ACTIVATABLE ANTIBODIES THAT BIND INTERLEUKIN-6 RECEPTOR AND METHODS OF USE THEREOF
The invention relates generally to activatable antibodies that include a masking moiety (MM), a cleavable moiety (CM), and an antibody (AB) that specifically binds to interleukin-6 receptor (IL-6R), and to methods of making and using these anti-IL-6R activatable antibodies in a variety of therapeutic, diagnostic and prophylactic indications.
1 - 39 . (canceled)
40 . A method of preventing, delaying the progression of,
treating, alleviating a symptom of, or otherwise ameliorating inflammation or an inflammatory disorder comprising:
administering a therapeutically effective amount of an activatable antibody to a subject in need thereof, wherein the activatable antibody comprises:
an antibody or an antigen binding fragment thereof (AB) that specifically binds to IL-6R;
a masking moiety (MM) that inhibits the binding of the AB to IL-6R in an uncleaved state; and
a cleavable moiety (CM) coupled to the AB, wherein the CM is a polypeptide that functions as a substrate for a protease.
41 . (canceled)
42 . A method of treating, preventing, delaying the progression of, alleviating a symptom of, or otherwise ameliorating an IL-6R-mediated disorder or disease comprising:
administering a therapeutically effective amount of an activatable antibody to a subject in need thereof, wherein the activatable antibody comprises:
an antibody or an antigen binding fragment thereof (AB) that specifically binds to IL-6R;
a masking moiety (MM) that inhibits the binding of the AB to IL-6R in an uncleaved state; and
a cleavable moiety (CM) coupled to the AB, wherein the CM is a polypeptide that functions as a substrate for a protease.
43 . A method of preventing, delaying the progression of, treating, alleviating a symptom of, or otherwise ameliorating an autoimmune disease or disorder in a subject comprising:
administering a therapeutically effective amount of an activatable antibody to a subject in need thereof, wherein the activatable antibody comprises:
an antibody or an antigen binding fragment thereof (AB) that specifically binds to IL-6R;
a masking moiety (MM) that inhibits the binding of the AB to IL-6R in an uncleaved state; and
a cleavable moiety (CM) coupled to the AB, wherein the CM is a polypeptide that functions as a substrate for a protease.
44 . The activatable antibody of claim 40 , wherein the AB comprises
a VH CDR1 sequence that comprises the VH CDR1 sequence of SEQ ID NO: 1,
a VH CDR2 sequence that comprises the VH CDR2 sequence of SEQ ID NO: 1,
a VH CDR3 sequence that comprises the VH CDR3 sequence of SEQ ID NO: 1,
a VL CDR1 sequence that comprises the VL CDR1 sequence of SEQ ID NO: 2,
a VL CDR2 sequence that comprises the VL CDR2 sequence of SEQ ID NO: 2, and
a VL CDR3 sequence that comprises the VL CDR3 sequence of SEQ ID NO: 2.
45 . The method of claim 40 , wherein the AB comprises
a VH CDR1 sequence that comprises the amino acid sequence SDHAWS (SEQ ID NO: 175);
a VH CDR2 sequence that comprises the amino acid sequence YISYSGITTYNPSLKSRVT (SEQ ID NO: 176);
a VH CDR3 sequence that comprises the amino acid sequence SLARTTAMDY (SEQ ID NO: 177);
a VL CDR1 sequence that comprises the amino acid sequence RASQDISS (SEQ ID NO: 178);
a VL CDR2 sequence that comprises the amino acid sequence TISSLQP (SEQ ID NO: 179); and
a VL CDR3 sequence that comprises the amino acid sequence QQGNTLPY (SEQ ID NO: 180).
46 . The method of claim 40 , wherein the activatable antibody has the structural arrangement from N-terminus to C-terminus as follows in the uncleaved state: MM-CM-AB or AB-CM-MM.
47 . The method of claim 40 , wherein the activatable antibody comprises a linking peptide between the MM and the CM.
48 . The method of claim 40 , wherein the activatable antibody comprises a linking peptide between the CM and the AB.
49 . The method of claim 40 , wherein the activatable antibody comprises a first linking peptide (LP1) and a second linking peptide (LP2), and wherein the activatable antibody has the structural arrangement from N-terminus to C-terminus as follows in the uncleaved state: MM-LP1-CM-LP2-AB or AB-LP2-CM-LP1-MM.
50 . The method of claim 49 , wherein the two linking peptides need not be identical to each other.
51 . The method of claim 49 , wherein at least one of LP1 or LP2 comprises an amino acid sequence selected from the group consisting of (GS) n , (GGS) n , (GSGGS) n (SEQ ID NO: 93) and (GGGS) n (SEQ ID NO: 94), where n is an integer of at least one.
