Il27rα binding molecules and methods of use

Abstract

To provide biologically active molecules comprising a single domain antibody (sdAb) that specifically binds to the extracellular domain of human IL27Rα, compositions comprising such antibodies, and methods of use thereof.SOLUTION: An IL27Rα-binding molecule is provided, which comprises a single domain antibody (sdAb) that specifically binds to the extracellular domain of IL2Rb and has a specific amino acid sequence.SELECTED DRAWING: None

Description

Technical Field

[0001] Cross - reference to Related Applications This application claims priority to U.S. Provisional Application No. 63 / 061,562, filed August 5, 2020; U.S. Provisional Application No. 63 / 078,745, filed September 15, 2020; and U.S. Provisional Application No. 63 / 135,884, filed January 11, 2021. The disclosures of these provisional applications are hereby incorporated by reference in their entirety for all purposes.

[0002] Field of the Invention The present disclosure relates to biologically active molecules comprising single - domain antibodies that specifically bind to the extracellular domain of the IL27 receptor α (IL27Rα), compositions comprising such single - domain antibodies, and methods of using the same.

Background Art

[0003] Background The cytokine IL - 27 is a heterodimeric cytokine consisting of p28 and EBI3, two non - covalently associated subunits. The p28 subunit belongs to the four - helix bundle cytokine family, while EBI3 is the shortest form of a soluble cytokine receptor having two representative cytokine - binding domains (Pflanz S, et al., Immunity. 2002 Jun;16(6):779 - 90).

[0004] Interleukin-27 receptor (IL27R) is a type I cytokine receptor for interleukin-27 (IL27). IL27R is a heterodimer composed of the IL27Rα subunit and glycoprotein 130 (IL6Rβ). IL27 is expressed by antigen-presenting cells and induces the differentiation of diverse T cell populations in the immune system. When IL27 binds to IL27R, intracellular signaling is initiated via several Jak family kinases that induce phosphorylation of STAT1 and STAT3. In activated T cells, IL27 mainly signals through STAT3

[23] , whereas in memory B cells, it mainly signals through STAT3. IL27 has been shown to have both pro-inflammatory and anti-inflammatory properties, and this pro-inflammatory or anti-inflammatory response is influenced by the context of IL27R-expressing cells.

[0005] The IL27Rα subunit (also known as the TCCR- or WSX-1 receptor) is an IL27 receptor subunit protected by intellectual property rights. Mature (signal peptide-free) IL27Rα is a 604 amino acid polypeptide with a 484 amino acid extracellular domain. The extracellular domain of IL27Rα has five domains: D1 - D5. D1 and D2 are primary cytokine binding domains, whereas the fibronectin type III (Fn3) domains D3, D4, and D5 are involved in ligand recognition to a much lesser extent than D1 and D2. Based on structural analysis, the Fn3 domains do not contribute to binding in the complex when the IL27 ligand is bound. Domains D1 and D2 are highly conserved, whereas the sequences of the Fn3 domains are more variable.

[0006] IL27 exhibits high (nanomolar) affinity for the IL27Ra subunit. The [IL27 / IL27Rα] complex associates with IL27Ra to complete the IL27 receptor signaling complex. The binding of gp130 to the [IL27 / IL27Rα] complex is considerably weaker than the interaction between IL-27 and IL-27R, which is somewhat common for cytokine receptor subunits (Pflanz S, et al., J Immunol. 2004 Feb 15;172(4):2225-31). The D5 domain of IL-27R and the D6 domain of gp130 come together in close proximity in the membrane for each receptor to adopt a "C" shape. This is necessary for the receptor complex to induce JAK binding in the intracellular domains of both receptors.

[0007] Monoclonal antibodies are the most widely used reagents for the detection and quantification of proteins. However, monoclonal antibodies are relatively large molecules of approximately 150 kDa, and due to their size, their use can potentially be limited in assays using several types of reagents that compete in the recognition of neighboring epitopes. A unique immunoglobulin class that contains heavy-chain domains and lacks light-chain domains (commonly referred to as "heavy-chain" antibodies (HCAb)) exists in camelid animals, including dromedary camels, llamas, alpacas, vicuñas, and guanacos, as well as in cartilaginous fish such as sharks. The isolated variable domain region of HCAb is known as VHH (an abbreviation of "variable-heavy-heavy" reflecting its structure) or Nanobody® (Ablynx). Single-domain VHH antibodies are approximately one-tenth the molecular weight of conventional mammalian IgG-class antibodies and have the advantage of being small in size (approximately 12 - 14 kD), which facilitates their binding to target epitopes that may not be accessible to conventional monoclonal IgG formats (Ingram et al., 2018). Furthermore, VHH single-domain antibodies are often characterized by high thermal stability, which facilitates the delivery of drugs to regions where it is difficult or impossible to maintain a cold chain. When these properties are combined with simple phage display recovery methods (unlike in the case of IgG antibodies) that do not require heavy-chain / light-chain pairing and simple manufacturing (e.g., manufacturing in bacterial expression systems), VHH single-domain antibodies are useful in various applications, including the development of imaging agents and therapeutic agents. Summary of the Invention

[0008] The present disclosure provides a polypeptide that specifically binds to IL27Ra.

[0009] The present disclosure provides a polypeptide that specifically binds to the extracellular domain of IL27Ra.

[0010] The present disclosure provides an IL27Ra binding molecule that specifically binds to the extracellular domain of human IL27Ra (hIL27Ra).

[0011] In some embodiments, the IL27Ra binding molecule comprises a single domain antibody (sdAb) that specifically binds to the extracellular domain of human IL27Ra.

[0012] In some embodiments, the IL27Ra binding molecule is an sdAb, and the sdAb comprises a set of CDRs corresponding to CDR1, CDR2, and CDR3 as shown in the rows of Table 1 below.

[0013] In some embodiments, the IL27Ra binding molecule comprises CDR1, CDR2, and CDR3 as described in the rows of Table 1 below, and each of CDR1, CDR2, and CDR3 may independently have at least 90% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity compared to the sequences described in the rows of Table 1 below, and may have 0, 1, 2, or 3 amino acid changes, optionally conservative amino acid changes.

[0014] In some embodiments, the IL27Ra binding molecule has at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 98%, or at least 99% (or is identical except for 1, 2, 3, or 4 amino acid substitutions which are optionally conservative amino acid substitutions) identity to any one of the polypeptide sequences of SEQ ID NOs: 2-25 as shown in Table 1 below, or consists of, optionally consists essentially of, or optionally comprises a single domain antibody (sdAb) having 100% identity thereto.

[0015] (Table 1) TIFF2025094022000001.tif60160TIFF2025094022000002.tif235160TIFF2025094022000003.tif235160TIFF2025094022000004.tif204160

[0016] In some embodiments, the foregoing set of CDRs is incorporated into a humanized VHH framework to become a "humanized" sdAb IL27Ra binding molecule.

[0017] Furthermore, the present disclosure provides methods of chemical or recombinant processes for preparing the IL27Ra binding molecules of the present disclosure.

