# US 10,765,776 B1 · Tissue-derived biomaterial composition and methods for ocular and other therapeutic applications

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- Number: US 10,765,776 B1
- Document ID: US10765776B1
- Status: Granted
- Kind: B1
- Application number: 15/791,720
- Filed: 2017-10-24
- Granted: 2020-09-08
- Earliest claimed: 2017-10-24
- Assignees: South Dakota Board of Regents (Sdbor)
- Applicants: South Dakota Board of Regents (SDBOR)
- Inventors: Gudiseva Chandrasekher; Somshuvra Bhattacharya
- Web page: https://patentgrep.com/patent/US10765776B1

## CPC

- 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}
- A61K 35/30: Nerves; Brain; Eyes; Corneal cells; Cerebrospinal fluid; Neuronal stem cells; Neuronal precursor cells; Glial cells; Oligodendrocytes; Schwann cells; Astroglia; Astrocytes; Choroid plexus; Spinal cord tissue
- A61L 27/3666: {Epithelial tissues other than skin}
- A61L 27/3687: {characterised by the use of chemical agents in the treatment, e.g. specific enzymes, detergents, capping agents, crosslinkers, anticalcification agents}
- 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}
- A61L 27/3808: {Endothelial cells}
- A61L 27/3813: {Epithelial cells, e.g. keratinocytes, urothelial cells}
- A61L 27/3869: {Epithelial tissues other than skin}
- A61L 27/3886: {comprising two or more cell types}
- A61L 27/52: Hydrogels or hydrocolloids
- C12N 5/0621: {Eye cells, e.g. cornea, iris pigmented cells (photoreceptors C12N5/062)}
- A61L 2400/06: Flowable or injectable implant compositions
- A61L 2430/16: for reconstruction of eye parts, e.g. intraocular lens, cornea
- C12N 2513/00: 3D culture
- C12N 2533/70: Polysaccharides
- C12N 2533/90: Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
- A61L: METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES (preservation of bodies or disinfecting characterised by the agents employed A01N; preserving, e.g. sterilising, food or foodstuffs A23; preparations for medical, dental or toiletry purposes A61K)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

The present disclosure relates to a composition and methods for preparing a tissue-derived, dissolved extracellular matrix. In implementations, the extracellular matrix composition includes components native to the cornea including liquefied corneal tissue. The extracellular matrix composition can be prepared by dissolving extracted corneas in an aqueous solution of inorganic base. The resulting solution is then dialyzed to achieve a physiological pH. In some implementations, the extracellular matrix composition can be seeded and/or chemically cross-linked to form a gel matrix. The extracellular matrix and/or the gel matrix can be seeded with a desired cell type (e.g., endothelial cells, epithelial cells, stromal keratocytes, etc.) to stimulate cell proliferation and regeneration.

## Claims

### Claim 1

A method of preparing an extracellular matrix composition, comprising:
extracting one or more corneas from one or more donor sources;
dissolving the one or more corneas in an aqueous solution of inorganic base to form a dissolved cornea solution;
dialyzing the dissolved cornea solution against at least one of a buffer solution or a serum-free medium to remove the inorganic base and adjust the pH of the dissolved cornea to physiological pH;
centrifuging the dissolved cornea solution; and
sterilizing the dissolved cornea solution.

### Claim 2 (depends on 1)

The method of preparing an extracellular matrix composition of claim 1, wherein the aqueous solution of inorganic base comprises a sodium hydroxide solution.

### Claim 3 (depends on 1)

The method of preparing an extracellular matrix composition of claim 1, further comprising seeding the dissolved cornea solution into a pectin-calcium chloride gel to form a gel matrix.

### Claim 4 (depends on 1)

The method of preparing an extracellular matrix composition of claim 1, further including seeding the dissolved cornea solution with cultured tissue cells to form a suspension.

### Claim 5 (depends on 4)

The method of preparing an extracellular matrix composition of claim 4, wherein the cultured tissue cells comprise at least one of epithelial cells, endothelial cells, or stromal keratocytes.

### Claim 6 (depends on 1)

The method of preparing an extracellular matrix composition of claim 1, further comprising cross-linking the dissolved cornea solution to a desired hardness to form a scaffold for sustained regeneration of cells.

### Claim 7 (depends on 6)

The method of preparing an extracellular matrix composition of claim 6, further comprising seeding the scaffold with cultured tissue cells.

### Claim 8

A method of preparing an extracellular matrix scaffold, comprising:
extracting one or more corneas from one or more donor sources;
dissolving the one or more corneas in an aqueous solution of an inorganic base to form a dissolved cornea solution;
dialyzing the dissolved cornea solution against at least one of a buffer solution or a serum-free medium to remove the inorganic base and adjust the pH of the dissolved cornea solution to physiological pH;
centrifuging the dissolved cornea solution;
sterilizing the dissolved cornea solution; and
forming a gel matrix from the dissolved cornea solution.

### Claim 9 (depends on 8)

The method of preparing an extracellular matrix scaffold of claim 8, wherein dialyzing the dissolved cornea solution against at least one of a buffer solution or a serum-free medium to remove the inorganic base and adjust the pH of the dissolved cornea to physiological pH includes dialyzing the dissolved cornea solution against at least DMEM/F12 medium to remove the inorganic base and adjust the pH of the dissolved cornea to physiological pH.

### Claim 10 (depends on 9)

The method of preparing an extracellular matrix scaffold of claim 9, wherein the gel matrix is formed by cross-linking the dissolved cornea solution to a desired hardness.

### Claim 11 (depends on 9)

The method of preparing an extracellular matrix scaffold of claim 9, further comprising seeding the scaffold with cultured tissue cells.

### Claim 12 (depends on 11)

The method of preparing an extracellular matrix scaffold of claim 11, wherein the cultured tissue cells comprise at least one of epithelial cells, endothelial cells, or stromal cells.

### Claim 13 (depends on 11)

The method of preparing an extracellular matrix scaffold of claim 11, wherein the cultured tissue cells comprise cultured corneal cells.

### Claim 14 (depends on 8)

The method of preparing an extracellular matrix scaffold of claim 8, wherein sterilizing the dissolved cornea solution comprises exposing the dissolved cornea solution to ultraviolet light.