52 . The method of claim 49 , wherein at least one of LP1 or LP2 comprises an amino acid sequence selected from the group consisting of GGSG (SEQ ID NO: 95), GGSGG (SEQ ID NO: 96), GSGSG (SEQ ID NO: 97), GSGGG (SEQ ID NO: 98), GGGSG (SEQ ID NO: 99), and GSSSG (SEQ ID NO: 100).
53 . The method of claim 49 , wherein LP1 comprises the amino acid sequence GSSGGSGGSGGSG (SEQ ID NO: 101), GSSGGSGGSGG (SEQ ID NO: 112), GSSGGSGGSGGS (SEQ ID NO: 113), GSSGGSGGSGGSGGGS (SEQ ID NO: 169), GSSGGSGGSG (SEQ ID NO: 170), or GSSGGSGGSGS (SEQ ID NO: 171).
54 . The method of claim 49 , wherein LP2 comprises the amino acid sequence GSS, GGS, GGGS (SEQ ID NO: 172), GSSGT (SEQ ID NO: 102) or GSSG (SEQ ID NO: 103).
55 . The method of claim 49 , wherein the AB has an equilibrium dissociation constant of about 100 nM or less for binding to IL-6R.
56 . The method of claim 40 , wherein the antigen binding fragment thereof is selected from the group consisting of a Fab fragment, a F(ab′)2 fragment, a scFv, a scAb, a dAb, a single domain heavy chain antibody, and a single domain light chain antibody.
57 . The method of claim 40 , wherein the AB comprises a heavy chain variable region comprising the variable region of the heavy chain amino acid sequence SEQ ID NO: 1.
58 . The method of claim 40 , wherein the AB comprises a light chain variable region comprising the variable region of the light chain amino acid sequence SEQ ID NO: 2.
59 . The method of claim 40 , wherein the AB comprises a heavy chain variable region comprising the variable region of the heavy chain amino acid sequence SEQ ID NO: 1 and a light chain variable region comprising the variable region of the light chain amino acid sequence SEQ ID NO: 2.
60 . The method of claim 40 , wherein the AB comprises a heavy chain amino acid sequence comprising SEQ ID NO: 1 and a light chain amino acid sequence comprising SEQ ID NO: 2.
61 . The method of claim 40 , wherein the activatable antibody comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 6-32, 109-111, 163-168, 181, and 182.
62 . The method of claim 40 , wherein the activatable antibody comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 6-32, and 163-168.
63 . The method of claim 40 , wherein the MM has an equilibrium dissociation constant for binding to the AB which is greater than the equilibrium dissociation constant of the AB to IL-6R.
64 . The method of claim 40 , wherein the MM does not interfere or compete with the AB for binding to IL-6R in a cleaved state.
65 . The method of claim 40 , wherein the MM is a polypeptide of up to 40 amino acids in length.
66 . The method of claim 40 , wherein the MM polypeptide sequence is different from that of IL-6R.
67 . The method of claim 40 , wherein the MM polypeptide sequence is no more than 50% identical to any natural binding partner of the AB.
68 . The method of claim 40 , wherein the MM comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 33-89.
69 . The method of claim 40 , wherein the protease is co-localized with IL-6R in a tissue, and wherein the protease cleaves the CM in the activatable antibody when the activatable antibody is exposed to the protease.
70 . The method of claim 40 , wherein the CM is a polypeptide of up to 15 amino acids in length.
71 . The method of claim 40 , wherein the CM is a substrate for an enzyme selected from the group consisting of a matrix metalloprotease (MMP), thrombin, a neutrophil elastase, a cysteine protease, legumain, matriptase, and uPA.
72 . The method of claim 40 , wherein the CM comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 90-92, 104, 105, 107, 116-128, 157-162, 173, 174, and 183-193.
73 . The method of claim 40 , wherein the activatable antibody comprises a spacer, wherein the spacer is joined directly to the MM and has the structural arrangement from N-terminus to C-terminus of spacer-MM-CM-AB.
74 . The method of claim 40 comprising an agent conjugated to the AB.
75 . The method of claim 74 , wherein the agent is a therapeutic agent, an antineoplastic agent, or a toxin or fragment thereof.
76 . The method of claim 74 , wherein the agent is conjugated to the AB via a linker.
77 . The method of claim 76 , wherein the linker is a cleavable linker.
78 . The method of claim 40 , wherein the activatable antibody comprises a detectable moiety.
79 . The method of claim 78 , wherein the detectable moiety is a diagnostic agent.