[0018] Furthermore, the present disclosure provides nucleic acids encoding the IL27Ra binding molecules. Table 2 below shows examples of DNA sequences encoding the IL27Ra binding molecules as described herein.

[0019] (Table 2) DNA sequences encoding the VHHs of Table 1 TIFF2025094022000005.tif202158TIFF2025094022000006.tif202158TIFF2025094022000007.tif202158TIFF2025094022000008.tif202158TIFF2025094022000009.tif163158

[0020] In some embodiments, IL27Ra is murine IL27Ra.

[0021] In some embodiments, the IL27Ra binding molecule comprises a single domain antibody (sdAb) that specifically binds to the extracellular domain of murine or rat IL27Ra (mIL27Ra).

[0022] In some embodiments, the IL27Ra binding molecule is an sdAb, and the sdAb comprises a set of CDRs corresponding to CDR1, CDR2, and CDR3 as shown in the rows of Table 3 below.

[0023] In some embodiments, the IL27Ra binding molecule comprises CDR1, CDR2, and CDR3 as set forth in the rows of Table 3 below, and CDR1, CDR2, and CDR3 may each independently have at least 90% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity compared to the sequences set forth in the rows of Table 3 below, and may have 0, 1, 2, or 3 amino acid changes, optionally conservative amino acid changes.

[0024] In some embodiments, the IL27Ra binding molecule has at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 98%, or at least 99% identity to any one of the polypeptide sequences of SEQ ID NO: 122, 126, 130, 134, 138, 142, 146, 150, 154, 158, 162, 166, 170, and 174 as shown in Table 3 below (or is identical except for 1, 2, 3, or 4 amino acids which are optionally conservative substitutions), or consists of, optionally consists essentially of, or optionally comprises a single domain antibody (sdAb) having 100% identity.

[0025] (Table 3) mIL2Rg VHH and CDR Amino Acid (AA) Sequences TIFF2025094022000010.tif135167TIFF2025094022000011.tif216167TIFF2025094022000012.tif216167TIFF2025094022000013.tif54167

[0026] In some embodiments, the aforementioned set of CDRs is incorporated into a humanized VHH framework to be a "humanized" sdAb IL27Rα binding molecule.

[0027] Furthermore, the present disclosure provides methods of chemical or recombinant processes for preparing the IL27Rα binding molecules of the present disclosure.

[0028] Furthermore, the present disclosure provides a nucleic acid encoding an IL27Ra binding molecule. Table 4 below shows examples of DNA sequences encoding hIL27Ra binding molecules as described in Table 3 above.

[0029] (Table 4) DNA sequences encoding the VHHs of Table 3 TIFF2025094022000014.tif91164TIFF2025094022000015.tif212164TIFF2025094022000016.tif212164TIFF2025094022000017.tif91164

[0030] Furthermore, the present disclosure provides recombinant viral vectors and non-viral vectors comprising a nucleic acid encoding an IL27Rα binding molecule of the present disclosure or a CDR of an IL27Rα binding molecule of the present disclosure.

[0031] Furthermore, the present disclosure provides host cells comprising a recombinant viral vector and a non-viral vector comprising a nucleic acid encoding an IL27Rα binding molecule of the present disclosure or a CDR of an IL27Rα binding molecule of the present disclosure.

[0032] Furthermore, the present disclosure provides host cells comprising a recombinant viral vector and a non-viral vector comprising a nucleic acid encoding an IL27Rα binding molecule of the present disclosure or a CDR of an IL27Rα binding molecule of the present disclosure.

[0033] Furthermore, the present disclosure provides a kit comprising an IL27Rα binding molecule of the present disclosure.

[0034] In another aspect, the present disclosure provides a construct for identifying IL27Rα-expressing cells, wherein the IL27Rα-binding molecule is conjugated to one or more imaging agents, optionally via a chemical linker or a polypeptide linker. Further, the present disclosure provides the aforementioned method of use in identifying IL27Rα-expressing cells in a subject, the method comprising administering to a subject in need of treatment an effective amount of an IL27Rα-binding molecule conjugated to an imaging agent, and evaluating the subject for the presence of the imaging agent conjugated to the IL27Rα-binding molecule.

[0035] In another aspect, the present disclosure provides an IL27Rα-binding molecule modified to have an extended in vivo duration of action, wherein the IL27Rα-binding molecule is conjugated to one or more carrier molecules.

[0036] The present disclosure provides an IL27Rα-binding molecule comprising a polypeptide sequence that specifically binds to the extracellular domain of IL27Rα, and methods of using the same in the isolation, depletion, or enrichment of IL27Rα-expressing cells in a biological sample. [Invention 1001] An IL27Rα-binding molecule that specifically binds to the extracellular domain of IL2Rb. [Invention 1002] The IL27Rα-binding molecule of Invention 1001, wherein the IL2Rb-binding molecule comprises a single-domain antibody (sdAb). [Invention 1003] The sdAb comprises complementarity-determining regions 1 (CDR1), CDR2, and CDR3 as shown in the rows of: TIFF2025094022000018.tif177167TIFF2025094022000019.tif151167 The IL27Rα-binding molecule of Invention 1002. [Invention 1004] The sdAb of the IL27Rα binding molecule of the present invention 1002 or 1003 having at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 98%, or at least 99% identity, or 100% identity to any one polypeptide sequence of SEQ ID NOs: 2 to 25. [The present invention 1005] The sdAb is as shown in the following table: The IL27Rα binding molecule of the present invention 1002 containing complementarity determining regions 1 (CDR1), CDR2, and CDR3 as shown in the horizontal row of TIFF2025094022000020.tif84170. [The present invention 1006] The sdAb of the IL27Rα binding molecule of the present invention 1002 having at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 98%, or at least 99% identity, or 100% identity to any one polypeptide sequence of SEQ ID NOs: 122, 126, 130, 134, 138, 142, 146, 150, 154, 158, 162, 166, 170, and 174. [The present invention 1007] The sdAb of any IL27Rα binding molecule of the present invention 1003 or 1005, which is humanized or otherwise contains CDRs grafted onto a heterologous framework. [The present invention 1008] The IL27Rα binding molecule of any one of the present inventions 1001 to 1007, further comprising a labeling agent, an imaging agent, and / or a therapeutic agent. [The present invention 1009] The IL27Rα binding molecule of any one of the present inventions 1001 to 1008 for use in the isolation, depletion, or enrichment of IL27Rα+ cells from a biological sample. [The present invention 1011] A nucleic acid sequence encoding the IL27Rα binding molecule of any one of the present inventions 1001 to 1008. [The present invention 1012] A recombinant viral vector or non-viral vector comprising the nucleic acid of the present invention 1011. [The present invention 1013] A host cell comprising the nucleic acid of the present invention 1011. [The present invention 1014] A kit comprising any one of the IL2Rb binding molecules of the present invention 1001 to 1008.

