HK1203211B - Cgrp antibodies - Google Patents

Description

CGRP antibodies

This application is a divisional application of PCT application PCT/US2011/039381, filed on June 7, 2011, with the invention name “CGRP antibody”. The date on which the PCT application entered the Chinese national phase is December 10, 2012, with application number 201180028611.1.

The present invention is in the field of medicine, in particular in the field of antibodies to calcitonin gene-related peptide (CGRP). More specifically, the present invention relates to CGRP antibodies and the use of CGRP antibodies in the treatment of osteoarthritis pain or migraine.

Calcitonin gene-related peptide (CGRP) is a 37-amino acid neuropeptide secreted by nerves in the central and peripheral nervous systems. CGRP is widely distributed in sensory nerves in the peripheral and central nervous systems and exhibits a variety of different biological activities. CGRP is reported to have functions. When released from the trigeminal nerve and other nerve fibers, CGRP is believed to mediate its biological responses by binding to specific cell surface receptors.

CGRP has been reported to play a role in migraines because it is released when sensory nerves are stimulated and has potent vasodilatory activity. When these fibers are stimulated, the release of CGRP increases vascular permeability and subsequent leakage of plasma proteins (plasma protein extravasation) into tissues innervated by trigeminal nerve fibers. In addition, studies have reported that infusion of CGRP in patients with migraines results in migraine-like symptoms.

CGRP has also been reported to play a role in osteoarthritis (OA). OA is a chronic condition that affects a high percentage of the population worldwide. Arthritis pain is characterized by hyperalgesia (excessive sensitivity to normal, harmless stimuli) and spontaneous pain (pain at rest). Inflammation of the joints leads to peripheral and central sensitization. In peripheral sensitization, nociceptors containing CGRP, which are usually high threshold, begin to respond to the slight pressure associated with normal joint movement. CGRP acts as a local promoter of the inflammatory process. This ongoing process ultimately leads to central sensitization of the spinal cord, causing neurons to become responsive to stimulation of inflamed and non-inflamed tissues. In a preclinical model of OA pain in rats, an increase in the number of CGRP-positive nerve fibers was observed, which positively correlated with the amount of pain.

The current standard of care for OA pain consists of nonsteroidal anti-inflammatory drugs (NSAIDs) and selective Cox-2 inhibitors. Over time, many patients will become insensitive to these treatments or cannot tolerate the high doses required for effective pain relief. Subsequent treatment consists of intra-articular injections (hyaluronan) or opiates. The final treatment is surgical total joint replacement. Many of these treatments have negative side effects, ranging from gastrointestinal (NSAIDs) to cardiovascular (Cox-2 inhibitors), are cumbersome and/or less effective (intra-articular injections), have a risk of abuse (opiates), or are limited to only one joint (intra-articular injections, surgery).

Although CGRP antibodies have been disclosed for the treatment of migraine and for the treatment of inflammatory pain (migraine - WO 2007/076336 and WO 2007/054809; OA - see antibody G1 of WO 2009/109908A1), there remains a need for alternatives. Preferably, the therapeutic alternatives are safe and effective. More preferably, the alternative treatment for OA can provide long-term, sustained pain relief across multiple joints that is tolerable to patients and does not pose a risk of dependence and/or abuse.

The present invention provides a human engineered CGRP antibody comprising a light chain variable region (LCVR) and a heavy chain variable region (HCVR), wherein the LCVR comprises LCDR1, LCDR2, and LCDR3 amino acid sequences, and the HCVR comprises HCDR1, HCDR2, and HCDR3 amino acid sequences selected from the group consisting of:

a) LCDR1 is RASQDIDNYLN (SEQ ID NO:3), LCDR2 is YTSEYHS (SEQ ID NO:4), LCDR3 is QQGDALPPT (SEQ ID NO:5), HCDR1 is GYTFGNYWMQ (SEQ ID NO:12), HCDR2 is AIYEGTGDTRYIQKFAG (SEQ ID NO:13), and HCDR3 is LSDYVSGFSY (SEQ ID NO:14);

b) LCDR1 is RASQDIDNYLN (SEQ ID NO:3), LCDR2 is YTSEYHS (SEQ ID NO:4), LCDR3 is QQGDALPPT (SEQ ID NO:5), HCDR1 is GYTFGNYWMQ (SEQ ID NO:12), HCDR2 is AIYEGTGKTVYIQKFAG (SEQ ID NO:15), and HCDR3 is LSDYVSGFSY (SEQ ID NO:14);

c) LCDR1 is RASKDISKYLN (SEQ ID NO:6), LCDR2 is YTSGYHS (SEQ ID NO:7), LCDR3 is QQGDALPPT (SEQ ID NO:5), HCDR1 is GYTFGNYWMQ (SEQ ID NO:12), HCDR2 is AIYEGTGKTVYIQKFAD (SEQ ID NO:16), and HCDR3 is LSDYVSGFGY (SEQ ID NO:39);

d) LCDR1 is RASRPIDKYLN (SEQ ID NO:8), LCDR2 is YTSEYHS (SEQ ID NO:4), LCDR3 is QQGDALPPT (SEQ ID NO:5), HCDR1 is GYTFGNYWMQ (SEQ ID NO:12), HCDR2 is AIYEGTGKTVYIQKFAG (SEQ ID NO:15), and HCDR3 is LSDYVSGFGY (SEQ ID NO:39); and

e) LCDR1 is RASQDIDKYLN (SEQ ID NO: 9), LCDR2 is YTSGYHS (SEQ ID NO: 7), LCDR3 is QQGDALPPT (SEQ ID NO: 5), HCDR1 is GYTFGNYWMQ (SEQ ID NO: 12), HCDR2 is AIYEGTGKTVYIQKFAG (SEQ ID NO: 15), and HCDR3 is LSDYVSGFGY (SEQ ID NO: 39),

Among them, the human engineered CGRP antibody binds to human CGRP.

