Delivery of bioactive compounds to an organism

• US 5,869,041 A
• Filed: 09/13/1996
• Issued: 02/09/1999
• Est. Priority Date: 01/12/1996
• Status: Expired due to Fees

• Current Assignee: Miriam Hospital (Lifespan Corporation)
• Original Assignee: Miriam Hospital (Lifespan Corporation)
• Inventors: Vandenburgh, Herman H.
• Primary Examiner(s): Crouch, Deborah
• Application Number:
  US08/712,111
• Time in Patent Office: 879 Days
• Field of Search: 435/325, 435/373, 435/377, 424/93.1, 424/93.2, 424/93.21, 514/44
• US Class Current: 424/93.21
• CPC Class Codes: A61K 38/1875 Bone morphogenic factor; Osteogenins; Osteogenic factor; Bone-inducing factor
• CPC Class Codes: A61K 38/27 Growth hormone [GH], i.e. somatotropin
• CPC Class Codes: A61K 48/00 Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
• CPC Class Codes: C12M 21/08 for producing artificial tissue or for ex-vivo cultivation of tissue prostheses A61F2/00, grafts A61L27/00
• CPC Class Codes: C12M 23/02 Form or structure of the vessel large containers B65D88/00
• CPC Class Codes: C12N 5/0658 Skeletal muscle cells, e.g. myocytes, myotubes, myoblasts
• CPC Class Codes: Y10S 623/915 Method or apparatus for preparing biological material
• CPC Class Codes: Y10S 977/924 using nanostructure as support of dna analysis

Abstract

Disclosed herein is a method of delivering a bioactive compound to an organism that involves growing individual cells in vitro under conditions that allow the formation of an organized tissue, at least a subset of the cells containing a foreign DNA sequence which mediates the production of the bioactive compound; and implanting the organized tissue into the organism, whereby the bioactive compound is produced and delivered to the organism. Also disclosed herein is an in vitro method for producing a tissue having in vivo-like gross and cellular morphology that involves providing precursor cells of the tissue; mixing the cells with a solution of extracellular matrix components to create a suspension; placing the suspension in a vessel having a three dimensional geometry approximating the in vivo gross and cellular morphology of the tissue and having attachment surfaces coupled thereto; allowing the suspension to coalesce; and culturing the cells under conditions in which the cells form an organized tissue connected to the attachment surfaces. Also disclosed herein is an apparatus for producing in vitro a tissue having in vivo-like gross and cellular morphology. This apparatus includes a vessel having a three dimensional geometry approximating the in vivo morphology of the tissue and tissue attachment surfaces coupled thereto.

Claims

1. A method of delivering a protein to a mammal comprising the steps of:

growing in vitro a plurality of cells from the same species as said mammal, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, and wherein said cells are mixed with an extracellular matrix to create a suspension;

placing said suspension in a vessel having a three-dimensional geometry approximating the in vivo gross morphology of a tissue of interest, said vessel having attachment surfaces thereon; and

allowing the suspension to coalesce;

culturing said coalesced suspension under conditions in which said cells connect to said attachment surfaces and form an organized tissue having an in vivo-like gross and cellular morphology of said tissue of interest, and wherein the organized tissue further is comprised of post-mitotic cells; and

implanting said tissue into said mammal, whereby said protein is produced and delivered to said mammal, whereby said protein is of a type or produced in an amount not produced by said tissue of interest lacking said foreign DNA sequence, wherein said protein is produced sufficiently to provide a therapeutic effect to the mammal upon implantation of said organized tissue into said mammal.

2. The method of claim 1, further comprising the steps of:

removing said tissue from said mammal to terminate delivery of said protein.

3. The method of claim 2, further comprising, following said removal step, the step of:

culturing said tissue in vitro under conditions which preserve its in vivo viability.

4. The method of claim 3, further comprising, following said culturing step, the step of:

reimplanting said tissue into said mammal to deliver said protein to said mammal.

5. The method of claim 1, wherein said tissue is implanted into the tissue of origin of at least one of said cells.

6. The method of claim 1, wherein said DNA sequence mediates the production of two proteins.

7. The method of claim 1, wherein said protein is a growth factor.

8. The method of claim 1, wherein, during said growing step, a force is exerted parallel to a dimension of said tissue.

9. The method of claim 8, wherein said force is exerted on said individual cells during growth in vitro and on said organized tissue during implantation in vivo.

10. The method of claim 1, wherein said tissue comprises skeletal muscle.

11. The method of claim 1, wherein said tissue comprises myotubes.

12. The method of claim 1, wherein said cells comprise myotubes.

13. The method of claim 1, wherein said mammal is a human.

14. A method of delivering a protein to a mammal comprising the steps of:

growing in vitro a plurality of mammalian cells from the same species as said mammal, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, and wherein said cells are mixed with an extracellular matrix to create a suspension;

placing said suspension in a vessel wherein the cells form an organized tissue of interest having a three dimensional cellular organization which is retained upon implantation into a mammal, and wherein the organized tissue is further comprised of post-mitotic cells; and

implanting said organized tissue into said mammal, whereby said protein is produced and delivered to said mammal sufficiently to provide a therapeutic effect to the mammal, whereby said protein is of a type or produced in an amount not produced by a tissue lacking said foreign DNA sequence.

15. A method of providing a protein to a mammal in therapeutic need thereof:

implanting into said mammal an organized tissue having a three-dimensional geometry approximating the in vivo gross morphology of a tissue of interest, wherein the organized tissue is comprised of post-mitotic cells from the same species as said mammal and wherein at least a subset of cells of said organized tissue comprises a foreign DNA sequence operatively linked to a promoter and encoding a protein, and wherein said protein is produced sufficiently to provide a therapeutic effect to the mammal upon implantation of said organized tissue into said mammal.