Mode for Carrying Out the Invention

[0037] Detailed Description of the Invention Introduction For a more facile understanding of the present disclosure, certain terms and phrases are defined below and throughout this specification. The definitions provided herein are non-limiting and must be construed in view of the knowledge known to those of ordinary skill in the art.

[0038] Before describing the methods and compositions, it is to be understood that the present disclosure is not limited to the particular methods or compositions described, and accordingly, may, of course, vary.

[0039] Where a range of values is provided, each value between the upper and lower limits of that range is also specifically disclosed to the extent of one tenth of the unit of the lower limit, unless the context clearly dictates otherwise. Each narrow range between any stated value or intervening value within the stated range and any other stated value or intervening value within the stated range is encompassed by the present invention. The upper and lower limits of these narrow ranges may independently be included in or excluded from the range, and each range that includes one or neither or both of the limits of the narrow range is also encompassed by the present invention, subject to any specifically excluded limit set forth in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of the included limits are also included in the present invention.

[0040] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, some possible and preferred methods and materials will now be described. All publications mentioned herein are incorporated herein by reference for the purpose of disclosing and describing the methods and / or materials described in the cited publications.

[0041] It should be noted that the singular forms “a,” “an,” and “the” as used in this specification and the appended claims include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to “a cell” includes a plurality of such cells, and reference to “the peptide” includes reference to one or more peptides and their equivalents known to those of ordinary skill in the art, such as polypeptides.

[0042] The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates and it may be necessary to independently confirm the actual publication dates.

[0043] Throughout this disclosure, it will be understood that amino acids are referred to according to the one-letter or three-letter notations. For the convenience of the reader, the one-letter and three-letter notations of amino acids are shown in Table 5 below.

[0044] (Table 5) Abbreviations of Amino Acids TIFF2025094022000021.tif110128

[0045] Standard methods in molecular biology are described in the scientific literature (see, e.g., Sambrook and Russell (2001) Molecular Cloning, 3rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.; and Ausubel, et al. (2001) Current Protocols in Molecular Biology, Vols. 1-4, John Wiley and Sons, Inc. New York, N.Y. These describe cloning and DNA mutagenesis in bacterial cells (Volume 1), cloning in mammalian cells and yeast (Volume 2), glycoconjugates and protein expression (Volume 3), and bioinformatics (Volume 4)). The scientific literature describes protein purification methods including immunoprecipitation, chromatography, electrophoresis, centrifugation, and crystallization, as well as chemical analysis, chemical modification, post-translational modification, production of fusion proteins, and protein glycosylation (see, e.g., Coligan, et al. (2000) Current Protocols in Protein Science, Vols. 1-2, John Wiley and Sons, Inc., NY).

[0046] Definitions Unless otherwise defined, the following terms are intended to have the meanings set forth below. Other terms are defined elsewhere throughout this specification.

[0047] Activate: As used herein, the term "activate" is used with respect to a receptor or receptor complex to reflect a biological effect, either directly and / or by involvement in a multi-component signaling cascade resulting from binding of an agonist ligand to the receptor in response to ligand binding.

[0048] Activity: As used herein, the term "activity" is used to describe a property of a molecule with respect to a test system (e.g., an assay), or a biological or chemical property (e.g., the degree of binding of a molecule to another molecule) or a physical property (e.g., alteration of cell membrane potential) of a material or cell. Examples of such biological functions include, but are not limited to, the catalytic activity of a biological agent, intracellular signaling, gene expression, the ability to stimulate cell proliferation, and the ability to modulate immunological activities such as the inflammatory response. "Activity" is typically expressed as the level of biological activity per unit of test agent, e.g., [catalytic activity] / [mg protein], [immunological activity] / [mg protein], international units of activity (IU), [STAT5 phosphorylation] / [mg protein], [proliferation] / [mg protein], plaque forming units (pfu), etc. As used herein, the term "proliferative activity" refers to an activity that promotes cell growth and replication, including dysregulated cell division, such as that observed in neoplastic diseases, inflammatory diseases, fibrosis, dysplasia, cell transformation, metastasis, and angiogenesis.

[0049] Administer / Administration: The terms "administer" and "administering" are used interchangeably herein to refer to the act of contacting a subject with an agent (e.g., an IL27Rα binding molecule or an engineered cell expressing an IL27Rα binding molecule, a chemotherapeutic agent, an antibody, or a pharmaceutical formulation comprising one or more of the foregoing) in vitro, in vivo, or ex vivo with a cell, tissue, organ, or biological fluid of the subject. Administration of the agent can be accomplished by any of a variety of methods recognized in the art, including, but not limited to, topical administration, intravascular injection (including intravenous infusion or intraarterial injection), intradermal injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, intracranial injection, intratumoral injection, transdermal delivery, transmucosal delivery, iontophoresis delivery, intralymphatic injection, intragastric infusion, intraprostatic injection, intravesical infusion (e.g., into the bladder), inhalation (e.g., using a respiratory inhaler including a dry powder inhaler), intraocular injection, intraperitoneal injection, intralesional injection, intraovarian injection, intracerebral infusion or injection, intracerebroventricular injection (ICVI), etc. The term "administration" includes contact of the agent with a cell, tissue, or organ, as well as contact of the agent with a fluid in contact with a cell, tissue, or organ.

[0050] Affinity: As used herein, the term "affinity" refers to the degree of specific binding between a first molecule (e.g., a ligand) and a second molecule (e.g., a receptor), and is measured by the equilibrium dissociation constant K off , which is the ratio of the dissociation rate constant (k on ) of the molecule and its target to the association rate constant (k D ) of the molecule and its target.

[0051] Agonist: As used herein, the term "agonist" refers to a first agent that specifically binds to a second agent (the "target") and interacts with the target to cause or promote an increase in the activation of the target. In some cases, an agonist is one or more genes, proteins, ligands, receptors, biological pathways, or cell cycle arrest or cell death, e.g., apoptosis, that can modulate cell activation, enhance activation, increase the sensitivity of the cell to activation by the second agent, or result in cell proliferation, or upregulate the expression of a pathway that results in cell death, and is an activator of a receptor protein. In some embodiments, an agonist is an agent that binds to a receptor, changes the receptor state, and as a result, produces a biological response that mimics the effect of the receptor's endogenous ligand. The term "agonist" includes partial agonists, full agonists, and superagonists. An agonist may be called a "full agonist" or a "partial agonist" when such agonist leads to a substantially complete biological response (i.e., a response associated with a natural ligand / receptor binding interaction) induced by the receptor under study. A "superagonist" is a type of agonist that can produce a maximal response greater than that of the endogenous agonist for a target receptor, and thus has an activity greater than 100% of the native ligand. A superagonist is typically a synthetic molecule that exhibits a response greater than 110%, or greater than 120%, or greater than 130%, or greater than 140%, or greater than 150%, or greater than 160%, or greater than 170% of the response of an evaluable quantitative or qualitative parameter of the native molecule when evaluated at a similar concentration in a comparable assay. It should be noted that the biological effects associated with full agonists may differ in degree and / or type from the biological effects of partial agonists or superagonists. In contrast to an agonist, an antagonist can specifically bind to a receptor but does not initiate a signal cascade, typically a signal cascade initiated by the receptor, and may modify the agonistic action at that receptor. An inverse agonist is an agent that produces a pharmacological response opposite to that of an agonist.

