ANTI-LAG-3 ANTIBODIES AND METHODS OF USE THEREOF

• US 20180127499A1
• Filed: 10/11/2017
• Published: 05/10/2018
• Est. Priority Date: 10/11/2016
• Status: Active Grant

• Current Assignee: Agenus Inc.
• Original Assignee: Agenus Inc.
• Inventors: Wilson, Nicholas Stuart, Savitsky, David Adam, Jennings, Shawn Michael, van Dijk, Marc, Mundt, Cornelia Anne
• Granted Patent: US 10,844,119 B2
• Time in Patent Office: Days
• CPC Class Codes: A61K 47/6873 the antibody targeting an immunoglobulin; the antibody being an anti-idiotypic antibody
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/2803 against the immunoglobulin superfamily
• CPC Class Codes: C07K 2317/20 characterized by taxonomic origin
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/34 Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
• CPC Class Codes: C07K 2317/55 Fab or Fab'
• CPC Class Codes: C07K 2317/76 Antagonist effect on antigen, e.g. neutralization or inhibition of binding

Abstract

The instant disclosure provides antibodies that specifically bind to LAG-3 (e.g., human LAG-3) and antagonize LAG-3 function. Also provided are pharmaceutical compositions comprising these antibodies, nucleic acids encoding these antibodies, expression vectors and host cells for making these antibodies, and methods of treating a subject using these antibodies.

Claims

1. An isolated antibody that specifically binds to human LAG-3, the antibody comprising a heavy chain variable region comprising complementarity determining regions CDRH1, CDRH2 and CDRH3 and a light chain variable region comprising complementarity determining regions CDRL1, CDRL2 and CDRL3, wherein:

(a) CDRH1 comprises the amino acid sequence of DX₁YX₂X₃(SEQ ID NO;

     140), wherein

X₁ is T or N,

X₂ is I or M, and

X₃ is H, Y or D;

(b) CDRH2 comprises the amino acid sequence of X₁IDPANX₂X₃X₄X₅X₆X₇PX₅X₉QX₁₀ (SEQ ID NO;

     142), wherein

X₁ is E, R, S, or K,

X₂ is D or G,

X₃ is N or H,

X₄ is T or S,

X₅ is K or H,

X₆ is Y or F,

X₇ is D or A,

X₈ is K or R,

X₉ is F or L, and

X₁₀ is G or D;

(c) CDRH3 comprises the amino acid sequence of YX₁X₂X₃YX₄VGGX₅DY (SEQ ID NO;

     144), wherein

X₁ is Y, F, or S,

X₂ is Y or D,

X₃ is K or R,

X₄ is D or E, and

X₅ is F or C;

(d) CDRL1 comprises the amino acid sequence of SVSSX₁ISSSX₂LX₃ (SEQ ID NO;

     147), wherein

X₁ is S or G,

X₂ is N or T, and

X₃ is H or Y;

(e) CDRL2 comprises the amino acid sequence of GTSNLAS (SEQ ID NO;

     104); and

(f) CDRL3 comprises the amino acid sequence of QQWX₁X₂YPX₃T (SEQ ID NO;

     149), wherein

X₁ is S, N, or R,

X₂ is S, T or R, and

X₃ is F, L, H, or W.

2. The isolated antibody of claim 1, wherein:

(a) CDRH1 comprises the amino acid sequence of DX₁YX₂X₃(SEQ ID NO;

     141), wherein;

X₁ is T or N;

X₂ is I or M; and

X₃ is H or Y;

(b) CDRH2 comprises the amino acid sequence of X₁IDPANX₂X₃X₄KX₅X₆PX₇FQX₈ (SEQ ID NO;

     143), wherein;

X₁ is E, R, or S;

X₂ is D or G;

X₃ is N or H;

X₄ is T or S;

X₅ is Y or F;

X₆ is D or A;

X₇ is K or R; and

X₈ is G or D;

(c) CDRH3 comprises the amino acid sequence of YX₁X₂X₃YDVGGX₄DY (SEQ ID NO;

