Compositions and Methods for Generating a Persisting Population of T Cells Useful for the Treatment of Cancer

• US 20150024482A1
• Filed: 02/22/2013
• Published: 01/22/2015
• Est. Priority Date: 02/22/2012
• Status: Active Grant

• Current Assignee: Trustees Of The University Of Pennsylvania (University of Pennsylvania)
• Original Assignee: Trustees Of The University Of Pennsylvania (University of Pennsylvania)
• Inventors: Zhao, Yangbing, Scholler, John, June, Carl H., Frigault, Matthew J.
• Granted Patent: US 10,040,846 B2
• Time in Patent Office: Days
• US Class Current: 435/325
• CPC Class Codes: A61K 39/4611 T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
• CPC Class Codes: A61K 39/4631 Chimeric Antigen Receptors [CAR]
• CPC Class Codes: A61K 39/464402 Receptors, cell surface antigens or cell surface determinants
• CPC Class Codes: A61K 39/464412 CD19 or B4
• CPC Class Codes: A61K 39/464468 Mesothelin [MSLN]
• CPC Class Codes: C07K 14/7051 T-cell receptor (TcR)-CD3 complex
• CPC Class Codes: C07K 14/70521 CD28, CD152
• CPC Class Codes: C07K 16/18 against material from animals or humans
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 2319/33 fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2510/00 Genetically modified cells
• CPC Class Codes: C12N 2740/15041 Use of virus, viral particle or viral elements as a vector
• CPC Class Codes: C12N 2840/00 Vectors comprising a special translation-regulating system
• CPC Class Codes: C12N 5/0636 T lymphocytes

Abstract

The present invention provides compositions and methods for generating a genetically modified T cells comprising a chimeric antigen receptor (CAR) having an antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the T cell exhibits prolonged exponential expansion in culture that is ligand independent and independent of the addition of exogenous cytokines or feeder cells.

Claims

1. An isolated nucleic acid sequence encoding a chimeric antigen receptor (CAR), wherein the CAR comprises an antigen binding domain, a hinge domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, and further wherein when the CAR is transduced into a T cell, the CAR contributes to at least one of:

increased antigen-independent activation of the transduced T cell, increased mean cell volume (MCV) of the transduced T cell, increased cell population expansion of the transduced T cell, increased proliferation of the transduced T cell, increased numbers of progeny of the transduced T cell, increased effector cytokine secretion, sustained expression of granzyme, increased persistence of the transduced T cell population in vitro, or increased persistence of the transduced T cell population in vivo.

2. The isolated nucleic acid sequence of claim 1, wherein the hinge domain is an IgG4 hinge domain.

3. The isolated nucleic acid sequence of claim 1, wherein the antigen binding domain is an anti-cMet binding domain, the hinge domain is IgG4, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

4. The isolated nucleic acid sequence of claim 3, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

1.

5. The isolated nucleic acid sequence of claim 1, wherein the antigen binding domain is an anti-mesothelin binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

6. The isolated nucleic acid sequence of claim 5, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

2.

7. The isolated nucleic acid sequence of claim 1, wherein the antigen binding domain is an anti-CD 19 binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is an CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

8. The isolated nucleic acid sequence of claim 7, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

3.

9. The isolated nucleic acid sequence of claim 1, wherein the antigen binding domain is an antibody or an antigen-binding fragment thereof.

10. A T cell comprising a nucleic acid sequence encoding a chimeric antigen receptor (CAR), the CAR comprising an antigen binding domain, a hinge domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, and wherein when the CAR is transduced into a T cell, the CAR contributes to at least one of:

increased antigen-independent activation of the transduced T cell, increased mean cell volume (MCV) of the transduced T cell, increased cell population expansion of the transduced T cell, increased proliferation of the transduced T cell, increased effector cytokine secretion, increased expression of granzyme, increased numbers of progeny of the transduced T cell, increased persistence of the transduced T cell population in vitro, or increased persistence of the transduced T cell population in vivo.

11. The T cell of claim 10, wherein the hinge domain is an IgG4 hinge domain.

12. The T cell of claim 10, wherein the antigen binding domain is an anti-cMet binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

13. The T cell of claim 12, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

1.

14. The T cell of claim 10, wherein the antigen binding domain is an anti-mesothelin binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is an CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

15. The T cell of claim 14, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

2.

16. The T cell of claim 10, wherein the antigen binding domain is an anti-CD 19 binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

17. The T cell of claim 16, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

3.

18. The T cell of claim 10, wherein the antigen binding domain is an antibody or an antigen-binding fragment thereof.

19. The T cell of claim 10, wherein the cell exhibits an anti-tumor immunity when the antigen binding domain binds to its corresponding antigen.

20. A vector comprising a nucleic acid sequence encoding a chimeric antigen receptor (CAR), the CAR comprising an antigen binding domain, a hinge domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, and wherein when the CAR is transduced into a T cell, the CAR contributes to at least one of:

increased antigen-independent activation of the transduced T cell, increased mean cell volume (MCV) of the transduced T cell, increased cell population expansion of the transduced T cell, increased proliferation of the transduced T cell, increased numbers of progeny of the transduced T cell, increased persistence of the transduced T cell population in vitro, or increased persistence of the transduced T cell population in vivo.

21. The vector of claim 20, wherein the hinge domain is an IgG4 hinge domain.

22. The vector of claim 20, wherein the antigen binding domain is an anti-cMet binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

23. The vector of claim 22, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

1.

24. The vector of claim 20, wherein the antigen binding domain is an anti-mesothelin binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

25. The vector of claim 24, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

2.

26. The vector of claim 20, wherein the antigen binding domain is an anti-CD 19 binding domain, the hinge domain is an IgG4 hinge domain, the transmembrane domain is a CD28 transmembrane domain, and the costimulatory signaling region is a CD28 signaling region.

27. The vector of claim 26, wherein the CAR comprises the amino acid sequence of SEQ ID NO:

3.

28. The vector of claim 20, wherein the antigen binding domain is an antibody or an antigen-binding fragment thereof.

29. A persisting population of genetically modified T cells, wherein the T cells comprise a nucleic acid sequence encoding a chimeric antigen receptor (CAR), the CAR comprising an antigen binding domain, a hinge domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, and wherein when the CAR is transduced into a T cell, the CAR contributes to at least one of:

increased antigen-independent activation of the transduced T cell, increased mean cell volume (MCV) of the transduced T cell, increased cell population expansion of the transduced T cell, increased proliferation of the transduced T cell, increased numbers of progeny of the transduced T cell, increased persistence of the transduced T cell population in vitro, or increased persistence of the transduced T cell population in vivo.

30. The persisting population of genetically modified T cells of claim 29, wherein the genetically modified T cells exhibit an anti-tumor immunity when the antigen binding domain binds to its corresponding antigen.

31. The persisting population of genetically modified T cells of claim 29, wherein the T cells exhibit a cytokine signature comprising at least one cytokine selected from the group consisting of IFN-γ

, TNF-α

, IL-17A, IL-2, IL-3, IL-4, GM-CSF, IL-10, IL-13, Granzyme B, Perforin, and any combination thereof.

32. The persisting population of genetically modified T cells of claim 29, wherein the T cells proliferate in the absence of exogenous cytokine or feeder cells.