[0052] Antagonist: As used herein, the terms "antagonist" or "inhibitor" refer to a molecule that counteracts the action of an agonist. An antagonist blocks, reduces, inhibits, or neutralizes the activity of an agonist and, in the absence of the specified agonist, can also block, inhibit, or reduce the constitutive activity of a target, e.g., a target receptor. An inhibitor is a molecule that, for example, decreases, blocks, prevents, delays, or inactivates, desensitizes, or down-regulates a cell, or a biological pathway including a gene, protein, ligand, receptor, immune checkpoint pathway, or a biological pathway including a gene, protein, ligand, receptor, immune checkpoint pathway.

[0053] Antibody: As used herein, the term "antibody" collectively refers to immunoglobulin derivatives including, but not limited to, (a) glycosylated or non-glycosylated immunoglobulins that specifically bind to a target molecule, and (b) antibody fragments such as single-domain antibodies. In some embodiments, the immunoglobulin derivative competes with the originating immunoglobulin in binding to the target molecule. The term "antibody" is not limited to immunoglobulins derived from any particular species and includes antibodies from cartilaginous fish including, but not limited to, mouse, human, horse, camelid, and shark. The term "antibody" includes antibodies that can be isolated from a natural source or an animal after immunization with an antigen, as well as monoclonal antibodies, bispecific antibodies, trispecific antibodies, chimeric antibodies, humanized antibodies, human antibodies, CDR-grafted antibodies, veneered antibodies, or deimmunized (e.g., for the purpose of removing T cell epitopes) antibodies, camelized (in the case of VHH), or molecules containing antibody binding domains (e.g., CDRs) on a non-immunoglobulin scaffold. The term "antibody" should not be construed as limited to any particular synthetic means and includes natural antibodies that can be isolated from a natural source, as well as antibodies isolated from transgenic animals into which a human immunoglobulin gene has been introduced or hybridomas prepared therefrom, antibodies isolated from host cells transformed with a nucleic acid construct that results in the expression of the antibody, and engineered antibody molecules prepared by "recombinant" means including antibodies isolated from combinatorial antibody libraries including phage display libraries. In one embodiment, an "antibody" is a mammalian immunoglobulin of the IgG1, IgG2, IgG3, or IgG4 class. In some embodiments, an antibody is a "full-length antibody" that includes variable and constant domains that provide binding and effector functions. As used herein, the term "single-domain antibody" (sdAb) refers to an antibody fragment consisting of a monomeric variable antibody domain that specifically binds to an antigen and can compete in binding with the originating parental antibody. The term "single-domain antibody" includes scFv and VHH molecules.As used herein, the term "VHH" typically refers to a single domain antibody derived from an antibody of a camelid animal (including camels, llamas, and alpacas) obtained from immunization of a camelid animal (see, e.g., Hamers-Casterman, et al. (1993) Nature 363:446-448). VHHs are also referred to as heavy chain antibodies or Nanobodies®. Single domain antibodies may also be derived from non-mammalian sources such as VHHs obtained from immunization of IgNAR antibodies of cartilaginous fish including, but not limited to, sharks.

[0054] Biological sample: As used herein, the term "biological sample" or "sample" refers to a sample obtained from (or derived from) a subject. By way of example, biological samples include materials selected from the group consisting of body fluids, blood, whole blood, plasma, serum, mucous secretions, saliva, cerebrospinal fluid (CSF), bronchoalveolar lavage fluid (BALF), eye fluids (e.g., vitreous humor, aqueous humor), lymph, lymph node tissue, spleen tissue, bone marrow, tumor tissue, including fractions enriched in immunoglobulins derived from one or more of such tissues or cell type-specific, enriched fractions.

[0055] IL27Rα cells: The terms "IL27Rα cells", "IL27Rα-expressing cells", "IL27Rα-positive cells", and "IL27Rα+" cells are used interchangeably herein to refer to cells that express and display the IL27Rα antigen on the extracellular surface of the cell membrane. Similarly, the terms "IL27Rα-negative cells", "IL27Rα- cells" are used interchangeably herein to describe cells that do not express and do not display the IL27Rα antigen on the cell surface.

[0056] CDR: As used herein, the term "CDR" or "complementary determining region" is intended to mean the discontinuous antigen-binding sites found within the variable regions of both the heavy chain immunoglobulin polypeptide and the light chain immunoglobulin polypeptide. CDRs are described by Kabat et al., J. Biol. Chem. 252:6609-6616 (1977); Kabat, et al., U.S. Department of Health and Human Services publication entitled "Sequences of proteins of immunological interest" (1991) (also referred to herein as "Kabat 1991" or "Kabat"); Chothia, et al. (1987) J. Mol. Biol. 196:901-917 (also referred to herein as "Chothia"); and MacCallum, et al. (1996) J. Mol. Biol. 262:732-745, and the definitions include overlaps or subsets of amino acid residues when compared to each other. Nevertheless, for purposes of referring to the CDRs of an antibody or a grafted antibody or variants thereof, the application of either definition is intended to be within the scope of the terms as defined and used herein. In the context of the present disclosure, unless otherwise specified, the numbering of CDR positions is assigned according to Kabat, or a hybrid of the Kabat and Chothia numbering rules.

[0057] Equivalent: As used herein, the term "equivalent" is used to describe the degree of difference between two measurements of an evaluable quantitative or qualitative parameter. For example, if a first measurement of an evaluable quantitative parameter and a second measurement of an evaluable parameter do not deviate beyond the range recognized by one of ordinary skill in the art as creating a statistically significant difference between the two results in this situation, the two measurements will be considered "equivalent." In some cases, a measurement may be considered "equivalent" if it deviates less than 35%, or less than 30%, or less than 25%, or less than 20%, or less than 15%, or less than 10%, or less than 7%, or less than 5%, or less than 4%, or less than 3%, or less than 2%, or less than 1% from another measurement. In certain embodiments, a measurement is equivalent to a standard if it deviates less than 15%, or less than 10%, or less than 5% from the standard.

[0058] Conservative amino acid substitution: As used herein, the term "conservative amino acid substitution" refers to an amino acid exchange in which a particular amino acid is changed to another amino acid having similar biochemical properties (e.g., charge, hydrophobicity, and size). For example, amino acids in each of the following groups: (1) hydrophobic amino acids: alanine, isoleucine, leucine, tryptophan, phenylalanine, valine, proline, and glycine; (2) polar amino acids: glutamine, asparagine, histidine, serine, threonine, tyrosine, methionine, and cysteine; (3) basic amino acids: lysine and arginine; and (4) acidic amino acids: aspartic acid and glutamic acid can be considered conservative amino acids of each other.

