COMPOSITIONS AND METHODS FOR IMMUNOTHERAPY
The present invention provides immunoresponsive cells, including T cells, cytotoxic T cells, regulatory T cells, and Natural Killer (NK) cells, expressing an antigen recognizing receptor and an inhibitory chimeric antigen receptor (iCAR). Methods of using the immunoresponsive cell include those for the treatment of neoplasia and other pathologies where an increase in an antigen-specific immune response is desired.
1 . A method of producing an immunoresponsive cell, the method comprising introducing into an immunoresponsive cell:
a) a first polynucleotide encoding a chimeric antigen receptor (CAR) comprising an extracellular antigen-binding domain comprising a single chain variable fragment (scFv) that binds to a first antigen that is expressed at the surface of a cell of the tumor, and an intracellular signaling domain that is capable of activating the immunoresponsive cell and comprises a signaling domain of CD28, and
b) a second polynucleotide encoding an inhibitory chimeric antigen receptor (iCAR) comprising an extracellular antigen-binding domain comprising a single chain variable fragment (scFv) that binds to a second antigen that is not expressed on the tumor cell surface, and a signaling domain of an immunoinhibitory receptor selected from the group consisting of CTLA-4, PD-1, LAG-3, 2B4, and BTLA.
2 . The method of claim 1 , wherein the first polynucleotide is included in a vector.
3 . The method of claim 1 , wherein the second polynucleotide is included in a vector.
4 . The method of claim 1 , wherein the immunoresponsive cell is selected from the group consisting of a T cell, a Natural Killer (NK) cell, a cell of the innate immune system, and a pluripotent stem cell from which a lymphoid cell may be differentiated.
5 . The method of claim 4 , wherein the immunoresponsive cell is a T cell.
6 . The method of claim 5 , wherein the T cell is selected from the group consisting of effector T cells, and memory T cells.
7 . The method of claim 6 , wherein the effector T cells are selected from the group consisting of helper T cells (CD4 + T cells), and cytotoxic T cells (CD8 + T cells).
8 . The method of claim 1 , wherein the first antigen is selected from the group consisting of CD19, CD7, CD10, CD20, CD22, CD30, CD33, CD34, CD38, CD41, CD44, CD49f, CD56, CD74, CD123, CD133, CD138, CAIX, CEA, CD5, EGP-2, EGP-40, EpCAM, Erb-B2, Erb-B3, Erb-B4, FBP, Fetal acetylcholine receptor, folate receptor-α, GD2, GD3, HER-2, IL-13R-α2, κ-light chain, LeY, L1 cell adhesion molecule, Mesothelin, Muc-1, Muc-16, oncofetal antigen (h5T4), PSCA, PSMA, ROR1, TAG-72, and VEGF-R2, optionally wherein the first antigen is selected from the group consisting of CD19, PSMA, mesothelin, and CD56.
9 . The method of claim 1 , wherein the iCAR further comprises a transmembrane domain selected from the group consisting of a CD4 polypeptide, a CD8 polypeptide, a CTLA-4 polypeptide, a PD-1 polypeptide, a LAG-3 polypeptide, a 2B4 polypeptide, and a BTLA polypeptide.
10 . The method of claim 1 , wherein the second antigen is selected from the group consisting of an Epithelial-mesenchymal transition (EMT) antigen, cytokeratin, human leukocyte antigens (HLAs), Opioid-binding protein/cell adhesion molecule (OPCML), HYAL2, Deleted in Colorectal Carcinoma (DCC), Scaffold/Matrix attachment region-binding protein 1 (SMAR1), CD33, CD38, and E-cadherin.
11 . A method of producing an immunoresponsive cell comprising introducing into an immunoresponsive cell a vector encoding:
a) a chimeric antigen receptor (CAR) comprising an extracellular antigen-binding domain comprising a single chain variable fragment (scFv) that binds to a first antigen that is expressed at the surface of a cell of the tumor, and an intracellular signaling domain that is capable of activating the immunoresponsive cell and comprises a signaling domain of CD28, and
b) an inhibitory chimeric antigen receptor (iCAR) comprising an extracellular antigen-binding domain comprising a single chain variable fragment (scFv) that binds to a second antigen that is not expressed on the tumor cell surface, and a signaling domain of an immunoinhibitory receptor selected from the group consisting of CTLA-4, PD-1, LAG-3, 2B4, and BTLA.
12 . The method of claim 11 , wherein the vector is a viral vector.
13 . The method of claim 11 , wherein the vector is a retroviral vector.
14 . The method of claim 11 , wherein the immunoresponsive cell is selected from the group consisting of a T cell, a Natural Killer (NK) cell, a cell of the innate immune system, and a pluripotent stem cell from which a lymphoid cell may be differentiated.
15 . The method of claim 14 , wherein the immunoresponsive cell is a T cell.
16 . The method of claim 15 , wherein the T cell is selected from the group consisting of effector T cells, and memory T cells.
17 . The method of claim 16 , wherein the effector T cells are selected from the group consisting of helper T cells (CD4 + T cells), and cytotoxic T cells (CD8 + T cells).
18 . The method of claim 11 , wherein the first antigen is selected from the group consisting of CD19, CD7, CD10, CD20, CD22, CD30, CD33, CD34, CD38, CD41, CD44, CD49f, CD56, CD74, CD123, CD133, CD138, CAIX, CEA, CD5, EGP-2, EGP-40, EpCAM, Erb-B2, Erb-B3, Erb-B4, FBP, Fetal acetylcholine receptor, folate receptor-a, GD2, GD3, HER-2, IL-13R-α2, κ-light chain, LeY, L1 cell adhesion molecule, Mesothelin, Muc-1, Muc-16, oncofetal antigen (h5T4), PSCA, PSMA, ROR1, TAG-72, and VEGF-R2, optionally wherein the first antigen is selected from the group consisting of CD19, PSMA, mesothelin, and CD56.
19 . The method of claim 11 , wherein the iCAR further comprises a transmembrane domain selected from the group consisting of a CD4 polypeptide, a CD8 polypeptide, a CTLA-4 polypeptide, a PD-1 polypeptide, a LAG-3 polypeptide, a 2B4 polypeptide, and a BTLA polypeptide.
20 . The method of claim 11 , wherein the second antigen is selected from the group consisting of an Epithelial-mesenchymal transition (EMT) antigen, cytokeratin, human leukocyte antigens (HLAs), Opioid-binding protein/cell adhesion molecule (OPCML), HYAL2, Deleted in Colorectal Carcinoma (DCC), Scaffold/Matrix attachment region-binding protein 1 (SMAR1), CD33, CD38, and E-cadherin.