The present invention provides a human engineered CGRP antibody comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO: 3, LCDR2 is SEQ ID NO: 4, LCDR3 is SEQ ID NO: 5, HCDR1 is SEQ ID NO: 12, HCDR2 is SEQ ID NO: 13, and HCDR3 is SEQ ID NO: 14, wherein the human engineered CGRP antibody binds to human CGRP. The present invention provides a human engineered CGRP antibody or antigen-binding fragment thereof comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO: 3, LCDR2 is SEQ ID NO: 4, LCDR3 is SEQ ID NO: 5, HCDR1 is SEQ ID NO: 12, HCDR2 is SEQ ID NO: 15, and HCDR3 is SEQ ID NO: 14, wherein the human engineered CGRP antibody binds to human CGRP. The present invention provides a human engineered CGRP antibody comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO: 6, LCDR2 is SEQ ID NO: 7, LCDR3 is SEQ ID NO: 5, HCDR1 is SEQ ID NO: 12, HCDR2 is SEQ ID NO: 16, and HCDR3 is SEQ ID NO: 39, wherein the human engineered CGRP antibody binds to human CGRP. The present invention provides a human engineered CGRP antibody comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO: 8, LCDR2 is SEQ ID NO: 4, LCDR3 is SEQ ID NO: 5, HCDR1 is SEQ ID NO: 12, HCDR2 is SEQ ID NO: 15, and HCDR3 is SEQ ID NO: 39, wherein the human engineered CGRP antibody binds to human CGRP. The present invention provides a human engineered CGRP antibody comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO:9, LCDR2 is SEQ ID NO:7, LCDR3 is SEQ ID NO:5, HCDR1 is SEQ ID NO:12, HCDR2 is SEQ ID NO:15, and HCDR3 is SEQ ID NO:39, wherein the human engineered CGRP antibody binds to human CGRP.

The invention provides _a human engineered CGRP antibody comprising a LCVR and _a HCVR, wherein LCDR1 is _{RASX1X2IX3X4YLN} (SEQ ID NO:10), LCDR2 is _YTSX5YHS (SEQ ID NO:11), LCDR3 is _QQGDALPPT (SEQ ID NO:5), and HCDR1 is GYTFGNYWMQ (SEQ ID NO:12), HCDR2 is _{AIYEGTGX6TX7YIQKFAX8} (SEQ ID NO: ₃₇ ), and HCDR3 is _LSDYVSGFX9Y (SEQ ID NO:38), wherein _Xi is Q, _R , or K; _X2 is D or P, _X3 is D or S, _X4 is N or K, _X5 is G or E, _X6 is K or D, _X7 is V or R, _X8 is D or G, and _X9 is G or S, wherein the antibody binds human CGRP.

In another embodiment, the present invention provides a human engineered CGRP antibody comprising a light chain variable region (LCVR) and a heavy chain variable region (HCVR), wherein the LCVR comprises

DIQMTQSPSSSLSASVGDRVTITCRASX ₁ X ₂ IX ₃ X ₄ YLNWYQQKPGKAPKLLIYYTSX ₅

YHSGVPSRFSGSGSGTDFTX ₆ TISSLQPEDX ₇ ATYYCQQGDALPPTFGX ₈ GTKX ₉ EIK

wherein _X1 is Q, K, or R; _X2 is D or P; _X3 is D or S; _X4 is K or N; _X5 is E or G; _X6 is F or L; _X7 is I or F; _X8 is Q or G; and _X9 is L or V. (SEQ ID NO: 42)

And HCVR contains

QVQLVQSGAEVKKPGX _{1SVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAI}

YEGTGX ₂ TX ₃ YIWKFAX ₄ RVTX ₅ TX ₆ DX ₇ STSTX ₈ YMELSSLRSEDTAVYYCARLSDY

VSGFX ₉ YWGQGTX ₁₀ VTVSS, where _XX = A or S; X ₂ = K or D; X ₃ = V or R; X ₄ = G or D;

_X5 = M or I; _X6 = R or A; _X7 = T or K; _X8 = V or A; _X9 = G or S; and _X10 = L or T. (SEQ ID NO: 43),

Wherein the antibody binds to human CGRP.

In another embodiment, the present invention provides a human engineered CGRP antibody comprising a light chain variable region (LCVR) and a heavy chain variable region (HCVR), wherein the LCVR and HCVR are amino acid sequences selected from the group consisting of:

a. LCVR is SEQ ID NO: 17 and HCVR is SEQ ID NO: 22;

b. LCVR is SEQ ID NO: 18 and HCVR is SEQ ID NO: 23;

c. LCVR is SEQ ID NO: 19 and HCVR is SEQ ID NO: 24;

d. LCVR is SEQ ID NO: 20 and HCVR is SEQ ID NO: 25; and

e. LCVR is SEQ ID NO: 21 and HCVR is SEQ ID NO: 26.

The present invention provides human engineered CGRP antibodies comprising a LCVR of SEQ ID NO: 17 and a HCVR of SEQ ID NO: 22. In preferred embodiments, the present invention provides human engineered CGRP antibodies comprising a LCVR of SEQ ID NO: 18 and a HCVR of SEQ ID NO: 23. The present invention provides human engineered CGRP antibodies comprising a LCVR of SEQ ID NO: 19 and a HCVR of SEQ ID NO: 24. The present invention provides human engineered CGRP antibodies comprising a LCVR of SEQ ID NO: 20 and a HCVR of SEQ ID NO: 25. The present invention provides human engineered CGRP antibodies comprising a LCVR of SEQ ID NO: 21 and a HCVR of SEQ ID NO: 26.

The present invention also provides a human engineered CGRP antibody comprising a light chain (LC) and a heavy chain (HC), wherein the LC and HC amino acid sequences are selected from the group consisting of:

a) LC is SEQ ID NO: 27 and HC is SEQ ID NO: 32;

b) LC is SEQ ID NO: 28 and HC is SEQ ID NO: 33;

c) LC is SEQ ID NO: 29 and HC is SEQ ID NO: 34;

d) LC is SEQ ID NO: 30 and HC is SEQ ID NO: 35; and

e) LC is SEQ OD NO: 31 and HC is SEQ ID NO: 36.

