CHIMERIC ANTIGEN RECEPTOR AND METHODS OF USE THEREOF

• US 20150368342A1
• Filed: 02/14/2014
• Published: 12/24/2015
• Est. Priority Date: 02/15/2013
• Status: Abandoned Application

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: WU, Chia-Yung, ONUFFER, James, LIM, Wendell A.
• Application Number:
  US14/766,105
• Publication Number:
  US 20150368342A1
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 2239/23 On/off switch
• CPC Class Codes: A61K 2239/24 Dimerizable CARs; CARs with adapter
• CPC Class Codes: A61K 2239/31 characterized by the route of administration
• CPC Class Codes: A61K 2239/38 characterised by the dose, timing or administration schedule
• 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 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 39/464412 CD19 or B4
• CPC Class Codes: A61K 39/464468 Mesothelin [MSLN]
• CPC Class Codes: A61K 47/6891 Pre-targeting systems involving an antibody for targeting specific cells
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748
• CPC Class Codes: C07K 14/7051 T-cell receptor (TcR)-CD3 complex
• CPC Class Codes: C07K 14/70521 CD28, CD152
• CPC Class Codes: C07K 14/70575 NGF/TNF-superfamily, e.g. CD70, CD95L, CD153, CD154 NGF C07K14/48, TNF C07K14/525
• CPC Class Codes: C07K 14/70578 NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95 NGF-receptor C07K14/71, TNF-receptor C07K14/7151
• CPC Class Codes: C07K 16/18 against material from animals or humans
• CPC Class Codes: C07K 16/2803 : against the immunoglobulin superfamily
• CPC Class Codes: C07K 16/2866 : against receptors for cytokines, lymphokines, interferons
• CPC Class Codes: C07K 16/2878 : against the NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95
• CPC Class Codes: C07K 2317/622 : Single chain antibody (scFv)
• CPC Class Codes: C07K 2319/00 : Fusion polypeptide
• CPC Class Codes: C07K 2319/03 : containing a transmembrane segment
• CPC Class Codes: C07K 2319/20 : containing a tag with affinity for a non-protein ligand
• 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: C07K 2319/70 : containing domain for protein-protein interaction
• CPC Class Codes: C07K 2319/74 : containing a fusion for binding to a cell surface receptor
• CPC Class Codes: C12N 2510/00 : Genetically modified cells
• CPC Class Codes: C12N 5/0636 : T lymphocytes
• CPC Class Codes: C12N 9/14 : Hydrolases (3)
• CPC Class Codes: C12N 9/90 : Isomerases (5.)
• CPC Class Codes: C12Y 502/01008 : Peptidylprolyl isomerase (5.2.1.8), i.e. cyclophilin

Abstract

The present disclosure provides a heterodimeric, conditionally active chimeric antigen receptor (CAR), and a nucleic acid comprising a nucleotide sequence encoding the CAR. The present disclosure provides cells genetically modified to produce the CAR. A CAR of the present disclosure can be used in various methods, which are also provided.

Claims

1. A heterodimeric, conditionally active chimeric antigen receptor (CAR) comprising:

a) a first polypeptide comprising;

i) a first member of a specific binding pair;

ii) a first modulatory domain;

iii) a first member of a dimerization pair; and

iv) a transmembrane domain interposed between the first member of a specific binding pair and the first modulatory domain; and

b) a second polypeptide comprising;

i) a transmembrane domain;

ii) a second modulatory domain;

iii) a second member of the dimerization pair; and

iv) an intracellular signaling domain;

or comprising;

a) a first polypeptide comprising;

i) a first member of a specific binding pair;

ii) a modulatory domain;

iii) a first member of a dimerization pair;

iv) a transmembrane domain interposed between the first member of a specific binding pair and the modulatory domain; and

b) a second polypeptide comprising;

i) a second member of the dimerization pair; and

ii) an intracellular signaling domain.

2. The heterodimeric, conditionally active CAR of claim 1, wherein the first polypeptide comprises a hinge region interposed between the first member of the specific binding pair and the transmembrane domain.

3. The heterodimeric, conditionally active CAR of claim 1, wherein the first member of the specific binding pair is an antibody or antibody fragment, a ligand, or a receptor.

4. The heterodimeric, conditionally active CAR of claim 2, wherein the hinge region is an immunoglobulin IgG hinge region or a hinge derived from CD8.

5. The heterodimeric, conditionally active CAR of claim 1, wherein the first and second modulatory domains are selected from 4-1BB (CD137), CD28, ICOS, BTLA, OX-40, CD27, CD30, GITR, HVEM, DAP10, DAP12, and CD28.

6. The heterodimeric, conditionally active CAR of claim 1, wherein the intracellular signaling domain is selected from ZAP70 and CD3-zeta.

7. The heterodimeric, conditionally active CAR of claim 1, wherein the intracellular signaling domain comprises an immunoreceptor tyrosine-based activation motif (ITAM).

8. The heterodimeric, conditionally active CAR of claim 1, wherein the first and second members of the dimerization pair form a homodimer in the presence of a small molecule dimerizer.

