Activatable antibodies that bind epidermal growth factor receptor and methods of use thereof

Patent metadata

Field	Value
Publication number	US-2018333507-A1
Application number	US-201815875947-A
Country	US
Kind code	A1
Filing date	Jan 19, 2018
Priority date	Apr 27, 2012
Publication date	Nov 22, 2018
Grant date	—

Abstract

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 epidermal growth factor receptor (EGFR), and to methods of making and using these anti-EGFR activatable antibodies in a variety of therapeutic, diagnostic and prophylactic indications.

First claim

1 - 70 . (canceled)

71 . An activatable antibody that in an activated state binds Epidermal Growth Factor Receptor (EGFR) comprising:
(a) an antibody or an antigen binding fragment thereof (AB) that specifically binds to EGFR, wherein the AB comprises:
(i) heavy chain complementarity determining regions (HCDRs), wherein the HCDRs are the HCDRs of a heavy chain amino acid sequence selected from the group consisting of: SEQ ID NO: 26, SEQ ID NO: 30, and SEQ ID NO: 34,
(ii) light chain complementarity determining regions (LCDRs), wherein the LCDRs are the LCDRs of light chain amino acid sequence SEQ ID NO: 68, and
(iii) a heavy chain comprising an amino acid sequence with at least one of the following amino acid sequence features: (1) a glutamine residue at the position corresponding to position 88 of SEQ ID NO: 26, SEQ ID NO: 30, and SEQ ID NO: 34, and (2) an alanine residue at the position corresponding to position 299 of SEQ ID NO: 26, SEQ ID NO: 30, and SEQ ID NO: 34;
(b) a masking moiety (MM) coupled to the AB that inhibits the binding of the AB of the activatable antibody in an uncleaved state to EGFR; and (c) a cleavable moiety (CM) coupled to the AB, wherein the CM is a polypeptide that functions as a substrate for a protease, wherein the activatable antibody in the uncleaved state has the structural arrangement from N-terminus to C-terminus as follows: MM-CM-AB or AB-CM-MM.

72 . The activatable antibody of claim 71 , wherein
the HCDRs of the AB comprise the amino acid sequences NYGVH, VIWSGGNTDYNTPFTS, and ALTYYDYEFAY; and the LCDRs of the AB comprise the amino acid sequences RASQSIGTNIH, YASESIS, and QQNNNWPTT.

73 . The activatable antibody of claim 71 , wherein the AB comprises:
(i) a heavy chain variable region, wherein the heavy chain variable region is the heavy chain variable region of a heavy chain amino acid sequence selected from the group consisting of SEQ ID NO: 26 and SEQ ID NO: 34, (ii) a light chain variable region, wherein the light chain variable region is the light chain variable region of light chain amino acid sequence SEQ ID NO: 68.

74 . The activatable antibody of claim 71 , wherein the antigen binding fragment thereof is selected from the group consisting of a Fab fragment, a F(ab′) 2 fragment, a scFv, and a scAb.

75 . The activatable antibody of claim 71 , wherein the activatable antibody comprises a combination of heavy and light chains selected from the group consisting of:
(a) a heavy chain comprising amino acid sequence of SEQ ID NO: 26 and a light chain comprising the amino acid sequence of SEQ ID NO: 28; (b) a heavy chain comprising amino acid sequence of SEQ ID NO: 2 and a light chain comprising the amino acid sequence of SEQ ID NO: 4; (c) a heavy chain comprising amino acid sequence of SEQ ID NO: 10 and a light chain comprising the amino acid sequence of SEQ ID NO: 4; (d) a heavy chain comprising amino acid sequence of SEQ ID NO: 34 and a light chain comprising the amino acid sequence of SEQ ID NO: 28; (e) a heavy chain comprising amino acid sequence of SEQ ID NO: 26 and a light chain comprising the amino acid sequence of SEQ ID NO: 68; and (f) a heavy chain comprising amino acid sequence of SEQ ID NO: 34 and a light chain comprising the amino acid sequence of SEQ ID NO: 68.

76 . The activatable antibody of claim 71 , wherein the activatable antibody comprises a heavy chain comprising amino acid sequence of SEQ ID NO: 26 and a light chain comprising the amino acid sequence of SEQ ID NO: 28.

