Compositions and methods for tcr reprogramming using fusion proteins

Abstract

Provided herein are T-cell receptor (TCR) fusion proteins (TFPs), T-cells engineered to express one or more TFPs, and methods of use thereof for the treatment of diseases, including cancer.

1 .- 20 . (canceled) 
     
     
         21 . A pharmaceutical composition comprising
 (I) a T cell from a human subject, wherein the T cell comprises a recombinant nucleic acid molecule encoding a T cell receptor (TCR) fusion protein (TFP) comprising
 (a) a TCR subunit comprising
 (i) an extracellular domain, 
 (ii) a TCR transmembrane domain, and 
 (iii) a TCR intracellular domain; and 
 
 (b) a murine, human or humanized scFv or single domain antibody comprising an antigen binding domain; and 
   (II) a pharmaceutically acceptable carrier;   wherein the TCR subunit and the antigen binding domain are operatively linked; wherein the TCR subunit does not comprise at least one of the following: a functional CD3 zeta intracellular domain, a costimulatory domain, and a heterologous stimulatory domain;   wherein the TFP functionally interacts with an endogenous TCR when expressed in a T cell; and   wherein the T cell exhibits increased cytotoxicity to a human cell expressing an antigen that specifically interacts with the antigen binding domain compared to a T cell not containing the TFP.   
     
     
         22 . The pharmaceutical composition of  claim 21 , wherein the antigen binding domain is connected to the TCR extracellular domain by a linker. 
     
     
         23 . The pharmaceutical composition of  claim 22 , wherein the linker comprises (G4S) n , wherein G is glycine, S is serine, and n is an integer from 1 to 4. 
     
     
         24 . The pharmaceutical composition of  claim 21 , wherein the antigen binding domain comprises an anti-CD19 binding domain or an anti-BCMA binding domain. 
     
     
         25 . The pharmaceutical composition of  claim 21 , wherein the antigen binding domain comprises
 (i) a light chain (LC) CDR1, LC CDR2 and LC CDR3 sequence of SEQ ID NO: 25, SEQ ID NO: 27 and SEQ ID NO: 29, respectively;   (ii) a heavy chain (HC) CDR1, HC CDR2 and HC CDR3 sequence of SEQ ID NO: 31, SEQ ID NO: 33 and SEQ ID NO: 35, respectively; or   (iii) a combination thereof.   
     
     
         26 . The pharmaceutical composition of  claim 21 , wherein the extracellular domain, the TCR transmembrane domain and the TCR intracellular domain are derived from a single TCR subunit. 
     
     
         27 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit is CD3 gamma, CD3 delta or CD3 epsilon. 
     
     
         28 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit is TCR alpha, TCR beta, TCR delta or TCR gamma. 
     
     
         29 . The pharmaceutical composition of  claim 26 , wherein the extracellular domain comprises a full-length TCR extracellular domain of the single TCR subunit. 
     
     
         30 . The pharmaceutical composition of  claim 21 , wherein the TCR subunit does not comprise a functional CD3 zeta intracellular domain. 
     
     
         31 . The pharmaceutical composition of  claim 21 , wherein the TCR subunit does not comprise a costimulatory domain. 
     
     
         32 . The pharmaceutical composition of  claim 21 , wherein the TCR subunit does not comprise a heterologous stimulatory domain. 
     
     
         33 . The pharmaceutical composition of  claim 21 , wherein the TCR subunit does not comprise at least two of the following: a functional CD3 zeta intracellular domain, a costimulatory domain, and a heterologous stimulatory domain. 
     
     
         34 . The pharmaceutical composition of  claim 21 , wherein the TCR subunit does not comprise a functional CD3 zeta intracellular domain, a costimulatory domain and a heterologous stimulatory domain. 
     