9. The heterodimeric, conditionally active CAR of claim 1, wherein the first and second members of the dimerization pair form a heterodimer in the presence of a small molecule dimerizer.

10. The heterodimeric, conditionally active CAR of claim 1, wherein the first and second members of the dimerization pair are selected from:

a) FK506 binding protein (FKBP) and FKBP;

b) FKBP and calcineurin catalytic subunit A (CnA);

c) FKBP and cyclophilin;

d) FKBP and FKBP-rapamycin associated protein (FRB);

e) gyrase B (GyrB) and GyrB;

f) dihydrofolate reductase (DHFR) and DHFR;

g) DmrB and DmrB;

h) PYL and ABI;

i) Cry2 and CIP;

j) GAI and GID1.

11. The heterodimeric, conditionally active CAR of claim 1, wherein:

i) the first and second modulatory domains are derived from 4-1BB;

ii) the first and second members of the dimerization pair are FKBP and FRB; and

ii) the signaling domain comprises an ITAM.

12. The heterodimeric, conditionally active CAR of claim 1, wherein the first member of the specific binding pair is a single-chain Fv.

13. The heterodimeric, conditionally active CAR of claim 1, wherein the first member of the specific binding pair binds an epitope present on a cell, on a solid surface, or a lipid bilayer.

14. The heterodimeric, conditionally active CAR of claim 13, wherein the cell is a cancer cell.

15. A mammalian cell genetically modified to produce the heterodimeric, conditionally active CAR of claim 1.

16. The cell of claim 15, wherein the cell is a stem cell, a progenitor cell, or a cell derived from a stem cell or a progenitor cell.

17. The cell of claim 15, wherein the cell is a T lymphocyte or an NK cell.

18. A nucleic acid comprising nucleotide sequences encoding the heterodimeric, conditionally active CAR of claim 1.

19. The nucleic acid of claim 18, wherein the nucleotide sequences are operably linked to a T lymphocyte-specific promoter or an NK cell-specific promoter.

20. The nucleic acid of claim 18, wherein the nucleic acid is in vitro transcribed RNA.

21. A recombinant expression vector comprising the nucleic acid of claim 18.

22. A method of activating a T lymphocyte, the method comprising contacting the T lymphocyte with a dimerizing agent and a second member of a specific binding pair, wherein the T lymphocyte is genetically modified to produce a heterodimeric, conditionally active CAR of claim 1, and wherein, in the presence of the dimerizing agent and the second member of a specific binding pair, the heterodimeric, conditionally active CAR dimerizes and activates the T lymphocyte, thereby producing an activated T lymphocyte.

23. The method of claim 22, wherein the second member of a specific binding pair is an antigen.

24. The method of claim 22, wherein said contacting occurs in vivo.

25. The method of claim 22, wherein the activated T lymphocyte mediates killing of a target cell.

26. The method of claim 22, wherein the activated T lymphocyte produces IL-2 and/or IFN-γ

.

27. The method of claim 25, wherein the target cell is a cancer cell.

28. The method of claim 22, wherein the first member of the specific binding pair of the heterodimeric, conditionally active CAR is an antibody specific for an epitope on a cancer cell.

29. A method of making the cell of claim 15, the method comprising genetically modifying a mammalian cell with an expression vector comprising nucleotide sequences encoding the heterodimeric, conditionally active CAR of claim 1, or genetically modifying a mammalian cell with an RNA comprising nucleotide sequences encoding the heterodimeric, conditionally active CAR of claim 1.

30. The method of claim 29, wherein said genetic modification is carried out ex vivo.

31. The method of claim 29, wherein the cell is a T lymphocyte, a stem cell, an NK cell, a progenitor cell, a cell derived from a stem cell, or a cell derived from a progenitor cell.

32. A method of treating a cancer in an individual, the method comprising:

i) genetically modifying T lymphocytes obtained from the individual with an expression vector comprising nucleotide sequences encoding the heterodimeric, conditionally active CAR of claim 1, wherein the antigen-binding domain of the heterodimeric, conditionally active CAR is specific for an epitope on a cancer cell in the individual, and wherein said genetic modification is carried out ex vivo;

ii) introducing the genetically modified T lymphocytes into the individual; and

iii) administering to the individual an effective amount of a dimerizing agent, wherein the dimerizing agent induces dimerization of the heterodimeric, conditionally active CAR, wherein said dimerization provides for activation of the genetically modified T lymphocytes and killing of the cancer cell, thereby treating the cancer.

33. The method of claim 32, wherein the dimerizing agent is a rapalog.

34. A method of modulating the activity of a host cell, the method comprising contacting the host cell with a dimerizing agent and a second member of a specific binding pair, wherein the T lymphocyte is genetically modified to produce a heterodimeric, conditionally active CAR of claim 1, and wherein, in the presence of the dimerizing agent and the second member of a specific binding pair, the heterodimeric, conditionally active CAR dimerizes and modulates at least one activity of the host cell.

35. The method of claim 34, wherein the activity is proliferation, cell survival, apoptosis, gene expression, or immune activation.

36. The method of claim 34, wherein the second member of a specific binding pair is an antigen.