USES AND ISOLATION OF VERY SMALL EMBRYONIC-LIKE (VSEL) STEM CELLS

• US 20110189136A1
• Filed: 10/30/2008
• Published: 08/04/2011
• Est. Priority Date: 12/08/2005
• Status: Active Grant

• Current Assignee: University of Louisville
• Original Assignee: University of Louisville
• Inventors: Smith, George, Kucia, Magdalena, Ratajczak, Mariusz, Ratajczak, Janina, Allen, Ronald, Marasco, Wayne, Bolli, Roberto
• Granted Patent: US 9,155,762 B2
• Time in Patent Office: Days
• US Class Current: 424/93.7
• CPC Class Codes: A61B 10/00 Other methods or instruments for diagnosis, e.g. instruments for taking a cell sample, for biopsy, for vaccination diagnosis vaccination prophylaxis, vaccination therapy A61B17/20; Sex determination; Ovulation-period determination menstruation tables G06C3/00; Throat striking implements
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• 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: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: A61K 35/34 Muscles; Smooth muscle cells; Heart; Cardiac stem cells; Myoblasts; Myocytes; Cardiomyocytes vascular smooth muscle A61K35/44
• CPC Class Codes: A61K 35/545 Embryonic stem cells; Pluripotent stem cells; Induced pluripotent stem cells; Uncharacterised stem cells
• CPC Class Codes: A61K 38/18 Growth factors; Growth regulators
• CPC Class Codes: C12N 5/0607 Non-embryonic pluripotent stem cells, e.g. MASC induced pluripotent stem cells C12N5/0696

Abstract

The presently disclosed subject matter provides populations of stem cells that are purified from bone marrow, peripheral blood, and/or other sources. Also provided are methods of using the stem cells for treating tissue and/or organ damage in a subject.

Claims

1. A method of collecting autologous VSELs from a subject comprising the steps of:

administering to the subject a stem cell potentiating agent;

collecting at least 10¹⁰ total nucleated cells from the peripheral blood of the subject;

enriching the total nucleated cells for CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs);

preserving the collected cells to maintain the cellular integrity of the cells.

2. The method of claim 1 comprising collecting at least 10¹¹ total nucleated cells from the peripheral blood of the subject.

3. The method of claim 1 comprising collecting at least 10¹² total nucleated cells from the peripheral blood of the subject.

4. The method of claim 1 wherein the enrichment process involves sorting the total nucleated cells based on their cellular marker expression pattern, cellular size, or combination thereof.

5. The method of claim 1 wherein the enrichment process uses a multiparameter process.

6. The method of claim 1 wherein the stem cell potentiating agent is G-CSF, GM-CSF, dexamethazone, a CXCR4 receptors inhibitor, or combination thereof.

7. The method of claim 6 wherein the stem cell potentiating agent is G-CSF.

8. The method of claim 7 comprising administering to the subject at least two doses of G-CSF of about 1 μ

g/kg/day to 8 μ

g/kg/day;

9. The method of claim 7, wherein the G-CSF is administered subcutaneously.

10. The method of claim 7, wherein about 480 μ

g per dose of G-CSF is administered subcutaneously to the subject.

11. The method of claim 8, wherein the at least two doses of G-CSF are administered on two consecutive days, with the subject receiving one dose per day.

12. The method of claim 8, wherein the subject is administered the at least two doses of G-CSF within a 2 to 6 day period.

13. The method of claim 8, wherein the subject receives the two doses of G-CSF administered on consecutive days.

14. The method of claim 8, wherein the subject is administered the at least two doses of G-CSF within about 12 to about 36 hours of each other.

15. The method of claim 1, further comprising earmarking the collected cells for use by the subject at the time of collection.

16. The method of claim 6, wherein the G-CSF is administered to a subject at a dose from about 4 to about 6 μ

g/kg/day.

17. The method of claim 6, wherein from about 50 μ

g to about 800 μ

g per dose of G-CSF is administered subcutaneously to the subject.

18. The method of claim 6, wherein from about 300 μ

g to about 500 μ

g per dose of G-CSF is administered subcutaneously to the subject.