     
         35 . The pharmaceutical composition of  claim 21 , wherein in the presence of a human cell expressing an antigen that specifically interacts with the antigen binding domain the T cell has greater than or more efficient cytotoxic activity than a T cell comprising a nucleic acid encoding a chimeric antigen receptor (CAR) comprising (a) the antigen binding domain operatively linked to (b) a CD28 extracellular domain (c) a CD28 transmembrane domain (d) a CD28 intracellular domain and (e) a CD3 zeta intracellular domain. 
     
     
         36 . The pharmaceutical composition of  claim 21 , wherein production of a pro-inflammatory cytokine by the T cell is lower compared to production of the pro-inflammatory cytokine by a T cell comprising a nucleic acid encoding a CAR comprising the antigen binding domain operatively linked to a CD28 extracellular domain, a CD28 transmembrane domain, a CD28 intracellular domain, and a CD3 zeta intracellular domain in the presence of a human cell expressing an antigen that specifically interacts with the antigen binding domain. 
     
     
         37 . The pharmaceutical composition of  claim 36 , wherein the pro-inflammatory cytokine is TNFα, GM-CSF or IL-2. 
     
     
         38 . The pharmaceutical composition of  claim 21 , wherein production of IFNγ or IL-2 by the T cell is increased compared to a T cell not containing the TFP in the presence of a human cell expressing an antigen that specifically interacts with the antigen binding domain. 
     
     
         39 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit comprises a sequence with at least 90% sequence identity to SEQ ID NO: 58. 
     
     
         40 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit comprises a sequence with at least 90% sequence identity to SEQ ID NO: 60. 
     
     
         41 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit comprises a sequence with at least 90% sequence identity to SEQ ID NO: 63. 
     
     
         42 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit is CD3 delta. 
     
     
         43 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit is TCR alpha. 
     
     
         44 . The pharmaceutical composition of  claim 26 , wherein the single TCR subunit is TCR beta. 
     
     
         45 . A method of treating cancer in a subject in need thereof comprising administering a T cell to the subject, wherein the T cell comprises a recombinant nucleic acid molecule encoding a T cell receptor (TCR) fusion protein (TFP) comprising
 (a) a TCR subunit comprising
 (i) a TCR extracellular domain, 
 (ii) a TCR transmembrane domain, and 
 (iii) a TCR intracellular domain; and 
   (b) an antibody domain comprising an antigen binding domain;   wherein the TCR subunit and the antigen binding domain are operatively linked;   wherein the TCR extracellular domain is a full length TCR extracellular domain;   wherein the TFP functionally interacts with an endogenous TCR when expressed in a T cell.   
     
     
         46 . The method of  claim 45 , wherein the TCR subunit does not comprise at least one of: a functional CD3 zeta intracellular domain, a costimulatory domain or a heterologous stimulatory domain. 
     
     
         47 . A method of treating cancer in a subject in need thereof comprising administering a T cell to the subject, wherein the T cell comprises a recombinant nucleic acid molecule encoding a T cell receptor (TCR) fusion protein (TFP) comprising
 (a) a TCR subunit comprising
 (i) an extracellular domain, 
 (ii) a TCR transmembrane domain, and 
 (iii) a TCR intracellular domain; and 
   (b) an antibody domain comprising an antigen binding domain;   wherein the TCR subunit and the antigen binding domain are operatively linked;   wherein the TCR subunit does not comprise at least one of: a functional CD3 zeta intracellular domain, a costimulatory domain or a heterologous stimulatory domain;   wherein the TCR transmembrane domain and the TCR intracellular domain are derived from a single TCR subunit; and   wherein the TFP functionally interacts with an endogenous TCR when expressed in a T cell.   
     
     
         48 . The method of  claim 47 , wherein the single TCR subunit is selected from the group consisting of CD3 epsilon, CD3 gamma, CD3 delta, TCR alpha, TCR beta, TCR gamma and TCR delta. 
     
     
         49 . The method of  claim 47 , wherein the extracellular domain, the TCR transmembrane domain and the TCR intracellular domain are derived from a single TCR subunit. 
     
     
         50 . The method of  claim 47 , wherein the extracellular domain is a full length TCR extracellular domain.