Anti-PACAP antibodies and uses thereof
The present invention is directed to antibodies and antigen binding fragments thereof having binding specificity for PACAP. The antibodies and antigen binding fragments thereof comprise the sequences of the V H , V L , and CDR polypeptides described herein, and the polynucleotides encoding them. Antibodies and antigen binding fragments described herein bind to and/or compete for binding to the same linear or conformational epitope(s) on human PACAP as an anti-PACAP antibody. The invention contemplates conjugates of anti-PACAP antibodies and binding fragments thereof conjugated to one or more functional or detectable moieties. Methods of making said anti-PACAP antibodies and antigen binding fragments thereof are also contemplated. Other embodiments of the invention contemplate using anti-PACAP antibodies, and binding fragments thereof, for the diagnosis, assessment, and treatment of diseases and disorders associated with PACAP and conditions where antagonism of PACAP-related activities, such as vasodilation, photophobia, mast cell degranulation, and/or neuronal activation, would be therapeutically beneficial.
1. A humanized anti-pituitary adenylate cyclase-activating peptide (anti-PACAP) antibody or antigen-binding antibody fragment thereof, comprising: (i) a heavy chain variable region (V H ) comprising a complementarity-determining region 1 (CDR1) sequence consisting of the amino acid sequence of SEQ ID NO: 1164, a complementarity-determining region 2 (CDR2) sequence consisting of the amino acid sequence of SEQ ID NO: 1166, and a complementarity-determining region 3 (CDR3) sequence consisting of the amino acid sequence of SEQ ID NO: 1168; and (ii) a light chain variable region (VL) comprising a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1184, a CDR2 sequence consisting of the amino acid sequence of SEQ ID NO: 1186, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1188, wherein: (i) the V H comprises an amino acid sequence with at least 85% sequence identity to the amino acid sequence of SEQ ID NO: 1162, and (ii) the VL comprises an amino acid sequence with at least 85% sequence identity to the amino acid sequence of SEQ ID NO: 1182, wherein said antibody or antigen-binding antibody fragment thereof binds to an epitope which comprises residues 7, 10, 12, 13, 14 and 17 of human PACAP38 comprising the amino acid sequence of SEQ ID NO: 1241 or of human PACAP27 comprising the amino acid sequence of SEQ ID NO: 1242, and optionally wherein said epitope is identified by alanine scanning.
2. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , wherein:
(i) the V H comprises an amino acid sequence with at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1162 and comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1164, a CDR2 sequence consisting of SEQ ID NO: 1166, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1168; and
(ii) the VL comprises an amino acid sequence with at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1182 and comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1184, a CDR2 sequence consisting of the amino acid sequence of SEQ ID NO: 1186, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1188.
3. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , wherein:
(i) the V H comprises an amino acid sequence with at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 1162 and comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1164, a CDR2 sequence consisting of SEQ ID NO: 1166, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1168; and
(ii) the VL comprises an amino acid sequence with at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 1182 and comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1184, a CDR2 sequence consisting of the amino acid sequence of SEQ ID NO: 1186, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1188.
4. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , wherein:
(i) the V H comprises an amino acid sequence with at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 1162 and comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1164, a CDR2 sequence consisting of SEQ ID NO: 1166, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1168; and
(ii) the VL comprises an amino acid sequence with at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 1182 and comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1184, a CDR2 sequence consisting of the amino acid sequence of SEQ ID NO: 1186, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1188.
5. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , wherein:
(i) the V H comprises the amino acid sequence of SEQ ID NO: 1162; and
(ii) the VL comprises the amino acid sequence of SEQ ID NO: 1182.
6. The humanized anti-PACAP antibody according to claim 1 , comprising:
(i) a heavy chain comprising an amino acid sequence with at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 1161; and
(ii) a light chain comprising an amino acid sequence with at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 1181,
wherein:
(i) the heavy chain comprises said V H , and said V H comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1164, a CDR2 sequence consisting of SEQ ID NO: 1166, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1168; and
(ii) the light chain comprises said VL, and said VL comprises a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1184, a CDR2 sequence consisting of the amino acid sequence of SEQ ID NO: 1186, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1188.
7. The humanized anti-PACAP antibody according to claim 6 , wherein:
(i) the heavy chain comprises the amino acid sequence of SEQ ID NO: 1161; and
(ii) the light chain comprises the amino acid sequence of SEQ ID NO: 1181.
8. The humanized anti-PACAP antibody according to claim 1 , comprising a human IgG constant domain, optionally wherein the human IgG is IgG1, IgG2, IgG3, or IgG4.
9. The humanized anti-PACAP antibody according to claim 1 , wherein the antibody comprises a Fc region comprising the amino acid sequence of SEQ ID NO: 1244, 1245, or 1246.
10. The humanized anti-PACAP antibody according to claim 1 , wherein the antibody comprises an Fc region which
(i) lacks N-glycosylation;
(ii) lacks O-glycosylation; or
(iii) lacks N-glycosylation and O-glycosylation.
11. The humanized anti-PACAP antigen-binding antibody fragment thereof according to claim 1 , wherein the antigen-binding antibody fragment is selected from the group consisting of scFvs, fragment antigen-binding (Fab) fragments, Fab′ fragments, monovalent antigen-binding antibody fragments, and F(ab′) 2 fragments, wherein said antigen-binding antibody fragments comprise a heavy chain variable region (V H ) and a light chain variable region (VL) comprising complementarity-determining regions according to claim 1 .
12. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , which binds to adenylate cyclase-activating peptide (PACAP) with:
(i) a dissociation constant (K D ) of less than or equal to 5×10 −10 M, optionally wherein the KD is determined by ELISA, bio-layer interferometry, kinetic exclusion assay, or surface plasmon resonance at 25° or 37° C.; or
(ii) an off-rate (kd) of less than or equal to 5×10 −4 s −1 .
13. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , wherein the K D of said antibody or antigen-binding antibody fragment thereof for PACAP is at least 10-fold lower than the K D of said antibody or antigen-binding antibody fragment thereof for Vasoactive Intestinal Peptide (VIP).
14. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , which inhibits at least one biological effect elicited by PACAP, wherein said at least one biological effect is:
(i) PACAP-induced cAMP production;
(ii) PACAP-induced vasodilation in a subject administered with said antibody or antigen-binding antibody fragment; or
(iii) PACAP-induced photophobia in a subject administered with said antibody or antigen-binding antibody fragment thereof, optionally wherein said antibody or antigen-binding antibody fragment thereof inhibits PACAP binding to PAC1-receptor expressing cells.
15. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , which when administered to a human subject having a PACAP-mediated condition associated with increased vasodilation and/or photophobia reduces said PACAP-mediated vasodilation and/or photophobia.
16. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , which:
(i) lacks N-glycosylation;
(ii) lacks O-glycosylation; or
(iii) lacks N-glycosylation and O-glycosylation.
17. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 , which is directly or indirectly attached to a detectable label.
18. The humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 17 , wherein the detectable label is a fluorescent dye, enzyme, substrate, bioluminescent material, radioactive material, chemiluminescent moiety, or mixture thereof.
19. A pharmaceutical composition comprising:
(a) at least one humanized anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 1 ; and
(b) a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, or mixture thereof; and optionally
(c) another active agent, optionally a chemotherapeutic, an analgesic, an anti-inflammatory agent, an immunosuppressant, a cytokine, an antiproliferative agent, an antiemetic, or a cytotoxin, and
optionally wherein said composition is lyophilized, stabilized, or formulated for administration by injection.
20. An anti-PACAP antibody or antigen-binding antibody fragment thereof, comprising:
(i) a V H comprising a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1164, a CDR2 sequence consisting of SEQ ID NO: 1166, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1168; and
(ii) a VL comprising a CDR1 sequence consisting of the amino acid sequence of SEQ ID NO: 1184, a CDR2 sequence consisting of the amino acid sequence of SEQ ID NO: 1186, and a CDR3 sequence consisting of the amino acid sequence of SEQ ID NO: 1188.
21. The anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 20 , wherein the V H comprises the amino acid sequence of SEQ ID NO: 1162, and the VL comprises the amino acid sequence of SEQ ID NO: 1182.
22. An anti-PACAP antibody comprising:
(i) a heavy chain comprising the amino acid sequence of SEQ ID NO: 1161; and
(ii) a light chain comprising the amino acid sequence of SEQ ID NO: 1181.
23. The anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 20 , wherein the antigen-binding antibody fragment is selected from the group consisting of scFvs, Fab fragments, Fab′ fragments, monovalent antigen-binding antibody fragments, and F(ab′) 2 fragments, wherein said antigen-binding antibody fragments comprise a heavy chain variable region (V H ) and a light chain variable region (VL) comprising complementarity-determining regions according to claim 20 .
24. A pharmaceutical composition comprising:
(a) at least one anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 20 ; and
(b) a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, or mixture thereof; and optionally
(c) another active agent, optionally a chemotherapeutic, an analgesic, an anti-inflammatory agent, an immunosuppressant, a cytokine, an antiproliferative a ent, an antiemetic or a cytotoxin, and
optionally wherein said composition is lyophilized, stabilized, or formulated for administration by injection.
25. A pharmaceutical composition comprising:
(a) at least one anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 21 ; and
(b) a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, or mixture thereof; and optionally
(c) another active agent, optionally a chemotherapeutic, an analgesic, an anti-inflammatory agent, an immunosuppressant, a cytokine, an antiproliferative agent, an antiemetic, or a cytotoxin, and
optionally wherein said composition is lyophilized, stabilized, or formulated for administration by injection.
26. A pharmaceutical composition comprising:
(a) at least one anti-PACAP antibody or antigen-binding antibody fragment thereof according to claim 23 ; and
(b) a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, or mixture thereof; and optionally
(c) another active agent, optionally a chemotherapeutic, an analgesic, an anti-inflammatory, an immunosuppressant, a cytokine, an antiproliferative agent, an antiemetic, or a cytotoxin, and
optionally wherein said composition is lyophilized, stabilized, or formulated for administration by injection.
27. A pharmaceutical composition comprising:
(a) at least one anti-PACAP antibody according to claim 22 ; and
(b) a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, or mixture thereof; and optionally
(c) another active agent, optionally a chemotherapeutic, an analgesic, an anti-inflammatory agent, an immunosuppressant, a cytokine, an antiproliferative agent, an antiemetic, or a cytotoxin, and
optionally wherein said composition is lyophilized, stabilized, or formulated for administration by injection.