[0059] Derived from: As used herein, the term "derived from" in the context of an amino acid sequence is intended to indicate that a polypeptide or nucleic acid has a sequence based on the sequence of a reference polypeptide or nucleic acid, and is not intended to be limiting with respect to the source or method by which the protein or nucleic acid is made. By way of example, the term "derived from" includes homologs or variants of the reference amino acid or DNA sequence.

[0060] Effective concentration (EC): As used herein, the term "effective concentration" or its abbreviation "EC" is used synonymously to refer to the concentration of an agent in an amount sufficient to vary a particular parameter in a test system. The abbreviation "E" refers to the magnitude of a particular biological effect observed in this test system when the test system is exposed to the test agent. When the magnitude of the response is expressed as a function of the concentration of the test agent ("C"), the abbreviation "EC" is used. In the context of a biological system, the term Emax refers to the maximum magnitude of a particular biological effect observed in response to the saturating concentration of an activating test agent. When the abbreviation EC is shown with a subscript (e.g., EC 40 , EC 50 etc.), the subscript refers to the percent of Emax of the biological response observed at this concentration. For example, the concentration of a test agent sufficient to induce a measurable biological parameter in a test system that is 30% of the maximum level of such a measurable biological parameter in response to such a test agent is referred to as the "EC 30 " of the test agent for such a biological parameter. Similarly, the term "EC 100 " is used to indicate the effective concentration of an agent that results in the maximum (100%) response of a measurable parameter in response to such an agent. Similarly, EC 50 (commonly used in the field of pharmacokinetics)The term refers to the concentration of an agent sufficient to bring about a maximum half (about 50%) change in a measurable parameter. The term "saturation concentration" refers to the maximum amount of a test agent that can be dissolved in a standard volume of a particular solvent (e.g., water) under standard conditions of temperature and pressure. In pharmacokinetics, the saturation concentration of a drug is typically used to indicate a sufficient concentration of the drug such that all available receptors are occupied by the drug, and EC 50 is the drug concentration that produces a maximum half-effect.

[0061] Concentrated: As used herein, the term "enriched" refers to a sample that has been non-naturally engineered such that (a) the species of interest (e.g., a molecule or a cell) is present at a concentration higher (e.g., at least 3-fold, or at least 5-fold, or at least 10-fold, or at least 50-fold, or at least 100-fold, or at least 1000-fold) than the concentration of that species in a starting sample, e.g., a biological sample (e.g., a sample in which the molecule naturally occurs, or a sample that exists after administration), or (b) the molecule is present at a concentration higher than the environment in which it was made (e.g., a recombinantly modified bacterial or mammalian cell).

[0062] Extracellular domain: As used herein, the term "extracellular domain" or its abbreviation "ECD" refers to the portion of a cell surface protein (e.g., a cell surface receptor) that is outside the plasma membrane of a cell. The cell surface protein may be a transmembrane protein, a cell surface protein, or a membrane-bound protein.

[0063] Identity: As used herein with respect to polypeptide or DNA sequences, the term "identity" refers to subunit sequence identity between two molecules. When the subunit positions of both molecules are occupied by the same monomeric subunit (i.e., the same amino acid residue or nucleotide), the molecules are identical at that position. Similarity between two amino acid sequences or two nucleotide sequences is a function of the number of identical positions. Generally, these sequences are aligned to obtain the highest order of match. If necessary, identity can be calculated using published techniques and widely available computer programs such as the BLAST 2.0 algorithm described in Altschul et al. (1990) J. Mol. Biol. 215: 403-410 and Altschul, et al. (1977) Nucleic Acids Res. 25: 3389-3402. Software for performing BLAST analysis can be publicly obtained through the website of the National Center for Biotechnology Information (NCBI). This algorithm involves identifying high scoring sequence pairs (HSPs) by first identifying short words of length W in the query sequence that match or align with a positive value threshold score "T" when aligned with words of the same length in the database sequence. T is called the neighborhood word score threshold (Altschul et.al., supra). These initial neighborhood word hits serve as seeds to initiate a search for longer HSPs that contain them. The word hits are then extended in both directions along each sequence as long as the cumulative alignment score increases. The cumulative score is calculated using the parameters "M" (reward score for a pair of matching residues; always, >0) and "N" (penalty score for a mismatched residue; always, <0) for nucleotide sequences. For amino acid sequences, a scoring matrix is used to calculate the cumulative score.(a) When the cumulative alignment score has decreased by an amount X from its maximum achievable value; when the cumulative score has become zero or less due to the accumulation of one or more residue alignments with negative scores; or (b) when reaching the end of either array, the extension of word hits in each direction stops. The BLAST algorithm parameters "W", "T", and "X" determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) functions similarly, but by default, uses a word size of 28 ("W"), an expectation value of 10 ("E"), M = 1, N = -2, and comparison of both strands. For amino acid sequences, the BLASTP program, by default, uses a word size of 3 (W), an expectation value of 10 (E), and the BLOSUM62 scoring matrix (see Henikoff & Henikoff, (1989) PNAS(USA) 89:10915-10919).

[0064] In an amount sufficient to effect a response: As used herein, the phrase "in an amount sufficient to elicit a response" refers to an amount of a test agent sufficient to effect a detectable change in the level of an indicator (e.g., a baseline level) measured prior to application of the test agent to a test system and the level of the indicator measured after application. In some embodiments, the test system is a cell, tissue, or organism. In some embodiments, the test system is an in vitro test system such as a fluorescence assay. In some embodiments, the test system is an in vivo system involving measurement of a change in the level of a parameter of a cell, tissue, or organism that reflects a biological function before and after application of a test agent to the cell, tissue, or organism. In some embodiments, the indicator reflects the biological function or developmental state of a cell evaluated in an assay in response to administration of an amount of a test agent. In some embodiments, the test system involves measurement of a change in the level of an indicator of a cell, tissue, or organism that reflects a biological state before and after application of one or more test agents to the cell, tissue, or organism. The term "in an amount sufficient to effect a response" may be sufficient to be a therapeutically effective amount, but may also be more or less than a therapeutically effective amount.

[0065] Inhibitor: As used herein, the term "inhibitor" refers to a molecule that, for example, reduces, blocks, prevents, delays, inactivates, desensitizes, or down-regulates the activity of a gene, protein, ligand, receptor, or cell. An inhibitor can also be defined as a molecule that reduces, blocks, or inactivates the constitutive activity of a cell or organism.

[0066] Intracellular domain: As used herein, the term "intracellular domain" or its abbreviation "ICD" refers to the portion of a cell surface protein (e.g., a cell surface receptor) that is inside the plasma membrane of a cell. The ICD may include the entire cytoplasmic portion of a transmembrane or membrane-bound protein, or may include intracellular proteins.

[0067] Isolated: As used herein, the term "isolated" is used with respect to a polypeptide of interest that is in an environment different from its natural environment if it occurs naturally. "Isolated" is intended to include a polypeptide of interest that is substantially enriched and / or the polypeptide of interest is partially or substantially purified in a sample. If the polypeptide is not natural, "isolated" indicates that the polypeptide is separated from the environment in which it was synthesized, e.g., isolated from a recombinant cell culture containing cells engineered to express the polypeptide, or isolated by a solution resulting from solid-phase synthesis means.

