Cleavable linker compositions and methods

Abstract

Provided herein are cleavable linkers, pharmaceutical compositions thereof, as well as nucleic acids, and methods for making and discovering the same. The cleavable linkers described herein have improved efficacy and safety.

1 - 30 . (canceled) 
     
     
         31 . An isolated polypeptide comprising a cleavable linker according to the amino acid sequence of SEQ ID NO: 4 (AAGLLAPPGGLSGRSDAG). 
     
     
         32 . The isolated polypeptide of  claim 31 , wherein the cleavable linker is cleavable by a protease. 
     
     
         33 . The isolated polypeptide of  claim 32 , wherein the protease comprises a tumor specific protease. 
     
     
         34 . The isolated polypeptide of  claim 32 , wherein the protease comprises a matrix metalloprotease (MMP) or a serine protease. 
     
     
         35 . The isolated polypeptide of  claim 34 , wherein the matrix metalloprotease comprises MMP2, MMP7, MMP9, MMP13, or MMP14. 
     
     
         36 . The isolated polypeptide of  claim 34 , wherein the serine protease comprises matriptase, urokinase, or hepsin. 
     
     
         37 . The isolated polypeptide of  claim 31 , wherein the isolated polypeptide further comprises an antigen binding domain that binds to a target antigen. 
     
     
         38 . The isolated polypeptide of  claim 37 , wherein the cleavable linker connects a peptide to the antigen binding domain that binds to the target antigen in a configuration according to Formula I: A 1 -L 1 -P 1  wherein A 1  comprises the antigen binding domain that binds to the target antigen; L 1  comprises the cleavable linker; P 1  comprises a peptide that impairs binding of the antigen binding domain to the target antigen; and wherein P 1  is connected C-terminal to L 1  and A 1  is connected N-terminal to L 1 , or P 1  is connected N-terminal to L 1  and A 1  is connected C-terminal to L 1 . 
     
     
         39 . The isolated polypeptide of  claim 38 , wherein P 1  is further linked to a half-life extending moiety, and wherein the half-life extending moiety is a single-domain antibody. 
     
     
         40 . The isolated polypeptide of  claim 38 , wherein A 1  comprises an antibody, a single chain variable fragment (scFv), a heavy chain variable domain (VH domain), a light chain variable domain (VL domain), a variable domain (VHH) of a camelid derived single domain antibody, a Fab, a Fab′, a Fab light chain polypeptide, or a Fab heavy chain polypeptide. 
     
     
         41 . The isolated polypeptide of  claim 40 , wherein A 1  comprises the scFv. 
     
     
         42 . The isolated polypeptide of  claim 41 , wherein the scFv comprises an anti-CD3e single chain variable fragment. 
     
     
         43 . A complex comprising the isolated polypeptide of  claim 37  and a second isolated polypeptide comprising a second antigen binding domain. 
     
     
         44 . The isolated polypeptide of  claim 43 , wherein the second isolated polypeptide is in a configuration according to Formula II: A 2 -L 2 -P 2  wherein A 2  comprises the second antigen binding domain; L 2  comprises a second cleavable linker; P 2  comprises a second peptide that impairs binding of the second antigen binding domain to a second target antigen; wherein the second antigen binding domain comprises a Fab light chain polypeptide or a Fab heavy chain polypeptide and the second target antigen comprises a tumor antigen. 
     
     
         45 . The isolated polypeptide of  claim 44 , wherein the second cleavable linker comprises the amino acid sequence of SEQ ID NO: 1 (LSGRSDAG). 
     
     
         46 . The isolated polypeptide of  claim 44 , wherein the second cleavable linker comprises the amino acid sequence of SEQ ID NO: 3 (ISSGLLSGRSDAG). 
     
     
         47 . The isolated polypeptide of  claim 44 , wherein the second cleavable linker comprises the amino acid sequence of SEQ ID NO: 26 (AGLLAPPGGLSGRSDAG). 
     
     
         48 . The isolated polypeptide of  claim 44 , wherein the second cleavable linker comprises the amino acid sequence of SEQ ID NO: 4 (AAGLLAPPGGLSGRSDAG). 
     
     
         49 . The isolated polypeptide of  claim 44 , wherein the second cleavable linker comprises the amino acid sequence of SEQ ID NO: 5 (SPLGLSGRSDAG). 
     
     
         50 . The isolated polypeptide of  claim 44 , wherein the second cleavable linker comprises the amino acid sequence of SEQ ID NO: 6 (LSGRSDAGSPLGLAG).