Engineered three-dimensional breast tissue, adipose tissue, and tumor disease model

• US 9,983,195 B2
• Filed: 04/03/2015
• Issued: 05/29/2018
• Est. Priority Date: 04/04/2014
• Status: Active Grant

• Current Assignee: Organovo Inc. (Organovo Holdings, Inc.)
• Original Assignee: Organovo Inc. (Organovo Holdings, Inc.)
• Inventors: King, Shelby Marie, Nguyen, Deborah Lynn Greene, Gorgen, Vivian A., Shepherd, Benjamin R., Presnell, Sharon C.
• Primary Examiner(s): Moloye, Titilayo
• Application Number:
  US14/678,392
• Publication Number:
  US 20150282885A1
• Time in Patent Office: 1,152 Days
• CPC Class Codes: B33Y 10/00 Processes of additive manufacturing
• CPC Class Codes: B33Y 80/00 Products made by additive manufacturing
• CPC Class Codes: C12N 2502/13 connective tissue cells; generic mesenchyme cells, e.g. so-called "embryonic fibroblasts"
• CPC Class Codes: C12N 2502/1305 Adipocytes
• CPC Class Codes: C12N 2502/1323 Adult fibroblasts
• CPC Class Codes: C12N 2502/1358 Bone marrow mesenchymal stem cells (BM-MSC)
• CPC Class Codes: C12N 2502/28 Vascular endothelial cells
• CPC Class Codes: C12N 2513/00 3D culture
• CPC Class Codes: C12N 2533/74 Alginate
• CPC Class Codes: C12N 5/0656 Adult fibroblasts
• CPC Class Codes: C12N 5/0693 Tumour cells; Cancer cells
• CPC Class Codes: C12N 5/0697 Artificial constructs associating cells of different lineages, e.g. tissue equivalents blood vessels C12N5/0691
• CPC Class Codes: G01N 33/5011 for testing antineoplastic activity
• CPC Class Codes: G01N 33/5082 Supracellular entities, e.g. tissue, organisms

Abstract

Described are three-dimensional, engineered, biological breast tissues, adipose tissues, and tumor models, including breast cancer models.

Claims

1. A three-dimensional, engineered, biological tumor model comprising:

a. stromal tissue; and

b. tumor tissue;

the tumor tissue comprising cancer cells, the tumor tissue in contact with the stromal tissue to form the three-dimensional, engineered, biological tumor model, wherein the tumor model does not comprise a perfusable vascular network, does not comprise red blood cells, and lacks innervation and neural tissue;

provided that the stromal tissue was bioprinted from a stromal bio-ink comprising human preadipocytes exposed to an adipocyte differentiation media for preadipocytes, and the tumor tissue was bioprinted from a tumor bio-ink.

2. The tumor model of claim 1, wherein the tumor tissue is surrounded on three or more sides by the stromal tissue to form the three-dimensional, engineered, biological tumor model.

3. The tumor model of claim 2, wherein the tumor tissue is completely surrounded on all sides by the stromal tissue to form the three-dimensional, engineered, biological tumor model.

4. The tumor model of claim 1, wherein the model is substantially free of pre-formed scaffold.

5. The tumor model of claim 1, wherein the stromal tissue comprises:

a. endothelial cells

b. fibroblasts, and

c. adipocytes or preadipocytes or both adipocytes and preadipocytes.

6. The tumor model of claim 1, wherein the tumor tissue further comprises one or more of:

endothelial cells, fibroblasts, adipocytes, and preadipocytes.

7. The tumor model of claim 1, wherein the tumor model is a human breast cancer model, the stromal tissue is human breast stroma comprising human mammary fibroblasts, human endothelial cells, and human adipocytes, and the tumor tissue is human breast tumor.

8. A three-dimensional, engineered, biological tumor model comprising:

a. stromal tissue; and

b. tumor tissue;

the tumor tissue comprising cancer cells, the tumor tissue in contact with the stromal tissue to form the three-dimensional, engineered, biological tumor model, wherein the tumor model does not comprise a perfusable vascular network, does not comprise red blood cells, and lacks innervation and neural tissue;

provided that the stromal tissue was bioprinted from a stromal bio-ink comprising human preadipocytes exposed to an adipocyte differentiation media for preadipocytes, and the tumor tissue was bioprinted from a tumor bio-ink;

provided that the stromal bio-ink and the tumor bio-ink each comprise a reversibly cross-linkable extrusion compound utilized to physically stabilize the tumor model architecture prior to cohesion of the cells.