[0068] Kabat numbering: As used herein, the term "Kabat numbering" is recognized in the art and refers to a system for numbering amino acid residues that are more variable (e.g., hypervariable) than other amino acid residues in the heavy and light chain regions of immunoglobulins (Kabat, et al., (1971) Ann. NY Acad. Sci. 190:382-93; Kabat, et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242). As used herein, the term "Chothia numbering" is recognized in the art and refers to a system for numbering amino acid residues based on the location of structural loop regions (Chothia et al. 1986, Science 233:755-758; Chothia & Lesk 1987, JMB 196:901-917; Chothia et al. 1992, JMB 227:799-817). For the purposes of the present disclosure, unless specifically specified otherwise, the positions of CDR2 and 3 in the variable region of an antibody follow Kabat numbering, or simply "Kabat". The position of CDR1 in the variable region of an antibody follows a hybrid of the Kabat and Chothia numbering schemes.

[0069] Ligand: As used herein, the term "ligand" refers to a molecule that specifically binds to a receptor and causes a change in the receptor, either to alter the activity of the receptor or the response of the cell expressing that receptor. In one aspect, the term "ligand" refers to a molecule or complex thereof that can act as an agonist or antagonist of a receptor. The term "ligand" as used herein includes both natural and synthetic ligands. "Ligand" also includes small molecules, cytokines, and peptidomimetics of antibodies. The complex of a ligand and a receptor is referred to as a "ligand-receptor complex". A ligand may comprise one domain of a polypeptide or fusion protein (e.g., either domain of an antibody / ligand fusion protein).

[0070] Modulate: As used herein, terms such as "modulate", "modulation", etc. refer to the ability of a test agent to cause, directly or indirectly, a positive or negative response in a system, including a biological system, or a biochemical pathway. The term "modulator" includes both agonists (including partial agonists, full agonists, and superagonists) and antagonists.

[0071] Nucleic acid: The terms "nucleic acid", "nucleic acid molecule", "polynucleotide", etc. are used interchangeably herein to refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof. Non-limiting examples of polynucleotides include linear and circular nucleic acids, messenger RNA (mRNA), complementary DNA (cDNA), recombinant polynucleotides, vectors, probes, primers, and the like.

[0072] Functionally linked: The term "functionally linked" is used herein to refer to the relationship between molecules, typically polypeptides or nucleic acids, arranged within a construct such that each of the functions of the component molecules is retained, but the activity of the individual components of the construct can be adjusted, either positively or negatively, by being functionally linked. For example, functionally linking a polyethylene glycol (PEG) molecule to a wild-type protein may result in a construct in which the biological activity of the protein is decreased relative to the wild-type molecule. However, they are still considered to be functionally linked. When the term "functionally linked" is applied to the relationship of multiple nucleic acid sequences encoding different functions, when the multiple nucleic acid sequences are combined into one nucleic acid molecule, for example, when this nucleic acid molecule is introduced into a cell using recombinant technology, it provides a nucleic acid that can transcribe and / or translate a specific nucleic acid sequence within the cell. For example, if a nucleic acid sequence encoding a signal sequence that facilitates the secretion of a polypeptide expresses a preprotein, it can be considered to be functionally linked to the DNA encoding the polypeptide. If a promoter or enhancer affects the transcription of a sequence, it is considered to be functionally linked to the coding sequence. Or, if a ribosome binding site is arranged to facilitate translation, it is considered to be functionally linked to the coding sequence. Generally, in the context of nucleic acid molecules, the term "functionally linked" means that the linked nucleic acid sequences are contiguous, and in the case of a secretion leader or attached subdomain of a molecule, contiguous and in the reading phase. However, certain genetic elements, such as enhancers, may function away from the sequences to which they exert an effect and need not be contiguous to those sequences, but may still be considered to be functionally linked.

[0073] Parent polypeptide: As used herein, the terms "parent polypeptide" or "parent protein" are used interchangeably to designate the source of a second polypeptide (e.g., a derivative, mutein or variant) that has been modified with respect to a first "parent" polypeptide. In some cases, the parent polypeptide is a wild-type or native protein. In some cases, the parent polypeptide may be a modified form of a native protein that has been further modified. The term "parent polypeptide" may refer to the polypeptide itself, or to a composition comprising the parent polypeptide (e.g., a glycosylated or PEGylated form and / or a fusion protein comprising the parent polypeptide).

[0074] Partial agonist: As used herein, the term "partial agonist" refers to a specifically binding molecule that binds to and activates a particular receptor, but has only partial activation of the receptor as compared to a full agonist. A partial agonist may exhibit both agonist and antagonist activity. For example, when both a full agonist and a partial agonist are present, the partial agonist competes with the full agonist for receptor binding and, as a result, acts as a competitive antagonist by causing a net decrease in receptor activation as compared to receptor contact with the full agonist in the absence of the partial agonist. When an insufficient amount of endogenous ligand is present, a partial agonist can be used to activate the receptor to produce a desired maximal sub-response in a subject. Alternatively, when an excess amount of endogenous ligand is present, a partial agonist can reduce over-stimulation of the receptor. The maximal response (E) produced by a partial agonist max) is referred to as its intrinsic activity and may be expressed on a percentage scale when a full agonist produces a 100% response. A partial agonist is greater than 10% but less than 100%, or greater than 20% but less than 100%, or greater than 30% but less than 100%, or greater than 40% but less than 100%, or greater than 50% but less than 100%, or greater than 60% but less than 100%, or greater than 70% but less than 100%, or greater than 80% but less than 100%, or greater than 90% but less than 100% of the activity of the reference polypeptide when evaluated at a similar concentration in a particular assay system.

[0075] Polypeptide: As used herein, the terms "polypeptide," "peptide," and "protein" are used interchangeably herein and refer to a polymeric form of amino acids of any length that can include amino acids specified by the genetic code and amino acids not specified by the genetic code, chemically or biochemically modified or derivatized amino acids, and polypeptides having modified polypeptide backbones. The term polypeptide includes, but is not limited to, fusion proteins having heterologous amino acid sequences; fusion proteins having heterologous leader sequences and homologous leader sequences; fusion proteins having or not having an N-terminal methionine residue; fusion proteins having amino acid sequences that facilitate purification, such as chelating peptides; fusion proteins having immunologically tagged proteins; fusion proteins including peptides having immunologically active polypeptide fragments (e.g., antigenic diphtheria or tetanus toxin or toxoid fragments).

