Methods of Inhibiting Tumor Cell Aggressiveness Using The Microenvironment of Human Embryonic Stem Cells

• US 20080050362A1
• Filed: 07/26/2007
• Published: 02/28/2008
• Est. Priority Date: 07/28/2006
• Status: Active Grant

• Current Assignee: Children's Memorial Hospital
• Original Assignee: Children's Memorial Hospital
• Inventors: Seftor, Elisabeth, Hendrix, Mary, Seftor, Richard, Postovit, Lynne-Marie
• Granted Patent: US 7,666,423 B2
• Time in Patent Office: Days
• US Class Current: 424/130.1
• CPC Class Codes: A61K 35/54 Ovaries; Ova; Ovules; Embryos; Foetal cells; Germ cells
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 15/00 Drugs for genital or sexual disorders for disorders of sex hormones A61P5/24; Contraceptives
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/495 Transforming growth factor [TGF]
• CPC Class Codes: C12N 15/1136 against growth factors, growth regulators, cytokines, lymphokines or hormones
• CPC Class Codes: C12N 2310/11 Antisense
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/3233 Morpholino-type ring
• CPC Class Codes: G01N 33/5011 for testing antineoplastic activity
• CPC Class Codes: G01N 33/53 Immunoassay; Biospecific binding assay; Materials therefor
• CPC Class Codes: G01N 33/574 for cancer
• CPC Class Codes: G01N 33/57415 of breast
• CPC Class Codes: G01N 33/5743 of skin, e.g. melanoma
• CPC Class Codes: G01N 33/57492 involving compounds localized on the membrane of tumor or cancer cells

Abstract

The invention provides compositions comprising one or more isolated factors from a microenvironment of human embryonic stem cells (hESCs), including, but not limited to, Lefty and inhibitors of Nodal. The invention also provides methods of utilizing factors derived from human embryonic stem cells (hESC) and their microenvironment to treat and prevent tumor formation and progression and to inhibit tumor cell aggressiveness. The invention further provides methods of inhibiting tumor cell growth and/or treating aggressive tumors in a mammal comprising administering to the mammal, having at least one tumor cell present in its body, an effective amount of an inhibitor of Nodal activity.

Claims

1. A composition comprising one or more isolated factors from a microenvironment of human embryonic stem cells.

2. The composition of claim 1 wherein the isolated factor or factors inhibit Nodal.

3. The composition of claim 1 wherein at least one factor is Lefty.

4. An isolated Lefty protein produced by conditioning a matrix with human embryonic stem cells.

5. A protein comprising glycosylated Lefty or a fragment or derivative thereof.

6. The protein of claim 5 wherein the glycosylated Lefty is isolated from the microenvironment of human embryonic stem cells.

7. A composition comprising the protein of claim 5.

8. A method of inhibiting tumor cell growth in a mammal comprising administering to the mammal the composition of claim 7 at a physiologically acceptable dosage.

9. The method of claim 8 wherein the dosage is between 0.1 and 200 ng/mL.

10. A method of using one or more factors from a microenvironment of human embryonic stem cells to inhibit tumor cell aggressiveness.

11. The method of claim 10, wherein at least one of the factors is an inhibitor of Nodal activity.

12. The method of claim 11, wherein the inhibitor of Nodal activity is Lefty.

13. The method of claim 11, wherein the inhibitor of Nodal activity is glycosylated Lefty.

14. The method of claim 11 wherein the inhibitor increases apoptosis and decreases cell proliferation.

15. A method of inhibiting tumor cell growth and/or treating aggressive tumors in a mammal comprising administering to the mammal, having at least one tumor cell present in its body, an effective amount of an inhibitor of Nodal activity.

16. The method of claim 15, wherein the inhibitor blocks binding of Nodal to a receptor.

17. The method of claim 15, wherein the inhibitor is one or more isolated factors from a microenvironment of human embryonic stem cells.