In an embodiment, the human engineered CGRP antibody comprises an LC of SEQ ID NO: 27 and an HC of SEQ ID NO: 32. In another embodiment, the human engineered CGRP antibody comprises an LC of SEQ ID NO: 28 and an HC of SEQ ID NO: 33. In another embodiment, the human engineered CGRP antibody comprises an LC of SEQ ID NO: 29 and an HC of SEQ ID NO: 34. In another embodiment, the human engineered CGRP antibody comprises an LC of SEQ ID NO: 30 and an HC of SEQ ID NO: 35. In another embodiment, the human engineered CGRP antibody comprises an LC of SEQ ID NO: 31 and an HC of SEQ ID NO: 36. In an embodiment, the human engineered CGRP antibody comprises two light chains and two heavy chains, wherein each LC amino acid sequence is SEQ ID NO: 27 and each HC amino acid sequence is SEQ ID NO: 32. In an embodiment, the human engineered CGRP antibody comprises two light chains and two heavy chains, wherein each LC amino acid sequence is SEQ ID NO: 28 and each HC amino acid sequence is SEQ ID NO: 33. In an embodiment, the human engineered CGRP antibody comprises two light chains and two heavy chains, wherein each LC amino acid sequence is SEQ ID NO: 29 and each HC amino acid sequence is SEQ ID NO: 34. In an embodiment, the human engineered CGRP antibody comprises two light chains and two heavy chains, wherein each LC amino acid sequence is SEQ ID NO: 30 and each HC amino acid sequence is SEQ ID NO: 35. In an embodiment, the human engineered CGRP antibody comprises two light chains and two heavy chains, wherein each LC amino acid sequence is SEQ ID NO: 31 and each HC amino acid sequence is SEQ ID NO: 36. The present invention also provides antigen-binding fragments of the antibodies described herein.

The present invention also provides human engineered CGRP antibodies having an IC50 <1.0 nM in an assay essentially as described in Example 4. In embodiments, the present invention provides human engineered CGRP antibodies comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO:6, LCDR2 is SEQ ID NO:7, LCDR3 is SEQ ID NO:5, HCDR1 is SEQ ID NO:12, HCDR2 is SEQ ID NO:16 and HCDR3 is SEQ ID NO:39, and wherein the human engineered CGRP antibody binds human CGRP with an IC50 <1.0 nM in an assay essentially as described in Example 4. In embodiments, the present invention provides human engineered CGRP antibodies comprising the LCVR amino acid sequence of SEQ ID NO:19 and the HCVR amino acid sequence of SEQ ID NO:24, wherein the human engineered CGRP antibody has an IC50 <1.0 nM in an assay essentially as described in Example 4. In an embodiment, the present invention provides a human engineered CGRP antibody comprising the LC amino acid sequence of SEQ ID NO: 29 and the HC amino acid sequence of SEQ ID NO: 34, wherein the human engineered CGRP antibody has an IC50 < 1.0 nM in an assay essentially as described in Example 4.

The present invention also provides human engineered CGRP antibodies having an IC50 < 0.5 nM in an assay essentially as described in Example 4. In embodiments, the present invention provides human engineered CGRP antibodies comprising a LCVR and a HCVR, wherein LCDR1 is SEQ ID NO: 6, LCDR2 is SEQ ID NO: 7, LCDR3 is SEQ ID NO: 5, HCDR1 is SEQ ID NO: 12, HCDR2 is SEQ ID NO: 16 and HCDR3 is SEQ ID NO: 39, and wherein the human engineered CGRP antibody binds human CGRP with an IC50 < 0.5 nM in an assay essentially as described in Example 4. In embodiments, the present invention provides human engineered CGRP antibodies comprising the LCVR amino acid sequence of SEQ ID NO: 19 and the HCVR amino acid sequence of SEQ ID NO: 24, wherein the human engineered CGRP antibody has an IC50 < 0.5 nM in an assay essentially as described in Example 4. In an embodiment, the present invention provides a human engineered CGRP antibody comprising the LC amino acid sequence of SEQ ID NO: 29 and the HC amino acid sequence of SEQ ID NO: 34, wherein the human engineered CGRP antibody has an IC50 < 0.5 nM in an assay essentially as described in Example 4.

The present invention also provides a pharmaceutical composition comprising the human engineered CGRP antibody or antigen-binding fragment thereof of the present invention and a pharmaceutically acceptable carrier, diluent or excipient. The present invention also provides a pharmaceutical composition comprising the CGRP antibody of the present invention, together with a pharmaceutically acceptable carrier and optionally other therapeutic ingredients.

In another aspect, the present invention provides a method for treating osteoarthritis pain, comprising administering to a patient in need thereof a human engineered CGRP antibody or antigen-binding fragment thereof of the present invention. In another aspect, the present invention provides a method for treating migraine, comprising administering to a patient in need thereof a human engineered CGRP antibody or antigen-binding fragment thereof of the present invention.

The present invention also provides a human engineered CGRP antibody or antigen-binding fragment thereof of the present invention for use in therapy. In another aspect, the present invention provides a human engineered CGRP antibody or antigen-binding fragment thereof of the present invention for use in treating osteoarthritis pain. In another aspect, the present invention provides a human engineered CGRP antibody or antigen-binding fragment thereof of the present invention for use in treating migraine.

The present invention also provides the use of the human engineered CGRP antibody or its antigen-binding fragment of the present invention for treating osteoarthritis pain. In another aspect, the present invention also provides the use of the human engineered CGRP antibody or its antigen-binding fragment of the present invention for treating migraine. The present invention also provides the use of the human engineered CGRP antibody or its antigen-binding fragment in the production of a medicament for treating osteoarthritis. The present invention also provides the use of the human engineered CGRP antibody or its antigen-binding fragment in the production of a medicament for treating migraine.