17. The method of claim 15 or 16, said protein being growth hormone.

28. An organized tissue comprising post-mitotic cells, said organized tissue having an in vivo gross cellular morphology and producing a protein of a type or produced in an amount not produced normally by a tissue of interest, produced according to the method of claim 17.

16. A method of providing a protein to a mammal in therapeutic need thereof:

implanting into said mammal an organized tissue comprising post-mitotic cells from the same species as said mammal and having a three-dimensional geometry approximating the in vivo gross morphology of a tissue of interest that is retained upon implantation of said organized tissue into a mammal, wherein at least a subset of the cells of said organized tissue comprise a foreign DNA sequence operatively linked to a promoter and encoding a protein, and wherein said protein is produced sufficiently to provide a therapeutic effect to the mammal upon implantation of said organized tissue into said mammal.

18. An in vitro method for producing an organized tissue which has an in vivo-like gross and cellular morphology comprising the steps of:

providing cells of said tissue, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, wherein said cells are mixed with an extracellular matrix to create a suspension;

placing said suspension in a vessel having a three-dimensional geometry approximating the in vivo gross morphology of a tissue of interest, said vessel having attachment surfaces thereon; and

allowing the suspension to coalesce; and

culturing said coalesced suspension under conditions in which said cells connect to said attachment surfaces and form an organized tissue having an in vivo-like gross and cellular morphology of said tissue of interest, wherein the organized tissue is further comprised of post-mitotic cells and wherein at least a subset of the cells said organized tissue comprise said DNA sequence.

19. The method of claim 18, wherein the step of providing comprises isolating primary cells of at least one of the cell types comprising said tissue of interest.

20. The method of claim 18, wherein the step of providing comprises utilizing immortalized cells of at least one of the cell types comprising said tissue.

21. The method of claim 18, wherein prior to the step of providing, a foreign DNA sequence operably linked to a promoter and encoding a protein is introduced to at least a subset of said cells.

22. The method of claim 18, wherein said cells comprise skeletal muscle cells.

23. The method of claim 18, wherein said coalesced suspension exerts a force on said cells parallel to a dimension of said vessel.

24. The method of claim 18, wherein said cells are aligned parallel to a dimension of said vessel.

25. The method of claim 24, wherein said vessel is substantially semi-cylindrical in shape.

26. The method of claim 25, wherein said attachment surfaces are positioned at opposite ends of said vessel.

27. The method of claim 18, wherein said organized tissue produces said protein.

29. An organized tissue producing a protein of a type or produced in an amount not produced normally by a tissue of interest, where the organized tissue is produced by the steps of:

mixing a plurality of cells with an extracellular matrix to create a suspension, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein;

placing said suspension in a vessel having a three-dimensional geometry approximating the in vivo gross morphology of said tissue of interest, said vessel having attachment surfaces thereon; and

allowing the suspension to coalesce; and

culturing said coalesced suspension under conditions in which said cells connect to said attachment surfaces and form an organized tissue having an in vivo-like gross and cellular morphology of said tissue of interest, and wherein the organized tissue is further comprised of post-mitotic cells, and;

wherein the protein is produced at detectable levels in said tissue.

30. An organized tissue, wherein the organized tissue has an in vivo-like gross and cellular morphology of a tissue of interest and produces a protein of a type or produced in an amount not produced normally by said tissue of interest comprising:

a plurality of cells, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, wherein said cells form an organized tissue approximating the in vivo gross morphology of said tissue of interest and wherein the organized tissue is further comprised of post-mitotic cells, and;

wherein the protein is produced at detectable levels in said tissue.

31. The organized tissue of claim 30, wherein said tissue is skeletal muscle.

32. An organized tissue producing a protein produced by the steps of:

mixing a plurality of mammalian cells, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, wherein said cells are mixed with an extracellular matrix to create a suspension;

placing said suspension in a vessel having a three-dimensional geometry approximating the in vivo gross morphology of a tissue of interest, said vessel having attachment surfaces thereon; and

allowing the suspension to coalesce; and

culturing said coalesced suspension under conditions in which said cells connect to said attachment surfaces, wherein said suspension of cells forms an organized tissue that has a three-dimensional structure that is retained upon implantation of said tissue into a mammal, and wherein the tissue is further comprised of post-mitotic cells, and wherein the protein is produced sufficiently to provide a therapeutic effect to said mammal once the organized tissue is implanted into said mammal.

33. An organized tissue having a three-dimensional cellular organization of a tissue of interest that is retained upon implantation of said tissue into a mammal, said tissue producing a protein of a type or in an amount not normally produced by a tissue of interest, comprising:

a plurality of cells, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, wherein said tissue has a three-dimensional geometry approximating the in vivo gross morphology of said tissue of interest, and wherein the organized tissue is further comprised of post-mitotic cells, and;

wherein the protein is produced to detectable levels in said tissue.

34. An organized tissue attached to a surface of a substrate, said tissue producing a protein, comprising:

a plurality of cells, wherein at least a subset of said cells comprise a foreign DNA sequence operably linked to a promoter and encoding a protein, wherein said cells form an organized tissue having a three-dimensional geometry approximating the in vivo gross morphology of a tissue of interest, and wherein said organized tissue is attached to the surface of a substrate, and wherein the organized tissue is further comprised of post-mitotic cells, and;

wherein the protein is produced to detectable levels in said tissue.

35. The organized tissue of claim 34, said substrate being selected from the group consisting of metal or plastic.

36. The organized tissue of claim 35, said metal substrate being steel mesh having a longitudinal axis and first and second points for attachment, and wherein said first and second attachment sites of said tissue are attached, respectively, to said first and second points of attachment.