# US 10,946,043 B2 · Methods of mediating macrophage phenotypes

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- Number: US 10,946,043 B2
- Document ID: US10946043B2
- Status: Granted
- Kind: B2
- Application number: 15/797,796
- Filed: 2017-10-30
- Published: 2018-02-22
- Granted: 2021-03-16
- Earliest claimed: 2013-11-26
- Assignees: Biomet Biologics, LLC
- Applicants: Biomet Biologies, LLC
- Inventors: Jennifer E. Woodell-May; Joel C. Higgins; William King
- Web page: https://patentgrep.com/patent/US10946043B2

## CPC

- C12N 5/0645: {Macrophages, e.g. Kuepfer cells in the liver; Monocytes}
- A61K 40/17: Monocytes; Macrophages
- A61K 40/22: Immunosuppressive or immunotolerising
- A61K 40/24: Antigen-presenting cells [APC]
- A61K 40/416: {Antigens related to auto-immune diseases; Preparations to induce self-tolerance}
- C12N 5/0667: {Adipose-derived stem cells [ADSC]; Adipose stromal stem cells}
- A61K 35/16: Blood plasma; Blood serum (umbilical cord blood A61K35/51)
- C12N 2501/20: Cytokines; Chemokines
- C12N 2502/1382: Adipose-derived stem cells [ADSC], adipose stromal stem cells
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- 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)

## Abstract

Methods of inducing a polarization of macrophages. The method includes obtaining a blood fraction, fractionating the blood fraction to produce a blood fraction, and contacting the blood fraction with a source of macrophages. A blood fraction including platelet-poor plasma polarizes the source of macrophages into M1 macrophages. A blood faction including a protein solution polarizes the source of macrophages into M2 macrophages.

## Claims

### Claim 1

A method for treating inflammation in a subject, comprising:
administering to the subject at a site of inflammation a composition comprising between about 1 pg/ml and about 60 pg/ml interleukin-4 (IL-4), wherein monocytes or macrophages at the site of inflammation are induced by the composition to polarize to an M2 phenotype.

### Claim 2 (depends on 1)

The method according to claim 1, composition comprises monocytes, macrophages, or both, from the subject.

### Claim 3 (depends on 1)

The method according to claim 1, wherein the monocytes or macrophages are polarized to an M2 phenotype within 10 days of administering the composition.

### Claim 4 (depends on 1)

The method according to claim 1, wherein the composition further comprises interleukin-10 (IL-10), transforming growth factor-beta (TGF-β), vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), interleukin-1 receptor antagonist (IL-1ra), or a combination thereof.

### Claim 5 (depends on 1)

The method according to claim 1, further comprising obtaining a cytokine cell suspension from a donor; and concentrating the cytokine cell suspension to form the composition.

### Claim 6 (depends on 5)

The method according to claim 5, wherein the cytokine cell suspension comprises whole blood, bone marrow aspirate, adipose tissue, a fraction or a concentrate thereof, or a mixture thereof.

### Claim 7 (depends on 5)

The method according to claim 5, wherein concentrating includes contacting the cytokine cell suspension with polyacrylamide beads.

### Claim 8 (depends on 1)

The method according to claim 1, wherein the composition is prepared from a cytokine cell suspension obtained from the subject.

### Claim 9 (depends on 1)

The method according to claim 1, wherein the composition comprises transforming growth factor-beta (TGF-11), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF), each at a concentration greater than the concentration of the respective protein in normal blood.

### Claim 10 (depends on 1)

The method according to claim 1, wherein the composition comprises interleukin-10 (IL-10) and transforming growth factor-β(TGF-β), each at a concentration greater than the concentration of the respective protein in normal blood.

### Claim 11 (depends on 1)

The method according to claim 1, wherein the composition comprises a protein selected from interleukin-1 receptor antagonist (IL-1ra), soluble tumor necrosis factor-receptor I (sTNF-RI), soluble tumor necrosis factor-receptor II (sTNF-RII), insulin-like growth factor 1 (IGF-I), epidermal growth factor (EGF), hepatocyte growth factor (HGF), platelet-derived growth factor AB (PDGF-AB), platelet-derived growth factor BB (PDGF-BB), vascular endothelial growth factor (VEGF), transforming growth factor-β1 (TGF-β1), soluble interleukin-1 receptor I (sIL-1RI), and soluble interleukin receptor II (sIL-1RII).

### Claim 12 (depends on 1)

The method according to claim 1, wherein the composition induces monocytes or macrophages in the composition to polarize to an M2 phenotype.

### Claim 13

A method for treating inflammation in a subject, comprising:
obtaining a differentiating composition prepared from a tissue and comprising about 1 pg/ml to about 20 pg/ml interleukin-10 (IL-10); and
administering the differentiating composition to a site of inflammation in the subject, wherein the differentiating composition induces monocytes or macrophages at the site of inflammation to differentiate to an M2 phenotype.

### Claim 14 (depends on 13)

The method according to claim 13, wherein the differentiating composition comprises interleukin-4 (IL-4), transforming growth factor-β (TGF-β), or interleukin-1 receptor antagonist, at a concentration greater than the concentration of the protein in normal blood.

### Claim 15 (depends on 13)

The method according to claim 13, wherein the differentiating composition is a concentrated plasma and cytokine cell suspension.

### Claim 16 (depends on 15)

The method according to claim 15, wherein the differentiating composition is prepared from whole blood, bone marrow aspirate, adipose tissue, urine, or a fraction, a mixture, or a concentrate of any one or more of the foregoing.

### Claim 17

A method of treating inflammation in a subject, comprising:
administering to a site of inflammation in the subject a protein solution prepared from tissue of the subject, the protein solution comprising about 1 pg/ml to about 60 pg/ml interleukin-4 (IL-4) and about 1 pg/ml to about 20 pg/ml interleukin-10 (IL-10) wherein the protein solution induces macrophages or monocytes at the site of inflammation to polarize to an M2 phenotype.

### Claim 18 (depends on 17)

The method according to claim 17, wherein the protein solution comprises transforming growth factor-β (TGF-β), vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), or a combination thereof.

### Claim 19 (depends on 18)

The method according to claim 18, wherein the concentration of TGF-β, VEGF, or HGF in the protein solution is greater than the concentration of the respective protein in normal blood.

### Claim 20 (depends on 17)

The method according to claim 17, wherein the protein solution further comprises monocytes isolated from the subject.