Naturally occurring full-length antibodies are immunoglobulin molecules that are composed of two heavy (H) chains and two light (L) chains interconnected by disulfide bonds. The amino-terminal portion of each chain includes a variable region of approximately 100-110 amino acids that is primarily responsible for antigen recognition through the complementary determining regions (CDRs) contained therein. The carboxyl-terminal portion of each chain defines a constant region that is primarily responsible for effector function.

Interspersed among the CDRs are more conserved regions known as framework regions (FRs). Each light chain variable region (LCVR) and heavy chain variable region (HCVR) consists of three CDRs and four FRs, arranged from amino-terminus to carboxyl-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The three CDRs of the light chain are referred to as "LCDR1, LCDR2, and LCDR3," and the three CDRs of the heavy chain are referred to as "HCDR1, HCDR2, and HCDR3." The CDRs contain the majority of residues that form specific interactions with the antigen. The numbering and positioning of the CDR amino acid residues within the LCVR and HCVR regions follows the well-known Kabat numbering convention.

Light chains are classified as either kappa or lambda, and are characterized by specific constant regions known in the art. Heavy chains are classified as gamma, mu, alpha, delta, or epsilon, defining the antibody's isotype, IgG, IgM, IgA, IgD, or IgE, respectively. IgG antibodies can be further divided into subclasses, e.g., IgG1, IgG2, IgG3, and IgG4. Each heavy chain type is characterized by a specific constant region, the sequence of which is known in the art.

As used herein, the term "monoclonal antibody" (Mab) refers to an antibody derived from a single copy or clone, including, for example, any eukaryotic, prokaryotic or phage clone, without limitation to the method of producing it. The Mab of the present invention is preferably present in a homogeneous or substantially homogeneous population. A complete Mab contains two heavy chains and two light chains. "Antigen-binding fragments" of such monoclonal antibodies include, for example, Fab fragments, Fab' fragments, F(ab') ₂ fragments and single-chain Fv fragments. The monoclonal antibodies of the present invention and their antigen-binding fragments can be produced by, for example, recombinant technology, phage display technology, synthetic technology (e.g., CDR-grafting), or a combination of such technologies, or other technologies known in the art. For example, mice can be immunized with human CGRP or a fragment thereof, and the antibodies obtained can be recovered and purified and evaluated by the methods disclosed in the following examples to determine whether they have similar or identical binding and functional properties to the antibody compounds disclosed herein. Antigen-binding fragments can also be prepared by conventional methods. Methods for producing and purifying antibodies and antigen-binding fragments are generally known in the art and can be found, for example, in Harlow and Lane (1988) Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, Chapters 5-8 and 15, ISBN 0-87969-314-2.

The phrase "human engineered CGRP antibody" refers to a monoclonal antibody that has been created and/or manipulated to have the binding and functional properties according to the present invention, binds to human CGRP, and has substantially human or fully human framework regions surrounding the CDRs derived from a non-human antibody. "Antigen-binding fragments" of such human engineered antibodies include, for example, Fab fragments, Fab' fragments, F(ab') ₂ fragments, and single-chain Fv fragments. "Framework region" or "framework sequence" refers to any one of framework regions 1 to 4. The human engineered antibodies and antigen-binding fragments thereof encompassed by the present invention include molecules in which any one or more of framework regions 1 to 4 are substantially or fully human, i.e., in which there is any possible combination of individual substantially or fully human framework regions 1 to 4. For example, this includes molecules in which framework region 1 and framework region 2, framework region 1 and framework region 3, framework regions 1, 2, and 3, etc. are substantially or fully human. Substantially human frameworks are those that have at least about 80% sequence identity to known human germline framework sequences. Preferably, the substantially human framework has at least about 85%, about 90%, about 95%, or about 99% sequence identity to a known human germline framework sequence.

Fully human frameworks are those that are identical to known human germline framework sequences. Human framework germline sequences are available from ImMunoGeneTics (IMGT) at the website http://imgt.cines.fr or from The Immunoglobulin Facts Book by Marie-Paule Lefranc and Gerard Lefranc, Academic Press, 2001, ISBN 012441351. For example, the germline light chain framework can be selected from the group consisting of: A11, A17, A18, A19, A20, A27, A30, L1, L11, L12, L2, L5, L15, L6, L8, O12, O2, and O8, and the germline heavy chain framework can be selected from the group consisting of: VH2-5, VH2-26, VH2-70, VH3-20, VH3-72, VHI-46, VH3-9, VH3-66, VH3-74, VH4-31, VHI-18, VHI-69, VI-13-7, VH3-11, VH3-15, VH3-21, VH3-23, VH3-30, VH3-48, VH4-39, VH4-59, and VH5-51.

Several different methods can be used to generate engineered antibodies other than those disclosed herein, showing similar functional properties according to the present invention. Specific antibody compounds disclosed herein can be used as templates or parental antibody compounds to prepare other antibody compounds. In one approach, the parental antibody compound CDRs are transplanted onto a human framework with a high degree of sequence identity to the parental antibody compound framework. Generally, the sequence identity of the new framework is at least about 80%, at least about 85%, at least about 90%, at least about 95% or at least about 99% identical to the sequence of the corresponding framework of the parental antibody compound. The transplant can result in a reduction in binding affinity compared to the parental antibody. If this is the case, the framework can be backmutated to the parental framework at certain positions based on the specific criteria disclosed by Queen et al. (1991) Proc. Natl. Acad. Sci. USA 88: 2869. Other references describing use in humanizing mouse antibodies include U.S. Pat. Nos. 4,816,397; 5,225,539 and 5,693,761; the computer programs ABMOD and ENCAD as described in Levitt (1983) J. Mol. Biol. 168:595-620; and the method of Winter and co-workers (Jones et al. (1986) Nature 321:522-525; Riechmann et al. (1988) Nature 332:323-327; and Verhoeyen et al. (1988) Science 239:1534-1536).

