Dual specificity polypeptide molecule

Abstract

The present invention relates to a bispecific polypeptide molecule comprising a first polypeptide chain and a second polypeptide chain providing a binding region derived from a T cell receptor (TCR) being specific for a major histocompatibility complex (MHC)-associated peptide epitope, and a binding region derived from an antibody capable of recruiting human immune effector cells by specifically binding to a surface antigen of said cells, as well as methods of making the bispecific polypeptide molecule, and uses thereof.

1 . A method of eliciting an immune response in a patient who has cancer, comprising administering to a patient a dual specificity polypeptide molecule comprising a first polypeptide chain and a second polypeptide chain, wherein:
 the first polypeptide chain comprises
 a first binding region of a variable domain (VD1) of an antibody specifically binding to a cell surface antigen of a human immune effector cell, and 
 a first binding region of a variable domain (VR1) of a TCR specifically binding to an MHC-associated peptide epitope, and 
 a first linker (LINK1) connecting said domains; 
   the second polypeptide chain comprises
 a second binding region of a variable domain (VR2) of a TCR specifically binding to an MHC-associated peptide epitope, and 
 a second binding region of a variable domain (VD2) of an antibody specifically binding to a cell surface antigen of a human immune effector cell, and 
 a second linker (LINK2) connecting said domains; 
 wherein said first binding region (VD1) and said second binding region (VD2) associate to form a first binding site (VD1)(VD2) that binds a cell surface antigen of a human immune effector cell; said first binding region (VR1) and said second binding region (VR2) associate to form a second binding site (VR1)(VR2) that binds said MHC-associated peptide epitope; 
 wherein said two polypeptide chains are fused to human IgG hinge domains and/or human IgG Fc domains or dimerizing portions thereof; and 
 wherein said two polypeptide chains are connected by covalent and/or non-covalent bonds between said hinge domains and/or Fc-domains; and 
 wherein said dual specificity polypeptide molecule is capable of simultaneously binding the cell surface molecule and the MHC-associated peptide epitope, 
 wherein the order of the binding regions in the two polypeptide chains is selected from VD1-VR1 and VR2-VD2 or VD1-VR2 and VR1-VD2, or VD2-VR1 and VR2-VD1 or VD2-VR2 and VR1-VD1, 
 wherein the domains are either connected by LINK1 or LINK2, and 
 wherein the cancer is selected from the group consisting of bladder cancer, breast cancer, colon cancer, kidney cancer, liver cancer, lung cancer, ovarian cancer, pancreatic cancer, stomach cancer, prostate cancer, cervical cancer, melanoma, thyroid cancer, brain cancer, leukemia, sarcoma, and lymphoma. 
   
     
     
         2 . The method according to  claim 1 , wherein the order of the binding regions in the polypeptide chains is selected from VD1-VR1 and VD2-VR2; and wherein the domains are connected by LINK1 or LINK2, respectively. 
     
     
         3 . The method according to  claim 1 , wherein the linker-sequences LINK1 and/or LINK2 contain at least one sequence motif selected from GGGS, GGGGS, TVLRT, TVSSAS, and TVLSSAS. 
     
     
         4 . The method according to  claim 1 , wherein said first and second polypeptide chains further comprise at least a hinge domain and an Fc domain or portions thereof derived from human IgG1, IgG2 or IgG4. 
     
     
         5 . The method according to  claim 1 , wherein said antibody-derived domains VD1 and VD2 display an engineered disulfide bridge introducing a covalent bond between VD1 and VD2 and where said cysteines are introduced into framework region (FR) 4 in case of VL and framework region 2 in case of VH. 
     
     
         6 . The method according to  claim 1 , wherein said cell surface molecule induces the activation of immune cells, or is at least one selected from the group consisting of CD3γ, CD3δ, CD3ε, CD4, CD7, CD8, CD10, CD11 b, CD11c, CD14, CD16, CD18, CD22, CD25, CD28, CD32a, CD32b, CD33, CD41, CD41b, CD42a, CD42b, CD44, CD45RA, CD49, CD55, CD56, CD61, CD64, CD68, CD94, CD90, CD117, CD123, CD125, CD134, CD137, CD152, CD163, CD193, CD203c, CD235a, CD278, CD279, CD287, Nkp46, NKG2D, GITR, FcεRI, TCRα/β, TCRγ/δ, and HLA-DR. 
     
     
         7 . A method of treating a patient with cancer, comprising administering to a patient a polypeptide molecule comprising a first polypeptide chain and a second polypeptide chain,
 wherein the first polypeptide chain comprises a first hinge domain and/or a first Fc domain, wherein the first polypeptide chain comprises at least 95% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 43, SEQ ID NO: 45, SEQ ID NO: 51, SEQ ID NO: 53, SEQ ID NO: 55, and SEQ ID NO: 57;   wherein the second polypeptide chain comprises a second hinge domain and/or a second Fc domain, wherein the second polypeptide chain comprises at least 95% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 52, SEQ ID NO: 54, SEQ ID NO: 56, and SEQ ID NO: 58; and   wherein the cancer is selected from the group consisting of bladder cancer, breast cancer, colon cancer, kidney cancer, liver cancer, lung cancer, ovarian cancer, pancreatic cancer, stomach cancer, prostate cancer, cervical cancer, melanoma, thyroid cancer, brain cancer, leukemia, sarcoma, and lymphoma.   
     