     145), wherein;

X₁ is Y, F, or S;

X₂ is Y or D;

X₃ is K or R; and

X₄ is F or C;

(d) CDRH3 comprises the amino acid sequence of YYYX₁YX₂VGGFDY (SEQ ID NO;

     146), wherein;

X₁ is K or R; and

X₂ is D or E;

(e) CDRL1 comprises the amino acid sequence of SVSSSISSSNLX₁(SEQ ID NO;

     148), wherein;

X₁ is H or Y; and

/or

(f) CDRL3 comprises the amino acid sequence of QQWX₁SYPX₂T (SEQ ID NO;

     150), wherein;

X₁ is S, N, or R; and

X₂ is F, L, or H.

9. The isolated antibody of claim 1, wherein:

(a) CDRH1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs;

78-82;

(b) CDRH2 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs;

83-93;

(c) CDRH3 comprises the amino acid sequence of YYYX₁YX₂VGGFDY (SEQ ID NO;

     146), wherein;

X₁ is K or R; and

X₂ is D or E;

(d) CDRH3 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs;

94-99;

(e) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs;

100-103; and

/or

(f) CDRL3 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs;

105-112.

14. The isolated antibody of claim 1, wherein CDRH1, CDRH2 and CDRH3 comprise the CDRH1, CDRH2 and CDRH3 amino acid sequences, respectively, set forth in SEQ ID NOs:

78, 83, and 94;

78, 85, and 95;

78, 86, and 96;

78, 86, and 97;

78, 91, and 94;

78, 92, and 96;

79, 84, and 95;

79, 88, and 95;

79, 89, and 95;

79, 90, and 95;

79, 90, and 98;

79, 90, and 99;

80, 85, and 96;

81, 87, and 96;

or, 82, 93, and 95, and/or CDRL1, CDRL2 and CDRL3 comprise the CDRL1, CDRL2 and CDRL3 amino acid sequences, respectively, set forth in SEQ ID NOs;

100, 104, and 105;

100, 104, and 106;

100, 104, and 107;

100, 104, and 109;

100, 104, and 110;

101, 104, and 108;

102, 104, and 105;

102, 104, and 112;

or, 103, 104, and 111.

16. The isolated antibody of claim 1, wherein CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences set forth in SEQ ID NOs:

78, 83, 94, 100, 104, and 105;

78, 85, 95, 100, 104, and 105;

78, 86, 96, 100, 104, and 105;

78, 86, 96, 100, 104, and 109;

78, 86, 96, 100, 104, and 110;

78, 86, 96, 101, 104, and 108;

78, 86, 96, 103, 104, and 111;

78, 86, 97, 102, 104, and 112;

78, 91, 94, 100, 104, and 107;

78, 92, 96, 100, 104, and 105;

78, 92, 96, 100, 104, and 109;

79, 84, 95, 100, 104, and 105;

79, 84, 95, 100, 104, and 106;

79, 84, 95, 102, 104, and 105;

79, 88, 95, 100, 104, and 105;

79, 89, 95, 100, 104, and 105;

79, 90, 95, 100, 104, and 105;

79, 90, 98, 100, 104, and 105;

79, 90, 99, 100, 104, and 105;

80, 85, 96, 100, 104, and 105;

81, 87, 96, 100, 104, and 105;

81, 87, 96, 100, 104, and 107;

or, 82, 93, 95, 100, 104, and 105, respectively.

20. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:

151, 222, 218, or 223, and/or a light chain variable region comprising the amino acid sequence of SEQ ID NO;

152 or 224.

21. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain variable region comprising an amino acid sequence which is at least 75%, 80%, 85%, 90%, 95%, or 100% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs:

56-72 and 220, and/or a light chain variable region comprising an amino acid sequence which is at least 75%, 80%, 85%, 90%, 95%, or 100% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs;

73-77 and 221.

24. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:

168-186 and 225-227, and/or a light chain comprising an amino acid sequence selected from the group consisting of SEQ ID NOs;

187-191 and 228.

