R 8  is selected from the group consisting of Phe, Phe(Br), Ile and Tyr, excluding sequences including R 4  as a terminal Tyr group; and    b) and instructions for use of the active agent to accelerate generation of tissue equivalents.    
     
     
         12 . The kit of  claim 11  further comprising tissue culture medium.  
     
     
         13 . The kit  claim 11  wherein the active agent is selected from the group consisting of angiotensinogen, SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO: 32, SEQ ID NO:33, SEQ ID NO: 34; SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, and SEQ ID NO:42.  
     
     
         14 . The kit of  claim 11  wherein the active agent is SEQ ID NO:1, SEQ ID NO:4, SEQ ID NO:18, SEQ ID NO:31, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, or SEQ ID NO:42.  
     
     
         15 . The kit of  claim 11  wherein the concentration of active agent is between about 0.1 ng/ml and about 1.0 mg/ml.  
     
     
         16 . An improved method for producing a tissue equivalent, the improvement comprising contacting the tissue equivalent with an amount effective to accelerate generation of tissue equivalents of at least one active agent comprising a sequence of the following general formula:  
       R1-Arg-R2-R3-R4-His-Pro-R5  wherein R1 is selected from the group consisting of H, Gly and Asp;    R2 is selected from the group consisting of Val, Pro, and Acpc;    R3 is selected from the group consisting of Tyr and Tyr(PO 3 ) 2 ;    R4 is selected from the group consisting of Ala, Val, Ile, Leu, and norLeu; and    R5 is Phe, Ile, or is absent.    
     
     
         17 . The method of  claim 16  wherein the active agent is selected from the group consisting of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:26, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, or SEQ ID NO:42.  
     
     
         18 . The method of  claim 16  wherein the concentration of active agent is between about 0.1 ng/kg and about 1.0 mg/kg.  
     
     
         19 . The method of  claim 16  where the tissue equivalent is selected from the group consisting of a skin, dermis, bone, bone marrow, pancreas, heart valve, vascular graft, cartilage, ligament, collagen lattice, liver, and kidney tissue equivalent.  
     
     
         20 . The method of  claim 16  where the tissue equivalent is selected from the group consisting of a collagen lattice and dermis tissue equivalent.  
     
     
         21 . An improved chemically defined medium for the culture of tissue equivalents, wherein the improvement comprises contacting the tissue equivalent with an amount effective to accelerate generation of tissue equivalents of at least one active agent comprising a sequence of the following general formula:  
       R1-Arg-R2-R3-R4-His-Pro-R5  wherein R is selected from the group consisting of H, Gly and A sp;    R2 is selected from the group consisting of Val, Pro, and Acpc;    R3 is selected from the group consisting of Tyr and Tyr(PO 3 ) 2 ;    R4 is selected from the group consisting of Ala, Val, Ile, Leu, and norLeu; and    R5 is Phe, Ile, or is absent.    
     
     
         22 . The chemically defined medium of  claim 21  wherein the active agent is selected from the group consisting of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:26, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, or SEQ ID NO:42.  
     
     
         23 . The method of  claim 21  wherein the concentration of active agent is between about 0.1 ng/kg and about 1.0 mg/kg.  
     
     
         24 . The method of  claim 21  where the tissue equivalent is selected from the group consisting of a skin, dermis, bone, bone marrow, pancreas, heart valve, vascular graft, cartilage, ligament, collagen lattice, liver, and kidney tissue equivalent.  
     
     
         25 . The method of  claim 21  where the tissue equivalent is selected from the group consisting of a collagen lattice and dermis tissue equivalent.  
     
     
         26 . An improved kit for the culture of tissue equivalents, wherein the improvement comprises providing 
 a). an amount effective to accelerate generation of tissue equivalents of at least one active agent comprising a sequence of the following general formula:    R1-Arg-R2-R3-R4-His-Pro-R5  wherein R1 is selected from the group consisting of H, Gly and Asp;    R2 is selected from the group consisting of Val, Pro, and Acpc;    R3 is selected from the group consisting of Tyr and Tyr(PO 3 ) 2 ;    R4 is selected from the group consisting of Ala, Val, Ile, Leu, and norLeu; and    R5 is Phe, Ile, or is absent; and      b) instructions for use of the active agent to accelerate generation of tissue equivalents.    
     
     
         27 . The kit of  claim 26  wherein the active agent is selected from the group consisting of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:26, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, or SEQ ID NO:42.  
     
     
         28 . The kit of  claim 26  wherein the concentration of active agent is between about 0.1 ng/kg and about 1.0 mg/kg.  
     
     
         29 . The kit of  claim 26  where the tissue equivalent is selected from the group consisting of a skin, dermis, bone, bone marrow, pancreas, heart valve, vascular graft, cartilage, ligament, collagen lattice, liver, and kidney tissue equivalent.  
     
     
         30 . The kit of  claim 16  where the tissue equivalent is selected from the group consisting of a collagen lattice and dermis tissue equivalent.

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