[0076] Receptor: As used herein, the term "receptor" refers to a polypeptide having a domain that specifically binds to a ligand, wherein at least one biological property of the polypeptide changes upon binding of the ligand. In some embodiments, the receptor is a cell membrane-bound protein that includes an extracellular domain (ECD) and a membrane-bound domain that serves to anchor the ECD to the cell surface. In some embodiments of cell surface receptors, the receptor is a transmembrane polypeptide in which an intracellular domain (ICD) and an extracellular domain (ECD) are linked by a transmembrane domain of the cell membrane, commonly referred to as a transmembrane domain (TM). When a cognate ligand binds to the receptor, a conformational change in the receptor occurs, resulting in a measurable biological effect. In some cases, when the receptor is a transmembrane polypeptide that includes an ECD, a TM, and an ICD, binding of the ligand to the ECD results in a measurable intracellular biological effect mediated by one or more domains of the ICD in response to the binding of the ligand to the ECD. In some embodiments, the receptor is a component of a multi-component complex that facilitates intracellular signaling. For example, a ligand may not be associated with any intracellular signaling on its own, but binding of the ligand promotes the formation of a hetero-oligomer (including hetero-dimers, hetero-trimers, etc.) or homo-oligomer (including homo-dimers, homo-trimers, homo-tetramers, etc.) complex, resulting in a measurable biological effect within the cell, such as activation of an intracellular signaling cascade (e.g., the Jak / STAT pathway) by binding to a cell surface receptor. In some embodiments, the receptor is a transmembrane single-chain polypeptide that includes ECD, TM, and ICD domains, and the ECD, TM, and ICD domains are derived from the same or different native receptor variants or synthetic functional equivalents thereof.

[0077] Recombinant: As used herein, the term "recombinant" is used as an adjective to refer to a method by which a polypeptide, nucleic acid, or cell has been modified using recombinant DNA technology. A "recombinant protein" is a protein produced using recombinant DNA technology and is abbreviated by using a lowercase "r" in front of the protein name to indicate the method by which the protein was produced (e.g., human growth hormone produced recombinantly is commonly abbreviated as "rhGH"). Similarly, a cell is called a "recombinant cell" if it has been modified by the incorporation (e.g., transfection, transduction, infection) of exogenous nucleic acids (e.g., ssDNA, dsDNA, ssRNA, dsRNA, mRNA, viral vector or non-viral vector, plasmid, cosmid, etc.) using recombinant DNA technology. Techniques and protocols for recombinant DNA technology, such as those found in Sambrook, et al. (1989) Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring Harbor Laboratory Press, Plainview, N.Y.) and other standard molecular biology laboratory manuals, are well known in the art.

[0078] Response: For example, the term "response" of a cell, tissue, organ, or organism encompasses a quantitative or qualitative change in an evaluable biochemical or physiological parameter (e.g., concentration, density, adhesion, proliferation, activation, phosphorylation, migration, enzyme activity, gene expression level, gene expression rate, energy consumption rate, level or state of differentiation), and this change is correlated with activation, stimulation, or treatment by an exogenous agent or an internal mechanism such as genetic programming, or contact with an exogenous agent or an internal mechanism such as genetic programming. In certain situations, terms such as "activation", "stimulation", etc. refer to cell activation that is regulated by an internal mechanism and also by external or environmental factors, whereas terms such as "inhibition", "downregulation", etc. refer to the opposite effect. A "response" may be evaluated in vitro, for example, by using assay systems, surface plasmon resonance, enzyme activity, mass spectrometry, amino acid or protein sequencing techniques. A "response" may be evaluated quantitatively in vivo by assessing objective physiological parameters such as body temperature, body weight, tumor mass, blood pressure, results of X-ray or other imaging techniques, or qualitatively by reported subjective emotional changes such as well-being, depression, excitement, or pain. In some embodiments, the proliferation level of CD3-activated primary human T cells may be evaluated in a bioluminescence assay that generates a luminescence signal proportional to the amount of ATP present, which is proportional to the number of live cells present during culture, as described in Crouch, et al. (1993) J. Immunol. Methods 160: 81-8, and may be evaluated generally in accordance with the instructions provided by the manufacturer using a commercially available assay, for example, the CellTiter-Glo® 2.0 Cell Viability Assay or the CellTiter-Glo® 3D Cell Viability kit, commercially available from Promega Corporation, Madison WI 53711 under catalog numbers G9241 and G9681.In some embodiments, the level of T cell activation in response to administration of a test agent may be confirmed by flow cytometry as described when confirmed by the level of STAT (e.g., STAT1, STAT3, STAT5) phosphorylation according to methods well known in the art.

[0079] Significantly reduced binding: As used herein, the term "showing significantly reduced binding" is used with respect to variants of a first molecule (e.g., a ligand or antibody) that show a significant reduction in affinity for a second molecule (e.g., a receptor or antigen) compared to the parental form of the first molecule. For an antibody variant, the variant "shows significantly reduced binding" if the variant binds to the native receptor with an affinity that is less than 20%, or less than about 10%, or less than about 8%, or less than about 6%, or less than about 4%, or less than about 2%, or less than about 1%, or less than about 0.5% of the affinity of the parental antibody from which the variant originated. Similarly, for a variant ligand, the variant ligand "shows significantly reduced binding" if the affinity of the variant ligand is less than 20%, or less than about 10%, or less than about 8%, or less than about 6%, or less than about 4%, or less than about 2%, or less than about 1%, or less than about 0.5% of the affinity of the parental ligand from which the variant ligand originated for binding to the receptor. Similarly, for a variant receptor, the variant ligand "shows significantly reduced binding" if the affinity of the variant receptor is less than 20%, or less than about 10%, or less than about 8%, or less than about 6%, or less than about 4%, or less than about 2%, or less than about 1%, or less than about 0.5% of the affinity of the parental receptor from which the variant receptor originated for binding.

[0080] Small molecule: The term "small molecule" refers to a compound (typically a pharmaceutically active compound) having a molecular weight of less than about 10 kDa, less than about 2 kDa, or less than about 1 kDa. Small molecules include, but are not limited to, inorganic molecules, organic molecules, organic molecules containing inorganic molecules, molecules containing radioactive atoms, and synthetic molecules. The term "small molecule" is well understood by those skilled in the art of pharmacy and is typically used to distinguish organic compounds from biologics.