26. The isolated antibody of claim 1, wherein the antibody comprises:

(a) a heavy chain variable framework region that is or is derived from an amino acid sequence encoded by a human gene, optionally wherein said amino acid sequence is selected from the group consisting of IGHV1-46*01 (SEQ ID NO;

     153), IGHV1-69-2*01 (SEQ ID NO;

     154), IGHV1-3*01 (SEQ ID NO;

     155), IGHV1-24*01 (SEQ ID NO;

     156), IGHV1-2*01 (SEQ ID NO;

     157), IGHV1-45*01 (SEQ ID NO;

     158), and IGHV1-18*01 (SEQ ID NO;

     159); and

/or

(b) a light chain variable framework region that is or is derived from an amino acid sequence encoded by a human gene, optionally wherein said amino acid sequence is selected from the group consisting of IGKV3-20*01 (SEQ ID NO;

     160), IGKV3D-15*01 (SEQ ID NO;

     161), IGKV3-15*01 (SEQ ID NO;

     161), IGKV3D-20*01 (SEQ ID NO;

     162), IGKV3D-7*01 (SEQ ID NO;

     163), IGKV1-9*01 (SEQ ID NO;

     164), and IGKV3-11*01 (SEQ ID NO;

     165).

27. The isolated antibody of claim 26, wherein the antibody comprises:

(a) a heavy chain variable framework region that is derived from the amino acid sequence IGHV1-46*01 (SEQ ID NO;

     153), wherein at least one amino acid in the amino acid sequence IGHV1-46*01 (SEQ ID NO;

     153) is substituted with an amino acid in an analogous position in a corresponding non-human heavy chain variable framework region, optionally wherein the amino acid substitution is at an amino acid position selected from the group consisting of 4, 5, 12, 23, 27, 28, 29, 30, 48, 69, 71, 75, 76, 80, 81, and 94, further optionally wherein the amino acid substitution is selected from the group consisting of 4M, 5K, 12V, 23T, 27F, 28N, 29I, 30K, 48I, 69I, 71A, 75S, 76N, 80L, 81Q, and 94T; and

/or

(b) a light chain variable framework region that is derived from the amino acid sequence IGKV3-20*01 (SEQ ID NO;

     160), wherein at least one amino acid in the amino acid sequence IGKV3-20*01 (SEQ ID NO;

     160) is substituted with an amino acid in an analogous position in a corresponding non-human light chain variable framework region, optionally wherein the amino acid substitution is at an amino acid position selected from the group consisting of 3, 22, 36, 43, 47, 58, 70, and 71, further optionally wherein the amino acid substitution is selected from the group consisting of 3L, 22T, 36F, 43S, 47W, 58V, 70S, and 71Y,

wherein the position of the amino acid substitution is indicated according to the Kabat numbering system.

51. The isolated antibody of claim 1, wherein the heavy chain variable region and the light chain variable region, respectively, comprise the amino acid sequences set forth in SEQ ID NOs:

56 and 73;

56 and 74;

56 and 75;

56 and 76;

56 and 77;

57 and 73;

57 and 74;

57 and 75;

57 and 76;

57 and 77;

58 and 73;

58 and 74;

58 and 75;

58 and 76;

58 and 77;

59 and 73;

59 and 74;

59 and 75;

59 and 76;

59 and 77;

60 and 73;

60 and 74;

60 and 75;

60 and 76;

60 and 77;

61 and 77;

62 and 77;

63 and 73;

64 and 73;

65 and 73;

220 and 73;

65 and 221;

220 and 221;

66 and 73;

67 and 73;

68 and 73;

69 and 73;

70 and 73;

71 and 73;

or 72 and 73.

53. The isolated antibody of claim 1, the antibody comprising:

(a) a heavy chain comprising the amino acid sequence of SEQ ID NO;

168 or 225, and a light chain comprising the amino acid sequence of SEQ ID NO;

187 or 228;

(b) a heavy chain comprising the amino acid sequence of SEQ ID NO;

169 or 226, and a light chain comprising the amino acid sequence of SEQ ID NO;

187 or 228;

or

(c) a heavy chain comprising the amino acid sequence of SEQ ID NO;

169 or 226, and a light chain comprising the amino acid sequence of SEQ ID NO;

187 or 228.

56. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain constant region selected from the group consisting of human IgG₁, IgG₂, IgG₃, IgG₄, IgA₁, and IgA₂, and optionally wherein the heavy chain constant region is:

(a) an IgG₁ heavy chain constant region comprising a N297A mutation, optionally wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO;

194;

(b) an IgG₁ heavy chain constant region comprising a N297Q mutation;

(c) a non-fucosylated IgG₁ heavy chain constant region; and

/or

(d) an IgG₄ heavy chain constant region comprises a S228P mutation, optionally wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO;

196,

wherein the amino acid residues are numbered according to the EU numbering system.

65. The isolated antibody of claim 1, wherein the antibody comprises a light chain constant region selected from the group consisting of human IgGκ

and IgGλ

, and optionally wherein the light chain constant region is an IgGκ

light chain constant region comprising the amino acid sequence of SEQ ID NO;

198 or 219.

68. An isolated antibody that cross-competes for binding to human LAG-3 with the antibody of claim 1.

69. An isolated antibody that binds to the same epitope of human LAG-3 as the antibody of claim 1.

76. The isolated antibody of claim 1, wherein the antibody:

(a) is a humanized antibody, murine antibody, or chimeric antibody;

(b) is antagonistic to human LAG-3

(c) deactivates, reduces, or inhibits an activity of human LAG-3;

(d) inhibits binding of human LAG-3 to MHC class II;

(e) induces IL-2 production by peripheral blood mononuclear cells (PBMCs) stimulated with staphylococcal enterotoxin A (SEA); and

/or

(f) induces TNFα

production by tumor infiltrating lymphocytes (TILs) stimulated with anti-CD3 and anti-CD28 antibodies.

82. The isolated antibody of claim 1 conjugated to a cytotoxic agent, cytostatic agent, toxin, radionuclide, or detectable label.

83. A pharmaceutical composition comprising the antibody of any one of claim 1 and a pharmaceutically acceptable carrier or excipient.

84. An isolated polynucleotide encoding a heavy and/or light chain of the antibody of claim 1.

85. A vector comprising the polynucleotide of claim 84.

86. A recombinant host cell comprising the polynucleotide of claim 84.

87. A method of producing an antibody that binds to human LAG-3, the method comprising culturing the host cell of claim 86 so that the polynucleotide is expressed and the antibody is produced.

88. A method of increasing T cell activation in response to an antigen in a subject, the method comprising administering to the subject an effective amount of the antibody of claim 1.

89. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of the antibody of claim 1.

3-8. -8. (canceled)

10-13. -13. (canceled)

15. (canceled)

17-19. -19. (canceled)

22-23. -23. (canceled)

25. (canceled)

28-50. -50. (canceled)

52. (canceled)

54-55. -55. (canceled)

57-64. -64. (canceled)

66-67. -67. (canceled)

70. An isolated antibody that specifically binds to human LAG-3, wherein the antibody binds to:

(a) an epitope located within a region of human LAG-3 consisting of the amino acid sequence of SEQ ID NO;

216;

(b) an epitope located within a region of human LAG-3 consisting of the amino acid sequence of SEQ ID NO;

215;

(c) an epitope located within a region of human LAG-3 consisting of the amino acid sequence of SEQ ID NO;

214;

(d) an epitope located within a region of human LAG-3 consisting of the amino acid sequence of SEQ ID NO;

213;

(e) an epitope located within a region of human LAG-3 consisting of the amino acid sequence of SEQ ID NO;

212; and

/or

(f) an epitope located within a region of human LAG-3 consisting of the amino acid sequence of SEQ ID NO;

211.

71-75. -75. (canceled)

77-81. -81. (canceled)

90-147. -147. (canceled)