Identification of residues to consider for backmutation can be performed as follows:

Framework amino acids of the human germline sequence used ("acceptor framework") were substituted with framework amino acids from the framework of the parent antibody compound ("donor framework") when the amino acids fell into the following categories:

(a) the amino acids in the human framework regions of the acceptor framework are unusual for human frameworks at said positions, while the corresponding amino acids in the donor immunoglobulin are typical for human frameworks at said positions;

(b) the amino acid is positioned immediately adjacent to one of the CDRs; or

(c) In three-dimensional immunoglobulin models, any side chain atom of a framework amino acid is within about 5-6 angstroms (center to center) of any atom of a CDR amino acid.

When each amino acid in the human framework region of the acceptor framework and the corresponding amino acid in the donor framework are generally uncommon for human frameworks at that position, such amino acid can be replaced with an amino acid that is typical for human frameworks at that position. This back mutation rule enables restoration of the activity of the parent antibody compound.

Another approach to generating human engineered antibodies that exhibit functional properties similar to those of the antibody compounds disclosed herein involves randomly mutating amino acids in the grafted CDRs without altering the framework, and screening the resulting molecules for binding affinity and other functional properties that are as good as or better than those of the parent antibody compound. It is also possible to introduce a single mutation at each amino acid position in each CDR and then evaluate the effect of such mutations on binding affinity and other functional properties. Individual mutations that produce improved properties can be combined to evaluate their effects when combined with each other.

In addition, a combination of the two approaches described above is also possible. After CDR grafting, in addition to introducing amino acid changes in the CDRs, specific framework regions can be backmutated. This methodology is described in Wu et al. (1999) J. Mol. Biol. 294: 151-162.

Applying the teachings of the present invention, those skilled in the art can use conventional techniques, such as site-directed mutagenesis, to replace amino acids in the CDR and framework sequences disclosed herein, thereby generating other variable region amino acid sequences derived from the sequences of the present invention. All alternative naturally occurring amino acids can be introduced at specific substitution sites. These other variable region amino acid sequences can then be screened using the methods disclosed herein to identify sequences with specified in vivo functions. Through this method, other sequences suitable for preparing human engineered antibodies and antigen-binding portions thereof according to the present invention can be identified. Preferably, amino acid substitutions within the framework are limited to 1, 2, or 3 positions in any one or more of the 4 light chains and/or heavy chain framework regions disclosed herein. Preferably, amino acid substitutions in the CDRs are limited to 1, 2, or 3 positions in any one or more of the 3 light chains and/or heavy chain CDRs. Combinations of various changes in these framework regions and CDRs are also possible.

The term "treating" refers to a process that involves slowing, interrupting, suppressing, controlling, halting, reducing or reversing the progression or severity of a symptom, disorder, condition or disease, but does not necessarily involve completely eliminating all disease-related symptoms, conditions or disorders associated with CGRP activity.

According to the Headache Classification Committee of the International Headache Society (International Headache Society, 2004), the term "migraine" as used herein refers to "migraine without aura" (formerly "common migraine") and "migraine with aura" (formerly "classic migraine"). For example, "migraine without aura" is generally characterized by being pulsatile, moderate or severe in intensity, aggravated by routine physical activity, unilateral in distribution, and associated with nausea, photophobia, and phonophobia. "Migraine with aura" may include interference with vision, interference with other senses, unilateral weakness, and in some cases difficulty speaking.

The human engineered antibodies of the present invention can be used as drugs in human medicine and are administered by various routes. Most preferably, such compositions are used for parenteral administration. Such pharmaceutical compositions can be prepared by methods generally known in the art (see, for example, Remington: The Science and Practice of Pharmacy, 19th edition (1995), A. Gennaro et al., Mack Publishing Co.), and the compositions comprise human engineered antibodies as disclosed herein and a pharmaceutically acceptable carrier, diluent or excipient.

The results of the following assays demonstrate that the exemplified monoclonal antibodies and antigen-binding fragments thereof of the present invention are useful for treating osteoarthritis pain.

Example 1 - Production of Antibodies

Antibodies I-V can be prepared and purified as follows. Appropriate host cells (e.g., HEK293EBNA or CHO) are transiently or stably transfected using an expression system for secreting the antibody, using an optimal, predetermined HC:LC vector ratio or a single vector system encoding the LC (e.g., SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, or SEQ ID NO: 31) and HC (e.g., SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, or SEQ ID NO: 36). The culture medium from which the antibody is secreted is purified using any of a variety of commonly used techniques. For example, the culture medium can be conveniently applied to a protein A or G sepharose FF column equilibrated with a compatible buffer (e.g., phosphate-buffered saline (pH 7.4)). The column is washed to remove non-specifically bound components. For example, the bound antibody is eluted by a pH gradient (e.g., 0.1 M sodium phosphate buffer, pH 6.8 to 0.1 M sodium citrate buffer, pH 3.0). Antibody fractions can be detected by SDS-PAGE, for example, and then mixed. Depending on the intended use, additional purification is optional. Conventional techniques can be used to concentrate and/or sterile filter antibodies. Soluble polymers and polymers can be effectively removed by conventional techniques (including size exclusion, hydrophobic interaction, ion exchange or hydroxyapatite chromatography). The antibody purity after these chromatography steps is greater than 99%. The product can be frozen immediately at -70°C or can be lyophilized. The amino acid sequences of these exemplary antibodies are provided below.