9. The tumor model of claim 8, wherein the tumor tissue is surrounded on three or more sides by the stromal tissue to form the three-dimensional, engineered, biological tumor model.

10. The tumor model of claim 9, wherein the tumor tissue is completely surrounded on all sides by the stromal tissue to form the three-dimensional, engineered, biological tumor model.

11. The tumor model of claim 8, wherein the model is substantially free of pre-formed scaffold.

12. The tumor model of claim 8, wherein the stromal tissue comprises:

a. endothelial cells,

b. fibroblasts, and

c. adipocytes or preadipocytes or both adipocytes and preadipocytes.

13. The tumor model of claim 8, wherein the tumor tissue further comprises one or more of:

endothelial cells, fibroblasts, adipocytes, and preadipocytes.

14. The tumor model of claim 8, wherein the tumor model is a human breast cancer model, the stromal tissue is human breast stroma comprising human mammary fibroblasts, human endothelial cells, and human adipocytes, and the tumor tissue is human breast tumor.

15. An array of three-dimensional, engineered, biological tumor models, each tumor model comprising:

stromal tissue and tumor tissue;

the tumor tissue comprising cancer cells, the tumor tissue in contact with the stromal tissue to form each three-dimensional, engineered, biological tumor model, wherein the tumor model does not comprise a perfusable vascular network, does not comprise red blood cells, and lacks innervation and neural tissue;

provided that the stromal tissue was bioprinted from a stromal bio-ink comprising human preadipocytes exposed to an adipocyte differentiation media for preadipocytes;

provided that the array is adapted for use in a high throughput assay.

16. The array of claim 15, wherein the tumor tissue is surrounded on three or more sides by the stromal tissue to form each three-dimensional, engineered, biological tumor model.

17. The array of claim 16, wherein the tumor tissue is completely surrounded on all sides by the stromal tissue to form each three-dimensional, engineered, biological tumor model.

18. The array of claim 15, wherein each tumor model is substantially free of pre-formed scaffold.

19. The array of claim 15, wherein each tumor model is in a well of a multi-well plate.

20. The array of claim 15, wherein the stromal tissue comprises:

endothelial cells, fibroblasts, and adipocytes or preadipocytes or both adipocytes and preadipocytes.

21. The array of claim 15, wherein the tumor tissue further comprises one or more of:

endothelial cells, fibroblasts, adipocytes, and preadipocytes.

22. The array of claim 15, wherein each tumor model is a human breast cancer model, the stromal tissue is human breast stroma comprising human mammary fibroblasts, human endothelial cells, and human adipocytes, and the tumor tissue is human breast tumor.

23. An array of three-dimensional, engineered, biological tumor models, each tumor model comprising:

stromal tissue and tumor tissue;

the tumor tissue comprising cancer cells, the tumor tissue in contact with the stromal tissue to form each three-dimensional, engineered, biological tumor model, wherein the tumor model does not comprise a perfusable vascular network, does not comprise red blood cells, and lacks innervation and neural tissue;

provided that the stromal tissue was bioprinted from a stromal bio-ink comprising a reversibly crosslinkable extrusion compound and human preadipocytes exposed to an adipocyte differentiation media for preadipocytes, and the tumor tissue was bioprinted from a tumor bio-ink comprising a reversibly crosslinkable extrusion compound;

provided that the array is adapted for use in a high throughput assay.

24. The array of claim 23, wherein the tumor tissue is surrounded on three or more sides by the stromal tissue to form each three-dimensional, engineered, biological tumor model.

25. The array of claim 24, wherein the tumor tissue is completely surrounded on all sides by the stromal tissue to form each three-dimensional, engineered, biological tumor model.

26. The array of claim 23, wherein each tumor model is substantially free of pre-formed scaffold.

27. The array of claim 23, wherein each tumor model is in a well of a multi-well plate.

28. The array of claim 23, wherein the stromal tissue comprises:

endothelial cells, fibroblasts, and adipocytes, preadipocytes, or both adipocytes and preadipocytes.

29. The array of claim 23, wherein the tumor tissue further comprises one or more of:

endothelial cells, fibroblasts, and adipocytes, preadipocytes, or both adipocytes and preadipocytes.

30. The array of claim 23, wherein each tumor model is a human breast cancer model, the stromal tissue is human breast stroma comprising human mammary fibroblasts, human endothelial cells, and human adipocytes, and the tumor tissue is human breast tumor.