Fusion protein comprising apelin and an anti-APLNR antibody

Patent metadata

Field	Value
Publication number	US-10947310-B2
Application number	US-201816218862-A
Country	US
Kind code	B2
Filing date	Dec 13, 2018
Priority date	Nov 20, 2013
Publication date	Mar 16, 2021
Grant date	Mar 16, 2021

Abstract

The present invention provides apelin receptor (APLNR) modulators that bind to APLNR and methods of using the same. The invention includes APLNR modulators such as antibodies, or antigen-binding fragments thereof, which inhibit or attenuate APLNR-mediated signaling. The invention includes APLNR modulators such as antibodies, or antibody fusion proteins thereof, that activate APLNR-mediated signaling. According to certain embodiments of the invention, the antibodies or antigen-binding fragments or antibody fusion proteins are fully human antibodies that bind to human APLNR with high affinity. The APLNR modulators of the invention are useful for the treatment of diseases and disorders associated with APLNR signaling and/or APLNR cellular expression, such as cardiovascular diseases, angiogenesis diseases, metabolic diseases and fibrotic diseases.

First claim

What is claimed is:

1. A protein comprising (i) an immunoglobulin (Ig) molecule or antigen-binding fragment thereof and (ii) an apelin peptide, wherein the Ig molecule comprises a heavy chain variable region (HCVR), and a light chain variable region (LCVR), wherein:
(a) the HCVR and LCVR together comprise six complementarity determining regions HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3 contained with an HCVR/LCVR amino acid sequence pair selected from the group consisting of SEQ ID NOs: 2/10, 18/26, 34/42, 50/58, 66/74, 82/90, 98/106, 114/122, 130/138, 146/154, 162/170, 178/186, 194/202, 210/218, 287/295, 303/311, 319/327, 335/343, 351/359 and 367/375; and
(b) the apelin peptide:
(i) comprises an amino acid sequence selected from the group consisting of: SEQ ID NO: 227, SEQ ID NO: 228, SEQ ID NO: 229, SEQ ID NO: 230, SEQ ID NO: 262, SEQ ID NO: 269, SEQ ID NO: 270, SEQ ID NO: 271, SEQ ID NO: 272, SEQ ID NO: 273, SEQ ID NO: 283, SEQ ID NO: 284, and SEQ ID NO: 285; or
(ii) is selected from the group consisting of amino acid residues 6-77, 40-77, 42-77, 43-77, 47-77, 59-77, 61-77, 63-77, 64-77, 65-77, 66-77, 67-77, 73-77, 1-25, 6-25, 42-64, 61-64, 61-74, 61-75, 61-76, 65-76, 65-75, 42-58, 42-57, 42-56, 42-55, 42-54, 42-53, and pyroglutamylated apelin65-77 of the preproapelin polypeptide of SEQ ID NO: 227; or
(iii) is selected from the group consisting of: apelin40-77 (apelin-38), apelin42-77 (apelin-36), apelin43-77 (apelin-35), apelin47-77 (apelin-31), apelin59-77 (apelin-19), apelin61-77 (apelin-17), apelin63-77 (apelin-15), apelin64-77 (apelin-14), apelin65-77 (apelin-13), apelin66-77 (apelin-12, or A12), apelin67-77 (apelin-11), apelin68-77 (apelin-10), apelin73-77 (apelin-5), apelin61-76 (apelin-K16P), apelin61-75 (apelin-K15M), apelin61-74 (apelin-K14P), and [Pyr1]Apelin-13.

2. The protein of claim 1 , wherein the apelin peptide is fused to the Ig molecule at the N or the C terminus of the Ig molecule.

3. The protein of claim 1 , wherein the apelin peptide is fused to the Ig molecule at the N terminus or the C terminus of the HCVR.

4. The protein of claim 1 , wherein the apelin peptide is fused to the Ig molecule at the N terminus or the C terminus of the LCVR.

5. The protein of claim 1 , wherein the CDRs of the HCVR/LCVR amino acid sequence pair are contiguous.

6. The protein of claim 1 , wherein the protein activates apelin receptor (APLNR)-mediated inhibition of cAMP accumulation.

7. The protein of claim 1 , wherein the protein decreases apelin receptor (APLNR)-mediated inhibition of cAMP with an EC50 of less than about 100 nM as measured in an in vitro cAMP accumulation assay.