Table 1 - Antibody SEQ ID NO

Example 2: Affinity (Kd) Measurement of Human Engineered CGRP Antibodies

On a Biacore T100 instrument pre-loaded with HBS-EP+ (GE Healthcare, 10mM Hepes pH7.4+150mM NaCl+3mM EDTA+0.05% surfactant P20) running buffer, surface plasmon resonance was used to determine the binding affinity of the exemplified antibodies to CGRP, and the analysis temperature was set to 37°C. A CM5 chip containing immobilized protein A (generated using standard NHS-EDC amino coupling) was used on all four flow cells (Fc) to adopt capture methodology. Antibody samples were prepared to 2 μg/mL by dilution with running buffer. Human CGRP samples were prepared at final concentrations of 5.0, 2.5, 1.3, 0.63, 0.31, and 0 (blank) nM by dilution with running buffer. Fresh, single-use aliquots of CGRP were used for each replicate to avoid multiple freeze-thaw cycles. Each analysis cycle consisted of (1) capturing the antibody sample in separate flow cells (Fc2, Fc3, and Fc4), (2) injecting 350 μL (210-sec) CGRP at 100 μL/min over all Fcs, (3) restoring the buffer flow for 10 min to monitor the dissociation phase, (4) regenerating the chip surface by injecting 5 μL (15-sec) glycine, pH 1.5, and (5) equilibrating the chip surface by injecting 5 μL (15-sec) HBS-EP+. The injection was repeated twice for each CGRP concentration. The data were processed using standard double controls and fitted to a 1:1 binding model using Biacore T100 Evaluation Software version 2.0.1 to determine the association rate (k _on , M ^-1 s ^-1 units), dissociation rate (k _off , s ^-1 units), and Rmax (RU units). The equilibrium dissociation constant (K _D ) was calculated according to the relationship K _D = k _off / _kon and is expressed in molar units. The values provided in Table 2 are the average of n experiments. Table 2 demonstrates that the antibodies exemplified by the present invention bind CGRP with an affinity of <200 pM.

Table 2 - Antibody Binding Affinity

Example 3 - Reducing pain in the MIA model

Injection of monoiodoacetic acid (MIA) into the knee joint of rats produces an acute inflammatory lesion, followed by chronic degradation of joint tissue in the injected joint. Pain caused by joint injury can be measured by differential weight bearing on the hind limbs using an incapacitance tester. The MIA model is well described in the literature and has been used to demonstrate the efficacy of various mechanisms and compounds versus pain. Efficacy is conventionally measured by the ability of a compound to partially normalize weight distribution.

To determine the ability of the antibodies exemplified in the present invention to reduce pain, male Lewis rats (Harlan, Indianapolis, IN) approximately 8 weeks old at the time of MIA injection were used. Rats were housed 2 or 3 per cage and maintained at a constant temperature and a 12-hour light/12-hour dark cycle. Except during data collection, animals had free access to food and water at all times. All experiments were performed according to protocols approved by the Eli Lilly Institutional Animal Care and Use Committee.

On day 0, each rat was injected with 0.3 mg of MIA in 50 μL of saline into the right knee and 50 μL of saline into the left knee. On the designated day after MIA, human IgG4 control or test CGRP antibodies in PBS were administered subcutaneously (n=6/group). Three days after administration of the CGRP antibody, pain was measured using an incapacitance test that measures the difference in hind paw weight bearing between the MIA-treated knee and the saline-injected knee. Each measurement was the average of three different measurements over 1 second each.

The data are expressed as the percentage inhibition of pain, which is calculated by dividing the mean of the CGRP antibody-treated group by the mean of the control antibody group, subtracting the value from 1, and multiplying the resulting value by 100. The CGRP-treated group and the vehicle group were also compared by one-way analysis of the means and Dunnett's test using JMP (versions 5.1 and 6) statistical analysis program (SAS Institute Inc., Cary, NC). Differences were considered significant if the P value was less than 0.05. As shown in Table 3, the data confirm that the CGRP antibodies exemplified by the present invention significantly reduce pain.

Table 3 - Percent Inhibition of MIA-Induced Pain

Example 4 - Inhibition of CGRP-induced cAMP formation in SK-N-MC cells

To compare the ability of the antibodies of the invention to antibody G1 (LCVR-SEQ ID NO:40 and HCVR-SEQ ID NO:41), inhibition of CGRP-induced cAMP formation in SK-N-MC was determined. Binding of CGRP to its receptor results in stimulation of cAMP production. The CGRP receptor is a heterotrimeric complex consisting of a calcitonin receptor-like receptor (CLR, a G protein-coupled receptor) and receptor activity regulating protein (RAMP)-1, coupled to the cytoplasm of the receptor component protein (RCP). The human neuroepithelioma cell line SK-N-MC naturally expresses these three molecules and can therefore be used to assess the effects of CGRP on signal transduction events. cAMP production is a standard measure of G protein-coupled receptor activation.

SK-N-MC cells were cultured in MEM containing 10% FBS, 1X MEM non-essential amino acids, 1X 100mM MEM sodium pyruvate, 1X Pen/Strep, and 2mM L-glutamine. After harvesting, cells were washed once and resuspended in assay buffer (stimulation buffer (HBSS with Mg and Ca, 5mM HEPES, 0.1% BSA, 100uM ascorbic acid) diluted 1:2 with Dulbecco's PBS containing a final concentration of 0.5mM IBMX) and seeded in 96-well plates at 15,000 cells per well. Test antibodies or control human IgG4 antibodies (serial 4-fold dilutions in assay buffer, 10-fold concentration) were added to the cells, followed by a fixed amount of human α-CGRP (2nM; Bachem H1470). The plates were incubated at room temperature for 1 hour. cAMP levels were measured by a homogeneous time-resolved fluorescence (HTRF) assay system (Cisbio).

The amount of cAMP induced by 2 nM human CGRP in the presence of different concentrations of antibody was calculated as a percentage inhibition compared to CGRP alone. The antibody concentration (IC50) that produces 50% inhibition of cAMP production was then calculated from a 4-parameter curve fitting model. According to the methods described herein, Antibody III ( _IC50 of 0.41 nM) inhibited the amount of cAMP induced by CGRP to a much greater extent than Antibody G1 ( _IC50 of 5.36 nM), possibly due to the faster _kon rate of _Antibody III (as measured by Biacore).