     
         8 . The method according to  claim 7 , wherein said first polypeptide chain is fused to said second polypeptide chain by covalent and/or non-covalent bonds between the first hinge domain and the second hinge domain, and/or between the first Fc domain and the second Fc domain. 
     
     
         9 . The method according to  claim 7 , wherein said first and second Fc domains each comprise at least one effector function silencing mutation. 
     
     
         10 . The method according to  claim 7 , wherein said first and second Fc domains each comprise a CH 3  domain comprising at least one mutation that facilitates the formation of heterodimers. 
     
     
         11 . The method according to  claim 7 , wherein said first and second Fc domains each comprise CH 2  and CH 3  domains comprising at least two additional cysteine residues. 
     
     
         12 . The method according to  claim 7 , wherein said cell surface molecule is at least one selected from the group consisting of CD3γ, CD3δ, CD3ε, CD4, CD7, CD8, CD10, CD11 b, CD11c, CD14, CD16, CD18, CD22, CD25, CD28, CD32a, CD32b, CD33, CD41, CD41b, CD42a, CD42b, CD44, CD45RA, CD49, CD55, CD56, CD61, CD64, CD68, CD94, CD90, CD117, CD123, CD125, CD134, CD137, CD152, CD163, CD193, CD203c, CD235a, CD278, CD279, CD287, Nkp46, NKG2D, GITR, FcεRI, TCRα/β, TCRγ/δ, and HLA-DR. 
     
     
         13 . The method according to  claim 7 ,
 wherein the first polypeptide chain comprises at least 99% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 43, SEQ ID NO: 45, SEQ ID NO: 51, SEQ ID NO: 53, SEQ ID NO: 55, and SEQ ID NO: 57; and   wherein the second polypeptide chain comprises at least 99% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 52, SEQ ID NO: 54, SEQ ID NO: 56, and SEQ ID NO: 58.   
     
     
         14 . A method of eliciting an immune response in a patient who has cancer, comprising administering to a patient a polypeptide molecule comprising a first polypeptide chain and a second polypeptide chain,
 wherein the first polypeptide chain comprises a first hinge domain and/or a first Fc domain, wherein the first polypeptide chain comprises at least 95% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 43, SEQ ID NO: 45, SEQ ID NO: 51, SEQ ID NO: 53, SEQ ID NO: 55, and SEQ ID NO: 57;   wherein the second polypeptide chain comprises a second hinge domain and/or a second Fc domain, wherein the second polypeptide chain comprises at least 95% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 52, SEQ ID NO: 54, SEQ ID NO: 56, and SEQ ID NO: 58; and   wherein the cancer is selected from the group consisting of bladder cancer, breast cancer, colon cancer, kidney cancer, liver cancer, lung cancer, ovarian cancer, pancreatic cancer, stomach cancer, prostate cancer, cervical cancer, melanoma, thyroid cancer, brain cancer, leukemia, sarcoma, and lymphoma.   
     
     
         15 . The method according to  claim 14 , wherein said first polypeptide chain is fused to said second polypeptide chain by covalent and/or non-covalent bonds between the first hinge domain and the second hinge domain, and/or between the first Fc domain and the second Fc domain. 
     
     
         16 . The method according to  claim 14 , wherein said first and second Fc domains each comprise at least one effector function silencing mutation. 
     
     
         17 . The method according to  claim 14 , wherein said first and second Fc domains each comprise a CH 3  domain comprising at least one mutation that facilitates the formation of heterodimers. 
     
     
         18 . The method according to  claim 14 , wherein said first and second Fc domains each comprise CH 2  and CH 3  domains comprising at least two additional cysteine residues. 
     
     
         19 . The method according to  claim 14 , wherein said cell surface molecule is at least one selected from the group consisting of CD3γ, CD3δ, CD3ε, CD4, CD7, CD8, CD10, CD11 b, CD11c, CD14, CD16, CD18, CD22, CD25, CD28, CD32a, CD32b, CD33, CD41, CD41b, CD42a, CD42b, CD44, CD45RA, CD49, CD55, CD56, CD61, CD64, CD68, CD94, CD90, CD117, CD123, CD125, CD134, CD137, CD152, CD163, CD193, CD203c, CD235a, CD278, CD279, CD287, Nkp46, NKG2D, GITR, FcεRI, TCRα/β, TCRγ/δ, and HLA-DR. 
     
     
         20 . The method according to  claim 14 ,
 wherein the first polypeptide chain comprises at least 99% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 43, SEQ ID NO: 45, SEQ ID NO: 51, SEQ ID NO: 53, SEQ ID NO: 55, and SEQ ID NO: 57; and   
       wherein the second polypeptide chain comprises at least 99% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 52, SEQ ID NO: 54, SEQ ID NO: 56, and SEQ ID NO: 58.