Treatment of graft-versus-host disease

• US 9,439,929 B2
• Filed: 05/27/2008
• Issued: 09/13/2016
• Est. Priority Date: 06/17/2005
• Status: Active Grant

• Current Assignee: SCM Lifescience Co., Ltd.
• Original Assignee: INHA Industry Partnership Institute
• Inventors: Song, Sun Uk, Lee, Moon Hee, Kim, Chul Soo
• Primary Examiner(s): DRISCOLL, LORA E BARNHART
• Application Number:
  US12/127,734
• Publication Number:
  US 20080292600A1
• Time in Patent Office: 3,031 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: A61K 2035/122 for inducing tolerance or supression of immune responses
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• CPC Class Codes: A61K 2239/31 characterized by the route of administration
• CPC Class Codes: A61K 35/17 Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes when activated by a specific antigen A61K39/00
• CPC Class Codes: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: A61K 38/2066 IL-10
• CPC Class Codes: A61K 39/461 characterised by the cell type used
• CPC Class Codes: A61K 39/4621 immunosuppressive or immunotolerising
• CPC Class Codes: A61K 39/46434 Antigens related to induction of tolerance to non-self
• CPC Class Codes: A61K 39/4644 Cancer antigens
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: C12N 5/0663 Bone marrow mesenchymal stem cells (BM-MSC)

Abstract

This present application describes a therapeutic agent for treating acute or chronic graft-versus-host disease using clonal marrow stem cells (cMSCs) as active ingredient.

Claims

1. A method of treating symptoms of graft-versus-host disease in a subject identified as suffering from graft-versus-host disease, comprising:

(A) obtaining a homogeneous population of single cell-derived clonal multipotent bone marrow cells based on cell density from a biological sample of bone marrow, comprising;

(i) allowing the biological sample to settle by gravity in a first container producing a first supernatant of lower density cells;

(ii) transferring the first supernatant directly without undergoing centrifugation to a second container of growth medium and allowing cells to settle to the bottom producing a second supernatant of lower density cells;

(iii) transferring the second supernatant directly without undergoing centrifugation to a third container of growth medium and allowing cells to settle to the bottom, producing a third supernatant of lower density cells;

(iv) transferring the third supernatant directly without undergoing centrifugation to another container of growth medium and allowing cells to settle to the bottom, producing another supernatant of lower density cells;

(v) allowing single-cell derived colonies to appear on the bottom of the container of step (iv);

(vi) isolating the single-cell derived colonies; and

(vii) expanding cells from the single-cell derived colonies in a further other container of growth medium to obtain the homogeneous population of single cell-derived clonal multipotent bone marrow cells, and

(B) administering to the subject in need thereof a therapeutically effective amount of the homogeneous population of single cell-derived clonal multipotent bone marrow cells obtained in (A) to inhibit T-cell activity from a donor marrow.

2. The method according to claim 1, wherein the cells are settled for one day in step (iv).

3. The method according to claim 1, wherein the isolated cells from the supernatant are expanded in a container.

4. A method of treating symptoms of graft-versus-host disease in a subject identified as suffering from graft-versus-host disease, comprising:

(A) obtaining a homogeneous population of single cell-derived clonal multipotent bone marrow cells based on cell density from a biological sample of bone marrow, comprising;

(i) allowing the biological sample to settle by gravity in a first container producing a first supernatant of lower density cells;

(ii) transferring the first supernatant directly without undergoing centrifugation to a second container of growth medium and allowing cells to settle to the bottom producing a second supernatant of lower density cells;

(iii) transferring the second supernatant directly without undergoing centrifugation to a third container of growth medium and allowing cells to settle to the bottom, producing a third supernatant of lower density cells;

(iv) transferring the third supernatant directly without undergoing centrifugation to a fourth container of growth medium and allowing cells to settle to the bottom, producing a fourth supernatant of lower density cells;

(v) transferring the fourth supernatant directly without undergoing centrifugation to another container of growth medium and allowing cells to settle to the bottom, producing another supernatant of lower density cells;

(vi) allowing single-cell derived colonies to appear on the bottom of the container of step (v);

(vii) isolating the single-cell derived colonies; and

(viii) expanding cells from the single-cell derived colonies in a further other container of growth medium to obtain the homogeneous population of single cell-derived clonal multipotent bone marrow cells, and

(B) administering to the subject in need thereof a therapeutically effective amount of the homogeneous population of single cell-derived clonal multipotent bone marrow cells obtained in (A) to inhibit T-cell activity from a donor marrow.

5. The method according to claim 4, wherein the cells are settled for one day in step (v).

6. The method according to claim 4, wherein the isolated cells from the supernatant are expanded in a container.

7. A method of treating symptoms of graft-versus-host disease in a subject identified as suffering from graft-versus-host disease, comprising:

(A) obtaining a homogeneous population of single cell-derived clonal multipotent bone marrow cells based on cell density from a biological sample of bone marrow, comprising;

(i) allowing the biological sample to settle by gravity in a first container producing a first supernatant of lower density cells;

(ii) transferring the first supernatant directly without undergoing centrifugation to a second container of growth medium and allowing cells to settle to the bottom producing a second supernatant of lower density cells;

(iii) transferring the second supernatant directly without undergoing centrifugation to a third container of growth medium and allowing cells to settle to the bottom, producing a third supernatant of lower density cells;

(iv) transferring the third supernatant directly without undergoing centrifugation to a fourth container of growth medium and allowing cells to settle to the bottom, producing a fourth supernatant of lower density cells;

(v) transferring the fourth supernatant directly without undergoing centrifugation to a fifth container of growth medium and allowing cells to settle to the bottom, producing a fifth supernatant of lower density cells;

(vi) transferring the fifth supernatant directly without undergoing centrifugation to another container of growth medium and allowing cells to settle to the bottom, producing another supernatant of lower density cells;

(vii) allowing single-cell derived colonies to appear on the bottom of the container of step (vi);

(viii) isolating the single-cell derived colonies; and

(ix) expanding cells from the single-cell derived colonies in a further other container of growth medium to obtain the homogeneous population of single cell-derived clonal multipotent bone marrow cells, and

(B) administering to the subject in need thereof a therapeutically effective amount of the homogeneous population of single cell-derived clonal multipotent bone marrow cells obtained in (A) to inhibit T-cell activity from a donor marrow.