Example 5 - Rat Dura Plasma Protein Extravasation (PPE) Model

The rat PPE model is a well-established preclinical model that can be used to evaluate the efficacy of anti-CGRP antibodies in treating migraine.

A 2 mg/mL solution of anti-CGRP antibody (Ab) was prepared in saline solution. All subsequent dilutions were performed with saline. A 2 mg/mL solution of monoclonal isotype control antibody (IgG) was also prepared in saline.

Male Sprague-Dawley rats (250 to 350 g) from Harlan Laboratories were anesthetized with Nembutal (60 mg/kg, ip.) and placed in a stereotaxic frame (David Kopf Instruments) with the incisor bar set to –2.5 mm. After a midline sagittal scalp incision, two pairs of bilateral holes were drilled through the skull (3.2 mm posterior, 1.8 and 3.8 mm lateral, all coordinates referenced to bregma). Paired stainless steel stimulating electrodes (Rhodes Medical Systems Inc.), insulated except for the tip, were inserted through the holes in both hemispheres to a depth of 9.2 mm below the dura mater . Test antibody or saline vehicle (1 mL/kg) was administered intravenously via the femoral vein. After 8 minutes, a solution of bovine serum albumin (BSA) (FITC-BSA, Sigma A9771) labeled with fluorescein isothiocyanate (FITC) dye (20 mg/kg, i.v.) was injected into the femoral vein as a marker for protein extravasation. 10 minutes after administration of the test antibody or vehicle, the left trigeminal ganglion was stimulated for 5 minutes using a Model S48 Grass Instrument stimulator at a current intensity of 1.0 mA (5 Hz, 5 ms duration). After 5 minutes of stimulation, the rats were killed by bleeding with 40 mL of saline. Bleeding also washed the residual FITC/BSA out of the blood vessels. The top of the skull was removed to collect the dura mater . Membrane samples were removed from both hemispheres, rinsed with water, and spread on microscope slides. The slides were dried on a slide dryer for 15 minutes and coverslipped with a 70% glycerol/water solution.

Equipped with a fluorescence microscope (Zeiss) of a grating monochromator and a spectrophotometer for the amount of the FITC-BSA dye in each dura mater sample. The microscope is equipped with an electric platform interfaced with a personal computer. This helps the computer-controlled platform to move, and the fluorescence measurement of 25 points (500 μm steps) is carried out to each dura mater sample. The extravasation induced by electrical stimulation of the trigeminal ganglion is the effect of the same side (that is, only occurring on one side of the dura mater where the trigeminal ganglion is stimulated). This allows the use of the other half (unstimulated) of the dura mater as a control. Calculate the extravasation ratio (that is, the amount of the extravasation from the dura mater on the stimulated side is compared to the ratio of the unstimulated side).

sequence

ACDTATCVTHRLAGLLSRSGGVVKNNFVPTNVGSKAF (SEQ ID NO: 1)

ACNTATCVTHRLAGLLSRSGGMVKSNFVPTNVGSKAF (SEQ ID NO: 2)

_X1 is Q, R or K;

_X2 is D or P;

X ₃ is D or S;

X ₄ is N or K; and

X ₅ is G or E

X ₆ is K or D;

X ₇ is V or R;

X ₈ is D or G; and

X ₉ is G or S

DIQMTQSPSSSLSASVGDRVTITCRASQDIDNYLNWYQQKPGKAPKLLIYYTSEYHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGDALPPTFGQGTKLEIK(SEQ ID NO:17)

DIQMTQSPSSSLSASVGDRVTITCRASQDIDNYLNWYQQKPGKAPKLLIYYTSEYHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGDALPPTFGQGTKLEIK(SEQ ID NO:18)

DIQMTQSPSSSLSASVGDRVTITCRASKDISKYLNWYQQKPGKAPKLLIYYTSGYHSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGDALPPTTFGGGTKVEIK(SEQ ID NO:19)

DIQMTQSPSSSLSASVGDRVTITCRASRPIDKYLNWYQQKPGKAPKLLIYYTSEYHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGDALPPTFGQGTKLEIK(SEQ ID NO:20)

DIQMTQSPSSSLSASVGDRVTITCRASQDIDKYLNWYQQKPGKAPKLLIYYTSGYHSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGDALPPTFGGGTKVEIK(SEQ ID NO:21)

QVQLVQSGAEVKKPGASVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGDTRYIQKFAGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARLSDYVSGFSYWGQGTLVTVSS(SEQ ID NO:22)

QVQLVQSGAEVKKPGASVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFAGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARLSDYVSGFSYWGQGTLVTVSS(SEQ ID NO:23)

QVQLVQSGAEVKKPGSSVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFADRVTITADKSTSTAYMELSSLRSEDTAVYYCARLSDYVSGFGYWGQGTTVTVSS(SEQ ID NO:24)

QVQLVQSGAEVKKPGASVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFAGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARLSDYVSGFGYWGQGTLVTVSS(SEQ ID NO:25)

QVQLVQSGAEVKKPGSSVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFAGRVTITADKSTSTAYMELSSLRSEDTAVYYCARLSDYVSGFGYWGQGTTVTVSS(SEQ ID NO:26)

DIQMTQSPSSSLSASVGDRVTITCRASQDIDNYLNWYQQKPGKAPKLLIYYTSEYHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGDALPPTFGQGTKLEIKRT VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ IDNO:27)

DIQMTQSPSSSLSASVGDRVTITCRASQDIDNYLNWYQQKPGKAPKLLIYYTSEYHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGDALPPTFGQGTKLEIKRT VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ IDNO:28)

DIQMTQSPSSSLSASVGDRVTITCRASKDISKYLNWYQQKPGKAPKLLIYYTSGYHSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGDALPPTTFGGGTKVEIKRT VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ IDNO:29)

