COMPOSITIONS AND METHODS FOR TCR REPROGRAMMING USING FUSION PROTEINS

1 .- 218 . (canceled)

219 . A recombinant nucleic acid molecule comprising:

a first nucleic acid sequence encoding a T cell receptor (TCR) fusion protein (TFP) wherein the TFP comprises:

(a) a TCR subunit comprising:

(i) at least a portion of a TCR extracellular domain, and

(ii) a TCR transmembrane domain, and

(b) an antigen binding domain; and

wherein the TCR subunit and the antigen binding domain are operatively linked, and a second nucleic acid sequence encoding an interleukin-15 (IL-15) polypeptide or a fragment thereof and/or an IL-15 (IL-15R) polypeptide or a fragment thereof.

220 . The recombinant nucleic acid molecule of claim 219 , wherein the TFP further comprises a TCR intracellular signaling domain.

221 . The recombinant nucleic acid molecule of claim 219 , wherein the first nucleic acid sequence and the second nucleic acid sequence are operatively linked by a first linker comprising a protease cleavage site.

222 . The recombinant nucleic acid molecule of claim 219 , wherein the IL-15 polypeptide is secreted when expressed in a T cell.

223 . The recombinant nucleic acid molecule of claim 219 , wherein the second nucleic acid sequence encodes an IL-15 polypeptide and an IL-15R alpha (IL-15Rα) polypeptide.

224 . The recombinant nucleic acid molecule of claim 223 , wherein the IL-15 polypeptide comprises a sequence of SEQ ID NO: 375 and the IL-15Rα polypeptide comprises a sequence of SEQ ID NO: 403.

225 . The recombinant nucleic acid molecule of claim 223 , wherein the IL-15 polypeptide and the IL-15Rα polypeptide are operatively linked by a second linker.

226 . The recombinant nucleic acid molecule of claim 225 , wherein the second linker comprises a sequence of SEQ ID NO: 378 or 405.

227 . The recombinant nucleic acid molecule of claim 223 , wherein the second nucleic acid sequence encodes a fusion protein comprising the IL-15 polypeptide linked to the N-terminus of the TL-15Rα polypeptide.

228 . The recombinant nucleic acid molecule of claim 227 , wherein the fusion protein comprises a sequence having at least 95% identity to SEQ ID NO: 389.

229 . The recombinant nucleic acid molecule of claim 227 , wherein the fusion protein is expressed on cell surface when expressed in a T cell.

230 . The recombinant nucleic acid molecule of claim 219 , wherein the recombinant nucleic acid molecule encodes an amino acid sequence of SEQ ID NO: 377, SEQ ID NO: 380, SEQ ID NO: 381, SEQ ID NO: 402, or SEQ ID NO: 404.

231 . The recombinant nucleic acid molecule of claim 220 , wherein the TCR intracellular signaling domain comprises a stimulatory domain from an intracellular signaling domain of CD3 gamma, CD3 delta, or CD3 epsilon.

232 . The recombinant nucleic acid molecule of claim 219 , wherein the antigen binding domain is connected to the TCR extracellular domain by a third linker sequence.

233 . The recombinant nucleic acid molecule of claim 220 , wherein at least two of the TCR extracellular domain, the TCR transmembrane domain, and the TCR intracellular signaling domain are from the same TCR subunit, and wherein the same TCR subunit is selected from the group consisting of CD3 epsilon, CD3 delta, and CD3 gamma.

234 . The recombinant nucleic acid molecule of claim 219 , wherein the antigen binding domain is a camelid antibody or binding fragment thereof; a murine antibody or binding fragment thereof; or a human or humanized antibody or binding fragment thereof.

235 . The recombinant nucleic acid molecule of claim 219 , wherein the antigen binding domain is a single-chain variable fragment (scFv) or a single domain antibody (sdAb) domain.

236 . The recombinant nucleic acid molecule of claim 219 , wherein the antigen binding domain is selected from the group consisting of an anti-CD19 binding domain, an anti-B-cell maturation antigen (BCMA) binding domain, and an anti-mesothelin (MSLN) binding domain, an anti-CD20 binding domain, an anti-CD70 binding domain, anti-MUC16 binding domain, an anti-Nectin-4 binding domain, an anti-GPC3 binding domain, and an anti-TROP-2 binding domain.

237 . A polypeptide encoded by the recombinant nucleic acid molecule of claim 219 .

238 . A vector comprising the recombinant nucleic acid molecule of claim 219 .

239 . A cell comprising the recombinant nucleic acid molecule of claim 219 .

240 . The cell of claim 239 , wherein the cell is a T cell.

241 . A cell comprising:

a first nucleic acid sequence encoding a T cell receptor (TCR) fusion protein (TFP) wherein the TFP comprises:

(a) a TCR subunit comprising:

(i) at least a portion of a TCR extracellular domain, and

(ii) a TCR transmembrane domain, and

(b) an antigen binding domain; and

wherein the TCR subunit and the antigen binding domain are operatively linked, and a second nucleic acid sequence encoding a fusion protein comprising an IL-15 polypeptide or a fragment thereof and an IL-15Rα polypeptide or a fragment thereof.

242 . The cell of claim 241 , wherein the first nucleic acid sequence and the second nucleic acid sequence are included in a single nucleic acid molecule.

243 . The cell of claim 241 , wherein the first nucleic acid sequence and the second nucleic acid sequence are included in two separate nucleic acid molecules.

244 . A pharmaceutical composition comprising the cell of claim 241 and a pharmaceutically acceptable carrier.

245 . A method of increasing the activity or persistence of a cell expressing a recombinant nucleic acid molecule comprising a sequence encoding a T cell receptor (TCR) fusion protein (TFP), the method comprising expressing an interleukin-15 (IL-15) polypeptide or a fragment thereof and/or an IL-15Rα polypeptide or a fragment thereof in the cell:

wherein the TFP comprises:

(a) a TCR subunit comprising:

(i) at least a portion of a TCR extracellular domain, and

(ii) a TCR transmembrane domain, and

(b) an antigen binding domain; and

wherein the TCR subunit and the antigen binding domain are operatively linked.

246 . A method of treating a disease or a condition in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the pharmaceutical composition of claim 244 .

247 . The method of claim 246 , wherein the disease or the condition is a cancer or a disease or a condition associated with expression of CD19, B-cell maturation antigen (BCMA), mesothelin (MSLN), CD20, CD70, MUC16, Trop-2, Nectin-4, or GPC3.