Irreversible electroporation to create tissue scaffolds

• US 10,286,108 B2
• Filed: 02/03/2017
• Issued: 05/14/2019
• Est. Priority Date: 04/29/2008
• Status: Active Grant

• Current Assignee: Virginia Tech Intellectual Properties, Inc. (Virginia Polytechnic Institute and State University)
• Original Assignee: Virginia Tech Intellectual Properties, Inc. (Virginia Polytechnic Institute and State University)
• Inventors: Davalos, Rafael V.
• Primary Examiner(s): Kim, Taeyoon
• Application Number:
  US15/423,986
• Publication Number:
  US 20170209620A1
• Time in Patent Office: 830 Days
• CPC Class Codes: A61K 35/12 Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells vaccines or medicinal preparations containing antigens or antibodies A61K39/00
• CPC Class Codes: A61L 2430/28 for liver reconstruction
• CPC Class Codes: A61L 2430/40 Preparation and treatment of biological tissue for implantation, e.g. decellularisation, cross-linking
• CPC Class Codes: A61L 27/3604 characterised by the human or animal origin of the biological material, e.g. hair, fascia, fish scales, silk, shellac, pericardium, pleura, renal tissue, amniotic membrane, parenchymal tissue, fetal tissue, muscle tissue, fat tissue, enamel
• CPC Class Codes: A61L 27/3625 Vascular tissue, e.g. heart valves
• CPC Class Codes: A61L 27/3633 Extracellular matrix [ECM]
• CPC Class Codes: A61L 27/3691 characterised by physical conditions of the treatment, e.g. applying a compressive force to the composition, pressure cycles, ultrasonic/sonication or microwave treatment, lyophilisation
• CPC Class Codes: A61L 27/38 containing added animal cells organs or tissue containing native cells A61L27/36
• CPC Class Codes: C12N 13/00 Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
• CPC Class Codes: C12N 2533/90 Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0068 General culture methods using substrates for specific animal cell type C12N5/06

Abstract

The present invention provides engineered tissue scaffolds, engineered tissues, and methods of using them. The scaffolds and tissues are derived from natural tissues and are created using non-thermal irreversible electroporation (IRE). Use of IRE allows for ablation of cells of the tissue to be treated, but allows vascular and neural structures to remain essentially unharmed. Use of IRE thus permits preparation of thick tissue scaffolds and tissues due to the presence of vasculature within the scaffolds. The engineered tissues can be used in methods of treating subjects, such as those in need of tissue replacement or augmentation.

Claims

1. An in vitro method of making an engineered tissue scaffold, said method comprising:

engineering a tissue scaffold by treating, in vitro, a tissue comprising cells and a native extracellular matrix with an electrical field;

applying the electrical field in a manner and for a sufficient time at a sufficient power to cause irreversible electroporation of the cells; and

producing an engineered tissue scaffold as a result of the irreversible electroporation.

2. The method of claim 1, wherein treating comprises placing a first electrode and a second electrode in contact with the tissue such that the portion of tissue to be decellularized by irreversible electroporation is positioned between the first and second electrodes.

3. The method of claim 2, wherein more than two electrodes are placed in contact with the tissue.

4. The method of claim 2, wherein treating with an electrical field comprises applying more than one electrical pulse between the first and second electrodes in an amount sufficient to induce irreversible electroporation of cells of the tissue.

5. The method of claim 1, further comprising removing cellular debris remaining from the irreversible electroporation.

6. The method of claim 5, wherein the cellular debris is removed using one or more physical, chemical, or enzymatic techniques.

7. The method of claim 1, wherein the irreversible electroporation does not cause substantial destruction of the extracellular matrix due to heat.

8. The method of claim 1, wherein the engineered tissue scaffold comprises a mixture of structural and functional proteins.

9. The method of claim 8, wherein the engineered tissue scaffold retains the three-dimensional architecture of the native extracellular matrix.

10. The method of claim 8, wherein the engineered tissue scaffold retains the mechanical properties of the native extracellular matrix.

11. The method of claim 1, wherein the engineered tissue scaffold has a thickness that approaches or mimics the thickness of the tissue before treatment.

12. The method of claim 1, wherein the engineered tissue scaffold has a thickness of about 5 mm or more.

13. The method of claim 1, wherein the tissue is an organ.

14. The method of claim 13, wherein the organ is a heart, a lung, a liver, a kidney, a urinary bladder, a brain, an ear, an eye, or skin.

15. The method of claim 1, wherein the tissue further comprises one or more vascular structures, and the irreversible electroporation does not cause irreversible damage to at least some of the one or more vascular structures.

16. The method of claim 15, wherein at least one of the one or more vascular structures is an artery, vein, or capillary which is left intact in the engineered tissue scaffold after the irreversible electroporation.

17. The method of claim 15, wherein at least one of the one or more vascular structures is left intact in the engineered tissue scaffold after the irreversible electroporation and is capable of conducting microcirculation and/or has a cross-sectional diameter of 5 to 10 micrometers.

18. The method of claim 15, wherein at least one of the one or more vascular structures is left intact in the engineered tissue scaffold after the irreversible electroporation and comprises a network of arteries, veins, and/or capillaries.

19. The method of claim 1, wherein the tissue further comprises one or more neural structures, and the irreversible electroporation does not cause irreversible damage to at least some of the one or more neural structures.

20. The method of claim 19, wherein at least one of the one or more neural structures is a neural tube which is left intact in the engineered tissue scaffold after the irreversible electroporation.