DIQMTQSPSSSLSASVGDRVTITCRASRPIDKYLNWYQQKPGKAPKLLIYYTSEYHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGDALPPTFGQGTKLEIKRT VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ IDNO:30)

DIQMTQSPSSSLSASVGDRVTITCRASQDIDKYLNWYQQKPGKAPKLLIYYTSGYHSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGDALPPTFGGGTKVEIKRT VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ IDNO:31)

QVQLVQSGAEVKKPGASVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGDTRYIQKFAGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARLSDYVSGFSYWGQG TLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKA KGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG(SEQID NO:32)

QVQLVQSGAEVKKPGASVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFAGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARLSDYVSGFSYWGQG TLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKA KGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG(SEQID NO:33)

QVQLVQSGAEVKKPGSSVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFADRVTITADKSTSTAYMELSSLRSEDTAVYYCARLSDYVSGFGYWGQG TTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKA KGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG(SEQID NO:34)

QVQLVQSGAEVKKPGASVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFAGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARLSDYVSGFGYWGQG TLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKA KGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG(SEQID NO:35)

QVQLVQSGAEVKKPGSSVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGKTVYIQKFAGRVTITADKSTSTAYMELSSLRSEDTAVYYCARLSDYVSGFGYWGQG TTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKA KGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG(SEQID NO:36)

EIVLTQSPATLSLSPGERATLSCKASKRVTTYVSWYQQKPGQAPRLLIYGASNRYLGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCSQSYNYPYTFGQGTKLEIK(SEQ ID NO:40)

EVQLVESGGGLVQPGGSLRLSCAASGFTFSNYWISWVRQAPGKGLEWVAEIRSESDASATHYAEAVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCLAYFDYGLAIQNYWGQGTLVTVSS(SEQ ID NO:41)

_{DIQMTQSPSSSLSASVGDRVTITCRASX 1 ​ ​ ​ ​ ​ ​ ​}

X ₁ =Q, K, or R;

X ₂ =D or P;

X ₃ D or S;

X ₄ =K or N;

X ₅ =E or G;

X ₆ =F or L;

X ₇ =I or F;

X ₈ =Q or G; and

X ₉ =L or V. (SEQ ID NO: 42)

QVQLVQSGAEVKKPGX ₁ SVKVSCKASGYTFGNYWMQWVRQAPGQGLEWMGAIYEGTGX ₂ TX ₃ YIQKFAX ₄ RVTX ₅ TX ₆ DX ₇ STSTX ₈ YMELSSLRSEDTAVYYCARLSDYVSGFX ₉ YWGQGTX ₁₀ VTVSS

X ₁ =A or S;

X ₂ =K or D;

X ₃ =V or R;

X ₄ =G or D;

X ₅ =M or I;

X ₆ =R or A;

X ₇ =T or K;

X ₈ =V or A;

X ₉ =G or S; and

X ₁₀ =L or T. (SEQ ID NO: 43)

Claims (13)

1. A human engineered CGRP antibody, the antibody comprising a light chain variable region (LCVR) and a heavy chain variable region (HCVR), wherein the LCVR comprises the amino acid sequences LCDR1, LCDR2, and LCDR3 and the HCVR comprises the amino acid sequences HCDR1, HCDR2, and HCDR3, wherein LCDR1 is SEQ ID NO:3, LCDR2 is SEQ ID NO:4, LCDR3 is SEQ ID NO:5, HCDR1 is SEQ ID NO:12, HCDR2 is SEQ ID NO:15, and HCDR3 is SEQ ID NO:14; or LCDR1 is SEQ ID NO:8, LCDR2 is SEQ ID NO:4, LCDR3 is SEQ ID NO:5, HCDR1 is SEQ ID NO:12, HCDR2 is SEQ ID NO:15, and HCDR3 is SEQ ID NO:39; or LCDR1 is SEQ ID NO:9, LCDR2 is SEQ ID NO:7, LCDR3 is SEQ ID NO:5, HCDR1 is SEQ ID NO:12, and HCDR2 is SEQ ID NO:39. NO:15, and HCDR3 is SEQ ID NO:39, wherein the human CGRP antibody binds to human CGRP. 2. The engineered human CGRP antibody of claim 1, wherein the LCVR amino acid sequence is SEQ ID NO:18 and the HCVR amino acid sequence is SEQ ID NO:23; or the LCVR amino acid sequence is SEQ ID NO:20 and the HCVR amino acid sequence is SEQ ID NO:25; or the LCVR amino acid sequence is SEQ ID NO:21 and the HCVR amino acid sequence is SEQ ID NO:26. 3. A pharmaceutical composition comprising a human engineered antibody and a pharmaceutically acceptable carrier as claimed in any one of claims 1 and 2. 4. A pharmaceutical composition comprising a human engineered antibody of any one of claims 1 and 2 and a pharmaceutically acceptable diluent. 5. A pharmaceutical composition comprising a human engineered antibody and a pharmaceutically acceptable excipient as claimed in any one of claims 1 and 2. 6. The pharmaceutical composition according to any one of claims 3-5, further comprising one or more therapeutic ingredients. 7. A human engineered CGRP antibody as claimed in any one of claims 1-2, for use in therapy. 8. The engineered CGRP antibody of a human as claimed in any one of claims 1-2, used to prepare a pharmaceutical treatment for treating migraines. 9. The engineered human CGRP antibody of any one of claims 1-2, used to prepare a pharmaceutical treatment for treating osteoarthritis pain. 10. Use of the engineered human CGRP antibody of any one of claims 1-2 in the production of a medicament for treating migraines. 11. Use of the engineered human CGRP antibody of any one of claims 1-2 in the production of a medicament for treating osteoarthritis pain. 12. Use of the pharmaceutical composition of any one of claims 3-6 in the production of a medicament for treating migraines. 13. Use of the pharmaceutical composition of any one of claims 3-6 in the production of a medicament for treating osteoarthritis pain.