Chimeric ILT receptor compositions and methods
Provided are chimeric ILT receptors (CIRs) that include a targeting region from ILT2 or ILT4, a transmembrane domain, and an intracellular domain (ICD). The ICD includes a signaling region (e.g., CD3 zeta (CD3ζ)) and optionally a costimulatory region (e.g., CD28, 4-1BB, OX40, and the like). Also provided are nucleic acids (e.g., expression vectors) encoding a subject CIR, and genetically modified cells (e.g., immune cells such as NK cells, T cells, iNKT cells, macrophages, and the like) expressing a subject CIR. For example, provided are genetically modified immune cells such as NK cells that include a nucleic acid encoding an ILT2 or ILT4 CIR. The subject CIRs are designed to activate cytotoxicity by immune cells such as NK cells, T cells, iNKT cells and macrophages against HLA-G expressing cancers.
1. A chimeric receptor protein, comprising:
(a) a targeting region, that targets HLA-G, comprising an immunoglobulin-like transcript 4 (ILT4) D1-D2 extracellular domain that comprises an amino acid sequence that is 90% or more identical to SEQ ID NO: 57, wherein the targeting region lacks an ILT4 D3-D4 extracellular domain;
(b) a CD8α stalk domain;
(c) a CD8α transmembrane domain; and
(d) an intracellular domain (ICD), comprising:
(i) a signaling region capable of transducing a signal, upon binding of said targeting region to HLA-G, into the interior of an immune effector cell to elicit effector cell function, wherein the signaling region comprises a CD3ζ signaling domain; and
(ii) a costimulatory region comprising a 4-1BB costimulatory domain.
2. The chimeric receptor protein of claim 1 , wherein the targeting region comprises an amino acid mutation at a position corresponding to Y96 of the ILT4 amino acid sequence set forth in SEQ ID NO: 57.
3. An isolated nucleic acid, comprising a nucleotide sequence encoding the chimeric receptor protein of claim 1 .
4. The isolated nucleic acid of claim 3 , wherein said nucleic acid is an expression vector.
5. An isolated genetically modified cell, expressing the chimeric receptor protein of claim 1 .
6. The isolated genetically modified cell of claim 5 , wherein the genetically modified cell is a natural killer (NK) cell, a T cell, an iNKT cell, or a macrophage.
7. A method of treatment, comprising administering the genetically modified cell of claim 5 to an individual in need, wherein the individual has diseased cells that express HLA-G.
8. The method of claim 7 , wherein the individual has a cancer with HLA-G expressing cancer cells.
9. A method of producing an isolated genetically modified cell, the method comprising:
introducing the nucleic acid of claim 3 into an isolated cell or an isolated population of cells, thus producing an isolated genetically modified cell.
10. The chimeric receptor protein of claim 1 , wherein the CD8α stalk domain comprises the amino acid sequence of SEQ ID NO: 107; the CD8α transmembrane domain comprises the amino acid sequence of SEQ ID NO: 100; the CD3ζ signaling domain comprises the amino acid sequence of SEQ ID NO: 33; and the 4-1BB costimulatory domain comprises the amino acid sequence of SEQ ID NO: 35.
11. The chimeric receptor protein of claim 1 , wherein the ILT4 D1-D2 extracellular domain comprises an amino acid sequence that is 95% or more identical to SEQ ID NO: 57.
12. The chimeric receptor protein of claim 10 , wherein the ILT4 D1-D2 extracellular domain comprises an amino acid sequence that is 95% or more identical to SEQ ID NO: 57.
13. The chimeric receptor protein of claim 1 , wherein the ILT4 D1-D2 extracellular domain comprises the amino acid sequence of SEQ ID NO: 57.
14. The chimeric receptor protein of claim 10 , wherein the ILT4 D1-D2 extracellular domain comprises the amino acid sequence of SEQ ID NO: 57.
15. The isolated nucleic acid of claim 3 , wherein said nucleic acid is a plasmid, a viral vector, or a transposon.
16. The isolated nucleic acid of claim 3 , wherein the nucleotide sequence encoding the chimeric receptor protein is operably linked to a constitutive promoter or to an inducible promoter.
17. The method of claim 7 , wherein the individual is a human.
18. The method of claim 7 , wherein the genetically modified cell is autologous to the individual.
19. The method of claim 7 , wherein the genetically modified cell is allogeneic to the individual.