77 . The activatable antibody of claim 71 , wherein the AB has an equilibrium dissociation constant of about 100 nM or less for binding to EGFR.

78 . The activatable antibody of claim 71 , wherein the MM has one or more of the characteristics selected from the group consisting of:
(i) the MM has an equilibrium dissociation constant for binding to the AB which is greater than the equilibrium dissociation constant of the AB to EGFR, (ii) the MM does not interfere or compete with the AB of the activatable antibody in a cleaved state for binding to EGFR, (iii) the MM is a polypeptide of no more than 40 amino acids in length, (iv) the MM polypeptide sequence is different from that of EGFR, (v) the MM polypeptide sequence is no more than 50% identical to any natural binding partner of the AB, (vi) the MM does not comprise more than 25% amino acid sequence identity to EGFR, and (vii) the MM does not comprise more than 10% amino acid sequence identity to EGFR.

79 . The activatable antibody of claim 71 , wherein the MM comprises the amino acid sequence CISPRGCPDGPYVMY (SEQ ID NO: 14);

80 . The activatable antibody of claim 71 , wherein the CM is a polypeptide of up to 15 amino acids in length.

81 . The activatable antibody of claim 71 , wherein the protease is co-localized with EGFR in a tissue, and wherein the protease cleaves the CM in the activatable antibody when the activatable antibody is exposed to the protease.

82 . The activatable antibody of claim 71 , wherein the CM comprises the amino acid sequence LSGRSDNH (SEQ ID NO: 13).

83 . The activatable antibody of claim 71 , wherein the activatable antibody comprises a linking peptide between the MM and the CM, a linking peptide between the CM and the AB, or both a linking peptide between the MM and the CM and a linking peptide between the CM and the AB.

84 . The activatable antibody of claim 71 , wherein the activatable antibody comprises a first linking peptide (LP1) and a second linking peptide (LP2), and wherein the activatable antibody in an uncleaved state has the structural arrangement from N-terminus to C-terminus as follows: MM-LP1-CM-LP2-AB or AB-LP2-CM-LP1-MM.

85 . The activatable antibody of claim 84 , wherein the two linking peptides need not be identical to each other.

86 . The activatable antibody of claim 84 , 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: 15) and (GGGS) n (SEQ ID NO: 16), where n is an integer of at least one.

87 . The activatable antibody of claim 84 , wherein at least one of LP1 or LP2 comprises an amino acid sequence selected from the group consisting of GGSG (SEQ ID NO: 17), GGSGG (SEQ ID NO: 18), GSGSG (SEQ ID NO: 19), GSGGG (SEQ ID NO: 20), GGGSG (SEQ ID NO: 21), and GSSSG (SEQ ID NO: 22).

88 . The activatable antibody of claim 84 , wherein LP1 comprises the amino acid sequence GSSGGSGGSGGSG (SEQ ID NO: 23).

89 . The activatable antibody of claim 84 , wherein LP2 comprises the amino acid sequence GSSGT (SEQ ID NO: 24) or GSSG (SEQ ID NO: 37).

90 . The activatable antibody of claim 71 , wherein the activatable antibody in an uncleaved state 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.

91 . The activatable antibody of claim 90 , wherein the spacer comprises the amino acid sequence QGQSGQ (SEQ ID NO: 38).

92 . The activatable antibody of claim 90 , wherein the spacer and MM comprises the amino acid sequence QGQSGQCISPRGCPDGPYVMY (SEQ ID NO: 59).

93 . The activatable antibody of claim 71 , wherein the AB is conjugated to an agent.

94 . The activatable antibody of claim 93 , wherein the agent has one or more of the characteristics selected from the group consisting of:
(a) the agent is a toxin or fragment thereof, (b) the agent is a dolastatin

Assignees

• Cytomx Therapeutics Inc

Inventors

• Lowman Henry Bernard
• Desnoyers Luc Roland
• Liu Shouchun
• West James William
• Sagert Jason Gary
• Vasiljeva Olga
• Menendez Elizabeth-Edna Mary

Classifications

• A61P35/00

  Antineoplastic agents · CPC title

• A61P37/02

  Immunomodulators · CPC title

• A61P29/00

  Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID] · CPC title

• A61P17/06

  Antipsoriatics · CPC title

• G01N33/5759

  involving compounds localised on the membrane of tumour or cancer cells · CPC title