8. The protein of claim 1 , wherein the protein decreases apelin receptor (APLNR)-mediated inhibition of cAMP with an EC50 of less than about 10 nM as measured in an in vitro cAMP accumulation assay.

9. The protein of claim 1 , wherein the protein activates apelin receptor (APLNR)-mediated pERK in an in vitro pERK assay.

10. The protein of claim 1 , wherein the protein increases apelin receptor (APLNR)-mediated pERK to total ERK ratio with an EC50 of less than about 10 nM as measured in an in vitro pERK assay.

11. The protein of claim 1 , wherein the protein increases apelin receptor (APLNR)-mediated pERK to total ERK ratio with an EC50 of less than about 1 nM as measured in an in vitro pERK assay.

12. The protein of claim 1 , wherein the protein comprises an amino acid sequence pair selected from the group consisting of: SEQ ID NOs: 130/235, 130/237, 239/138, 241/138, 243/138, 245/138, 247/122, 114/249, 114/251, 253/26, 255/26, 257/26, 259/26, 274/138, 275/138, 276/138, 277/138, 278/138, 279/26, 280/26, 281/26, and 282/26.

13. The protein of claim 1 , wherein the Ig molecule comprises a scFv comprising a HCVR/LCVR amino acid sequence pair selected from the group consisting of SEQ ID NOs: 2/10, 18/26, 34/42, 50/58, 66/74, 82/90, 98/106, 114/122, 130/138, 146/154, 162/170, 178/186, 194/202, 210/218, 287/295, 303/311, 319/327, 335/343, 351/359 and 367/375.

14. The protein of claim 13 , wherein the Ig molecule comprises a scFv comprising a HCVR/LCVR amino acid sequence pair selected from the group consisting of SEQ ID NOs: 34/42, 50/58, 66/74, 82/90, 287/295, 303/311, 319/327, 335/343, 351/359 and 367/375.

15. The protein of claim 1 , wherein the six complementarity determining regions HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3 comprise the amino acid sequences, respectively, selected from the group consisting of SEQ ID NOs: 4-6-8-12-14-16, 20-22-24-28-30-32, 36-38-40-44-46-48, 52-54-56-60-62-64, 68-70-72-76-78-80, 84-86-88-92-94-96, 100-102-104-108-110-112, 116-118-120-124-126-128, 132-134-136-140-142-144, 148-150-152-156-158-160, 164-166-168-172-174-176, 180-182-184-188-190-192, 196-198-200-204-206-208, 212-214-216-220-222-224, 289-291-293-297-299-301, 305-307-309-313-315-317, 321-323-325-329-331-333, 337-339-341-345-347-349, 353-355-357-361-363-365, and 369-371-373-377-379-381.

16. The protein of claim 15 , wherein the six complementarity determining regions HCDR1-HCDR2-HCDR3-LCDR1-LCDR2-LCDR3 comprise the amino acid sequences, respectively, selected from the group consisting of SEQ ID NOs: 36-38-40-44-46-48, 52-54-56-60-62-64, 68-70-72-76-78-80, 84-86-88-92-94-96, 289-291-293-297-299-301, 305-307-309-313-315-317, 321-323-325-329-331-333, 337-339-341-345-347-349, 353-355-357-361-363-365, and 369-371-373-377-379-381.

17. The protein of claim 1 , wherein the apelin peptide is fused to the Ig molecule or antigen-binding fragment thereof via a linker.

18. The protein of claim 17 , wherein the linker is a (G4S) n linker, wherein n is from 1 to 10.

19. The protein of claim 18 , wherein the linker is a (G4S) 3 linker.

20. A pharmaceutical composition comprising the protein of claim 1 , and a pharmaceutically acceptable carrier or diluent.

Assignees

• Regeneron Pharma

Inventors

• Stevis Panayiotis
• Murphy Andrew J
• Gromada Jesper
• Ray Yonaton
• Kim Jee H
• Lobov Ivan B

Classifications

• C07K16/28 Primary

  against receptors, cell surface antigens or cell surface determinants · CPC title

• C07K7/06

  having 5 to 11 amino acids · CPC title

• A61K38/08 Primary

  Peptides having 5 to 11 amino acids {(A61K38/043 - A61K38/046 take precedence)} · CPC title

• C07K19/00

  Hybrid peptides {, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes} · CPC title

• C07K16/46

  Hybrid immunoglobulins (hybrids of an immunoglobulin with a peptide not being an immunoglobulin C07K19/00) · CPC title