19. The method of claim 1, wherein an apheresis process is used to collect the at least 10¹⁰ total nucleated cells from the peripheral blood of the subject.

20. The method of claim 19, wherein the apheresis process is conducted the day after the second dose of G-CSF is administered.

21. The method of claim 19, wherein the apheresis process is conducted about 12 to about 36 hours after the second dose of G-CSF is administered.

22. The method of claim 1, wherein the subject is a human subject that has met at least one condition selected from the group consisting of being between 10 and 200 kg in weight and between 2 to 80 years old.

23. The method of claim 22, wherein the collecting step is conducted when the subject is an adult or a non-neonate.

24. A cellular therapy product comprising an autologous mixture of CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs) and non-VSELs, wherein the non-VSELs comprise progenitor cells and optionally functional cells.

25. The cellular therapy product of claim 24 comprising from about 10% to about 90% peripheral blood VSELs and from about 10% to about 90% non-VSELs.

26. The cellular therapy product of claim 24 comprising from about 10% to about 80% peripheral blood VSELs and from about 20% to about 90% non-VSELs.

27. The cellular therapy product of claim 24 comprising from about 10% to about 60% peripheral blood VSELs and from about 40% to about 90% non-VSELs.

28. The cellular therapy product of claim 24, wherein the non-VSELs are selected from the group consisting of hematopoietic progenitor cells, neural progenitor cells, glial progenitor cells, oligodendrocyte progenitor cells, skin progenitor cells, hepatic progenitor cells, muscle progenitor cells, bone progenitor cells, mesenchymal stem or progenitor cells, pancreatic progenitor cells, progenitor chondrocytes, stromal progenitor cells, cultured expanded stem or progenitor cells, cultured differentiated stem or progenitor cells, or combinations thereof.

29. The cellular therapy product of claim 24, wherein the functional cells are selected from the group consisting of terminally differentiated hematopoietic cells, terminally differentiated neural cells, terminally differentiated glial cells, terminally differentiated oligodendrocytes, terminally differentiated skin cells, terminally differentiated hepatic cells, terminally differentiated muscle cells, terminally differentiated bone cells, terminally differentiated adipocytes, terminally differentiated pancreatic cells, chondrocytes, stromal cells, cultured differentiated stem or progenitor cells, or combinations thereof.

30. A method of enhancing the engraftment of stem cells comprising administering to a subject an autologous mixture of CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs), progenitor cells, and optionally functional cells.

31. A cellular therapy product comprising at least 10⁴ CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs).

32. The cellular therapy product of claim 31 comprising at least 10⁵ CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

VSELs.

33. The cellular therapy product of claim 31 comprising at least 10⁶ CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

VSELs.

34. A method of enhancing the engraftment of stem cells comprising administering to a subject the cellular therapy product of claim 31.

35. A process of making stem cells and progenitor cells available to a person, comprising the steps of:

the person proactively electing to have his CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs) collected, wherein the person has no immediate perceived health condition requiring immediate treatment using his own collected stem cells and progenitor cells;

collecting at least 10¹⁰ total nucleated cells from the peripheral blood of the person using an apheresis process;

at the time of collection, earmarking the collected cells for use by the person; and

preserving the collected cells in storage.

36. A process for collecting and banking CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs) of a person, comprising the steps of collecting at least 10¹⁰ total nucleated cells from peripheral blood of a person with no immediate perceived health condition requiring immediate treatment using an apheresis process;

at the time of collection, earmarking the collected VSELs for use by the person; and

preserving the collected cells by storage in a cell bank.

37. A method of treating an injury to a tissue in a subject, the method comprising administering to the subject a composition comprising CD34⁺/lin⁻

/CD45⁻

or Sca-1⁺/lin⁻

/CD45⁻

very small embryonic-like stem cells (VSELs) in a pharmaceutically acceptable carrier, in an amount and via a route sufficient to allow at least a fraction of the population of VSELs to engraft the tissue and differentiate therein, wherein the injury is selected from the group consisting of an ischemic injury, a myocardial infarction, and stroke.

38. The method of claim 37, wherein the isolated VSELs are isolated from a source selected from the group consisting of bone marrow, peripheral blood, spleen, cord blood, and combinations thereof.