8. The method according to claim 7, wherein the cells are settled for one day in step (vi).

9. The method according to claim 7, wherein the isolated cells from the supernatant are expanded in a container.

10. A method of treating graft-versus-host disease in a subject identified as suffering from graft-versus-host disease, comprising:

(A) obtaining a homogeneous population of single cell-derived clonal multipotent bone marrow cells based on cell density from a biological sample of bone marrow, comprising;

(i) allowing the biological sample to settle by gravity in a first container producing a first supernatant of lower density cells;

(ii) transferring the first supernatant directly without undergoing centrifugation to a second container of growth medium and allowing cells to settle to the bottom producing a second supernatant of lower density cells;

(iii) transferring the second supernatant directly without undergoing centrifugation to a third container of growth medium and allowing cells to settle to the bottom, producing a third supernatant of lower density cells;

(iv) transferring the third supernatant directly without undergoing centrifugation to another container of growth medium and allowing cells to settle to the bottom, producing another supernatant of lower density cells;

(v) allowing single-cell derived colonies to appear on the bottom of the container of step (iv);

(vi) isolating the single-cell derived colonies; and

(vii) expanding cells from the single-cell derived colonies in a further other container of growth medium to obtain the homogeneous population of single cell-derived clonal multipotent bone marrow cells, and

(B) administering to the subject in need thereof a therapeutically effective amount of the homogeneous population of single cell-derived clonal multipotent bone marrow cells obtained in (A) to inhibit T-cell activity from a donor marrow.

11. The method according to claim 10, wherein the cells are settled for one day in step (iv).

12. The method according to claim 10, wherein the isolated cells from the supernatant are expanded in a container.

13. A method of treating graft-versus-host disease in a subject identified as suffering from graft-versus-host disease, comprising:

(A) obtaining a homogeneous population of single cell-derived clonal multipotent bone marrow cells based on cell density from a biological sample of bone marrow, comprising;

(i) allowing the biological sample to settle by gravity in a first container producing a first supernatant of lower density cells;

(ii) transferring the first supernatant directly without undergoing centrifugation to a second container of growth medium and allowing cells to settle to the bottom producing a second supernatant of lower density cells;

(iii) transferring the second supernatant directly without undergoing centrifugation to a third container of growth medium and allowing cells to settle to the bottom, producing a third supernatant of lower density cells;

(iv) transferring the third supernatant directly without undergoing centrifugation to a fourth container of growth medium and allowing cells to settle to the bottom, producing a fourth supernatant of lower density cells;

(v) transferring the fourth supernatant directly without undergoing centrifugation to another container of growth medium and allowing cells to settle to the bottom, producing another supernatant of lower density cells;

(vi) allowing single-cell derived colonies to appear on the bottom of the container of step (v);

(vii) isolating the single-cell derived colonies; and

(viii) expanding cells from the single-cell derived colonies in a further other container of growth medium to obtain the homogeneous population of single cell-derived clonal multipotent bone marrow cells, and

(B) administering to the subject in need thereof a therapeutically effective amount of the homogeneous population of single cell-derived clonal multipotent bone marrow cells obtained in (A) to inhibit T-cell activity from a donor marrow.

14. The method according to claim 13, wherein the cells are settled for one day in step (v).

15. The method according to claim 13, wherein the isolated cells from the supernatant are expanded in a container.

16. A method of treating graft-versus-host disease in a subject identified as suffering from graft-versus-host disease, comprising:

(A) obtaining a homogeneous population of single cell-derived clonal multipotent bone marrow cells based on cell density from a biological sample of bone marrow, comprising;

(i) allowing the biological sample to settle by gravity in a first container producing a first supernatant of lower density cells;

(ii) transferring the first supernatant directly without undergoing centrifugation to a second container of growth medium and allowing cells to settle to the bottom producing a second supernatant of lower density cells;

(iii) transferring the second supernatant directly without undergoing centrifugation to a third container of growth medium and allowing cells to settle to the bottom, producing a third supernatant of lower density cells;

(iv) transferring the third supernatant directly without undergoing centrifugation to a fourth container of growth medium and allowing cells to settle to the bottom, producing a fourth supernatant of lower density cells;

(v) transferring the fourth supernatant directly without undergoing centrifugation to a fifth container of growth medium and allowing cells to settle to the bottom, producing a fifth supernatant of lower density cells;

(vi) transferring the fifth supernatant directly without undergoing centrifugation to another container of growth medium and allowing cells to settle to the bottom, producing another supernatant of lower density cells;

(vii) allowing single-cell derived colonies to appear on the bottom of the container of step (vi);

(viii) isolating the single-cell derived colonies; and

(ix) expanding cells from the single-cell derived colonies in a further other container of growth medium to obtain the homogeneous population of single cell-derived clonal multipotent bone marrow cells, and

(B) administering to the subject in need thereof a therapeutically effective amount of the homogeneous population of single cell-derived clonal multipotent bone marrow cells obtained in (A) to inhibit T-cell activity from a donor marrow.

17. The method according to claim 16, wherein the cells are settled for one day in step (vi).

18. The method according to claim 16, wherein the isolated cells from the supernatant are expanded in a container.