Optimized PNE-based chimeric receptor T cell switches and uses thereof

1. An optimized chimeric antigen receptor-effector cell (CAR-EC) platform, comprising:

(a) (1) a CAR-EC switch that comprises a chimeric antigen receptor binding peptide (CAR-BP) and a targeting moiety, wherein said targeting moiety is a Fab fragment specific for a membrane distal epitope of a cell surface molecule on a target cell, and wherein said CAR-BP is linked to the N-terminus of said Fab fragment with a linker, and

(2) a CAR-EC that comprises a CAR, wherein said CAR comprises a transmembrane domain, an intracellular signaling domain, and an extracellular domain that binds said CAR-BP, wherein said CAR-EC is a T cell, wherein said extracellular domain comprises a hinge region and an scFv that is specific for said CAR-BP; or

(b) (1) a CAR-EC switch that comprises a chimeric antigen receptor binding peptide (CAR-BP) and a targeting moiety, wherein said targeting moiety is a Fab fragment specific for a membrane proximal epitope of a cell surface molecule on a target cell, and wherein said CAR-BP is linked to the C-terminus of said Fab fragment with a linker, and

(2) a CAR-EC that comprises a CAR, wherein said CAR comprises a transmembrane domain, an intracellular signaling domain, and an extracellular domain that binds said CAR-BP, wherein said CAR-EC is a T cell, wherein said extracellular domain comprises a hinge region and an scFv that is specific for said CAR-BP;

wherein the hinge region is between 1-20 amino acid residues in length; and wherein the linker is between 4-16 amino acid residues in length.

2. A method of treating relapsed cancer, comprising administering to the subject the chimeric antigen receptor switch and the chimeric antigen receptor effector cell of the optimized chimeric antigen receptor-effector cell platform of claim 1 .

3. A method of treating cancer, comprising administering to a subject an optimized first chimeric antigen receptor effector cell (CAR-EC) platform of claim 1 .

4. The optimized CAR-EC platform of claim 1 , wherein the CAR-BP comprising a length of between about 10 and about 20 amino acids.

5. The optimized CAR-EC platform of claim 1 , wherein the CAR-EC switch comprises a plurality of CAR-BPs.

6. The optimized CAR-EC platform of claim 1 , wherein the CAR-BP comprises a peptide selected from a non-human peptide, a non-mammalian peptide, a synthetic non-naturally occurring peptide, a non-endogenous peptide, a non-immunogenic peptide, and a peptide that has low immunogenicity in a human subject.

7. The optimized CAR-EC platform of claim 1 , wherein the targeting moiety comprises (a) a Fab fragment that binds a cell surface molecule selected from CD19, CD20, CD22, CD33, CEA, CLL1, BCMA, CS1, CD123 and Her2, or (b) a sequence selected from SEQ ID NOS: 21-38, or a portion thereof, or a homolog thereof.

8. The optimized CAR-EC platform of claim 1 , wherein the targeting moiety comprises a Fab fragment specific for CD19.

9. The optimized CAR-EC platform of claim 1 , wherein the CAR-BP comprises a GCN4 peptide that is at least 85% identical to SEQ ID NO: 3.

10. The optimized CAR-EC platform of claim 1 or 9 , wherein the CAR-BP is attached to the N-terminus of the heavy chain, the light chain, or both the heavy and the light chains of the Fab fragment.

11. The optimized CAR-EC platform of claim 1 or 9 , wherein the CAR comprises an anti-GCN4 scFv.

12. The optimized CAR-EC platform of claim 1 , wherein (a) the number of CAR-BPs on the CAR-EC is between about 1 and about 8; (b) the CAR density on the CAR-EC membrane is a magnitude of about 100 to about 10{circumflex over ( )} 10 CAR per CAR-EC; and/or (c) the number of engagements between the CAR-EC and the target cell is about 1 to about 100.