18. The method of claim 15, wherein the inhibitor is glycosylated Lefty.

19. The method of claim 16, wherein the inhibitor is an antibody.

20. The method of claim 15, wherein the inhibitor blocks expression of Nodal protein.

21. The method of claim 15, wherein the inhibitor is an activin-like kinase receptor inhibitor.

22. The method of claim 21, wherein the activin-like kinase receptor inhibitor is an ALK 4/5/7 inhibitor.

23. The method of claim 22 wherein the ALK 4/5/7 inhibitor is SB431542.

24. The method of claim 15 wherein the inhibitor is a Cripto inhibitor.

25. The method of claim 20, wherein the inhibitor is an antisense oligonucleotide.

26. The method of claim 25, wherein the antisense oligonucleotide is an anti-Nodal Morpholino.

27. The method of claim 15, wherein the inhibitor is Lefty.

28. The method of claim 15, wherein the inhibitor is recombinant Lefty.

29. The method of claim 15, wherein the inhibitor is glycosylated Lefty.

30. The method of claim 20, wherein the inhibitor is a Notch inhibitor.

31. The method of claim 30, wherein the Notch inhibitor is a Notch siRNA.

32. The method of claim 30, wherein the inhibitor is a Notch4 inhibitor.

33. The method of claim 32, wherein the inhibitor is a Notch4 siRNA.

34. The method of claim 15, wherein the inhibitor is derived from a microenvironment of human embryonic stem cells.

35. A method of inhibiting tumor cell growth in a mammal comprising administering to the mammal, having at least one tumor cell present in its body, an effective amount of a preconditioned microenvironment, which has been in contact with human embryonic stem cells.

36. A method of detecting aggressive tumor cells comprising the steps of:

a. obtaining a sample from a patient;

b. assaying the sample for the presence of Nodal and Lefty; and

c. detecting aggressive tumor cells if Nodal is present and Lefty is absent in the sample.

37. The method of claim 36, wherein assaying for the presence of Nodal and the absence of Lefty comprises a nucleic acid based assay.

38. The method of claim 36, wherein assaying for the presence of Nodal and the absence of Lefty comprises a protein based assay.

39. A method of identifying a compound for treating aggressive tumors, comprising:

a. providing a plurality of cells that express Nodal;

b. assaying the cells for Nodal activity in the presence and absence of a candidate compound; and

c. identifying the compound as a compound for treating aggressive tumors if the Nodal activity is less in the presence of the candidate compound than in the absence of the candidate compound.

40. A method for monitoring the effectiveness of a pharmaceutical composition as an agent for treating aggressive tumors in a patient comprising the steps of:

a. obtaining a sample from a patient;

b. assaying the sample for the presence of Nodal;

c. administering an amount a pharmaceutical composition to the patient;

d. repeating steps (b) and (c) on subsequently-collected samples from the patient; and

e. comparing the amount of Nodal detected in the sample from step (a) with the amount of Nodal detected in the samples from step (d), wherein the effectiveness of the pharmaceutical composition is monitored by detecting changes in the amount of Nodal in the subsequently-collected samples compared with the sample taken in step (a).

41. A method for detecting the presence of aggressive tumor cells comprising the steps of:

a. obtaining a sample of tumor cells from a patient;

b. conducting a sequence based methylation analysis of the Nodal CpG island in the tumor cells;

c. comparing the degree of methylation in the CpG island of Nodal in the tumor cells to that of non-aggressive or non-tumor cells;

d. correlating hypermethylation of Nodal with the presence of aggressive tumor cells.

42. A method for detecting the presence cells having a dedifferentiated, multipotent plastic phenotype in a mammal comprising the steps of:

a. obtaining a sample from a mammal;

b. assaying the sample for the presence of Nodal;

c. correlating the presence of Nodal with the presence cells having a dedifferentiated, multipotent plastic phenotype.

43. The method of claim 42 wherein the sample is a bodily fluid.

44. The method of claim 43 wherein the bodily fluid is serum.