T cell receptor-deficient T cell compositions
The invention is directed to modified T cells, methods of making and using isolated, modified T cells, and methods of using these isolated, modified T cells to address diseases and disorders. In one embodiment, this invention broadly relates to TCR-deficient T cells, isolated populations thereof, and compositions comprising the same. In another embodiment of the invention, these TCR-deficient T cells are designed to express a functional non-TCR receptor. The invention also pertains to methods of making said TCR-deficient T cells, and methods of reducing or ameliorating, or preventing or treating, diseases and disorders using said TCR-deficient T cells, populations thereof, or compositions comprising the same.
1. A method of producing one or more compositions comprising human T cells which express a heterologous ligand binding domain or a heterologous receptor other than a T cell receptor (TCR), wherein said human T cells are derived from primary human T cells and wherein said method comprises:
(i) obtaining one or more compositions comprising isolated primary human T cells, each of said compositions being comprised of primary human T cells isolated from at least one human donor;
(ii) modifying the primary human T cells comprised in said one or more compositions comprising primary human T cells in order to functionally impair or reduce expression of one or more components of the TCR complex endogenously expressed by said human primary T cells; and
(iii) modifying the primary human T cells in the one or more compositions comprising human T cells by introducing a nucleic acid which encodes a desired exogenous ligand binding domain or a desired exogenous receptor, wherein said exogenous ligand binding domain and exogenous receptor do not comprise a TCR and further wherein said exogenous ligand binding domain or exogenous receptor firectly or indirectly mediates T cell signaling upon binding of the ligand binding domain or the exogenous receptor to a ligand expressed by a target cell;
thereby producing one or more compositions comprising human T cells which express at least one desired ligand binding domain or desired receptor and further wherein one or more components of the TCR complex endogenously expressed by said human primary T cells is functionally impaired or the expression of one or more components of the TCR complex endogenously expressed by said human primary T cells is reduced.
2. The method of claim 1 , wherein said desired ligand binding domain or desired receptor binds to tumor cells.
3. The method of claim 1 , wherein said desired ligand binding domain or desired receptor binds to infected cells.
4. The method of claim 1 , wherein said desired ligand binding domain or desired receptor binds to autoimmune cells.
5. The method of claim 1 , wherein said nucleic acid further encodes a signaling domain.
6. The method of claim 1 , wherein reduced expression of the targeted TCR component or the functional impairment or reduced expression of the endogenous TCR by the T cells in said one or more compositions obtained after effecting steps (i)-(iii) is confirmed by effecting one or more of the following procedures:
(a) detecting the level of expression of said targeted TCR component in T cells comprised in said one or more compositions,
(b) detecting the proliferation of T cells obtained in said one or more compositions in the presence of HLA-mismatched PMBCs, and
(c) detecting the level of gamma interferon produced by T cells comprised in said one or more compositions in response to HLA-mismatched donor cells.
7. The method of claim 1 , wherein said one or more compositions comprising human T cells are derived from different human donors.
8. The method of claim 1 , wherein said one or more compositions comprising human T cells are derived from the same human donor.
9. The method of claim 1 , wherein said one or more components of the TCR complex include TCR-α, TCR-β, TCR-γ, TCR-δ, CD3-γ, CD3-δ, CD3-ζ, and CD3-ε.
10. The method of claim 1 , wherein in step (i) the functional impairment or reduced expression of the one or more components of the TCR complex is effected by introducing into the primary human T cells at least one short hairpin RNA (shRNA) which targets a nucleic acid that encodes at least one component of a functional TCR expressed by said primary human T cells.
11. The method of claim 10 , wherein said one or more components of the TCR complex include TCR-α, TCR-β, TCR-γ, TCR-δ, CD3-γ, CD3δ, CD3-ζ, and CD3-ε.
12. The method of claim 1 , wherein the compositions are in a form whereby they may be stored and can be used for future therapy.
13. The method of claim 11 , wherein the compositions are in a form whereby they may be stored and can be used for future therapy.
14. The method of claim 1 , wherein the primary human T cells express CD4 or CD8.
15. The method of claim 13 , wherein the primary human T cells express CD4 or CD8.
16. The method of claim 1 , wherein the primary human T cells of (i) comprise human peripheral blood mononuclear cells or cytotoxic T cells.
17. The method of claim 15 , wherein the primary human T cells of step (i) of claim 1 comprise human peripheral blood mononuclear cells or cytotoxic T cells.
18. The method of claim 1 , wherein the ligand binding domain is obtained from NKG2D, NKG2A, NKG2C, NKG2F, LLT1, AICL, CD26 or NKRP1.
19. The method of claim 1 , wherein the ligand binding domain is obtained from an anti-tumor chimeric antigen receptor or anti-tumor antibody.
20. The method of claim 5 , wherein the signaling domain is obtained from CD3 ζ.
21. The method of claim 1 , wherein the receptor binds to a ligand expressed on tumor or infected cells.
22. The method of claim 1 , wherein the receptor binds to any one of MIC-A, MIC-B, an estrogen receptor, a progesterone receptor, RON or one or more members of the ULBP family.
23. The method of claim 22 , wherein the one or more members of the ULBP family include Rae-1, H-60, HCMV UL18, Rae-1β, ULBP2, and ULBP3.
24. The method of claim 1 , wherein the receptor comprises a ligand binding domain from NKG2D.