[0081] Specifically bind: As used herein, the term "specifically binds" refers to the degree of affinity that a first molecule exhibits for a second molecule. In the context of a binding pair (e.g., ligand / receptor, antibody / antigen), the first molecule of the binding pair is said to specifically bind to the second molecule of the binding pair when the first molecule of the binding pair does not bind in significant amounts to other components present in the sample. The first molecule of the binding pair is said to specifically bind to the second molecule of the binding pair when the affinity of the first molecule for the second molecule is at least 2-fold, or at least 5-fold, or at least 10-fold, or at least 20-fold, or at least 100-fold the affinity of the first molecule for other components present in the sample. In certain embodiments, when the first molecule of the binding pair is an antibody, the equilibrium dissociation constant between the antibody and the antigen is, for example, about 10 6 M or greater, or about 10 8 M or greater, or about 10 10 M or greater, or about 10 11 M or greater, about 10 12 M or greater, then the antibody specifically binds to the antigen (or protein, antigen, ligand, or epitope of the antigen determinant of the receptor). In one embodiment, when the ligand is an IL27Rα-binding sdAb and the receptor comprises IL27Rα, the equilibrium dissociation constant of the IL27Rα-binding sdAb / IL27RαECD is about 10 5 M or greater, or about 10 6 M or greater, or about 10 7M-mode, or approximately 10 8 M-mode, or approximately 10 9 M-mode, or approximately 10 10 M-mode, or approximately 10 11In the case of M-mode, the IL27Rα-binding sdAb binds specifically. Specific binding can be evaluated using techniques known in the art, including but not limited to competitive ELISA assays, radioligand binding assays (e.g., saturation binding, Scatchard plots, non-linear curve fitting programs, and competitive binding assays); non-radioligand binding assays (e.g., fluorescence polarization (FP), fluorescence resonance energy transfer (FRET)); liquid-phase ligand binding assays (e.g., real-time polymerase chain reaction (RT-qPCR), and immunoprecipitation); and solid-phase ligand binding assays (e.g., multiwell plate assays, on-bead ligand binding assays, on-column ligand binding assays, and filter assays)), as well as surface plasmon resonance assays (e.g., see Drescher et al., (2009) Methods Mol Biol 493:323-343 and commercially available measurement devices such as Biacore 8K, Biacore 8K+, Biacore S200, Biacore T200 (Cytiva, 100 Results Way, Marlborough MA 01752)). In some embodiments, the disclosure provides molecules that specifically bind to an IL27Rα isoform (e.g., an IL27Rα-binding sdAb). As used herein, the binding affinity of an IL27Rα-binding molecule for IL27Rα may be determined and / or quantified by surface plasmon resonance ("SPR"). When evaluating the binding affinity of an IL27Rα-binding molecule for IL27Rα, one member of the binding pair may be immobilized and the other member of the binding pair may be provided in the mobile phase.In some embodiments, a sensor chip on which a protein of interest is immobilized on the surface is conjugated with a substance that facilitates the binding of the protein of interest, such as a nitrilotriacetic acid (NTA)-derivatized surface plasmon resonance sensor chip (e.g., Sensor Chip NTA available from Cytiva Global Life Science Solutions USA LLC, Marlborough MA under catalog number BR100407), an anti-His tag antibody (e.g., anti-histidine CM5 chip commercially available from Cytiva, Marlborough MA), protein A, or biotin. As a result, it is often necessary to modify the protein to bind to the substance conjugated to the chip surface to evaluate the binding. For example, one member of the binding pair to be evaluated was conjugated to NTA by incorporating a chelating peptide containing a polyhistidine sequence (e.g., 6xHis (SEQ ID NO:195) or 8xHis (SEQ ID NO:196)) for retention on the chip. In some embodiments, the IL27Rα binding molecule may be immobilized on the chip and IL27Rα (or an ECD fragment thereof) may be provided in the mobile phase. Or, IL27Rα (or an ECD fragment thereof) may be immobilized on the chip and the IL27Rα binding molecule may be provided in the mobile phase. In either case, it should be noted that modifying some proteins for immobilization on the coated SPR chip may interfere with the binding characteristics of one or both components of the binding pair to be evaluated by SPR. In such cases, it may be necessary to switch the mobile and binding elements of the binding pair or use a chip with a binder that facilitates non-interfering conjugation of the protein to be evaluated.Alternatively, when evaluating the binding affinity of an IL27Rα-binding molecule to IL27Rα using SPR, the IL2Rb-binding molecule may be derivatized by addition of a C-terminal polyHis sequence (e.g., 6xHis (SEQ ID NO:195) or 8xHis (SEQ ID NO:196)) and immobilized on an NTA-derivatized sensor chip, and the IL27Rα receptor subunit whose ligand-binding affinity is being evaluated is provided in the mobile phase. Means for incorporating a polyHis sequence at the C-terminus of an IL27Rα-binding molecule generated by recombinant DNA technology are well known to those of ordinary skill in the relevant fields of biotechnology. In some embodiments, the binding affinity of an IL27Rα-binding molecule to IL27Rα using SPR generally follows the disclosure of the examples.

[0082] Subject: The terms "recipient", "individual", "subject", and "patient" are used interchangeably herein and refer to any mammalian subject, particularly a human, for whom diagnosis, treatment, or therapy is desirable. For purposes of treatment, "mammal" refers to any animal classified as a mammal, including humans, laboratory and domestic animals, and zoo, sports, or pet animals such as dogs, horses, cats, cows, sheep, goats, pigs, etc. In some embodiments, the mammal is a human.

[0083] Substantially pure: As used herein, the term "substantially pure" indicates that one component of a composition constitutes more than about 50%, or more than about 60%, or more than about 70%, or more than about 80%, or more than about 90%, or more than about 95% of the total content of the composition. A "substantially pure" protein constitutes more than about 50%, or more than about 60%, or more than about 70%, or more than about 80%, or more than about 90%, or more than about 95% of the total content of the composition.

[0084] T cell: As used herein, the term "T cell" (also "T-cell" or "T cell") is used in its conventional sense to refer to lymphocytes that differentiate in the thymus, have specific cell surface antigen receptors, and include those that control the initiation or suppression of cellular and humoral immunity, as well as those that lyse cells bearing antigens. In some embodiments, T cells include naive CD8 + T cells, cytotoxic CD8 + T cells, naive CD4 + T cells, helper T cells, such as T H 1, T H 2, T H 9, T H 11, T H 22, T FH ; regulatory T cells, such as T R 1, Treg, inducible Treg; memory T cells, such as central memory T cells, effector memory T cells, NKT cells, tumor-infiltrating lymphocytes (TIL), and including, but not limited to, CAR-T cells, recombinant modified TIL, and TCR-engineered cells, such manipulated variants of such T-cells are included, but not limited thereto. In some embodiments, T cells are T cells that express an IL27Rα isoform, which are referred to synonymously as IL27Rα cells, IL27Rα+ cells, IL27Rα T cells, or IL27Rα+ T cells).

[0085] Terminus / Terminal: As used herein in the context of polypeptide structure, "N-terminus" (or "amino terminus") and "C-terminus" (or "carboxyl terminus") refer, respectively, to the amino- and carboxyl-terminal ends of a polypeptide. In contrast, the terms "N-terminal" and "C-terminal" refer, respectively, to the relative positions in a polypeptide amino acid sequence with respect to the N-terminus and C-terminus, and may include the residues at the N-terminus and C-terminus, respectively. "Immediately N-terminal" refers to the position of the first amino acid residue relative to the second amino acid residue in a contiguous polypeptide sequence, with the first amino acid being closer to the N-terminus of the polypeptide. "Immediately C-terminal" refers to the position of the first amino acid residue relative to the second amino acid residue in a contiguous polypeptide sequence, with the first amino acid being closer to the C-terminus of the polypeptide.

[0086] Transmembrane domain: The term "transmembrane domain" or "TM" refers to the polypeptide domain of a transmembrane polypeptide (e.g., a transmembrane receptor) that is embedded in the cell membrane when the transmembrane polypeptide is bound to the cell membrane and is peptidyl-bonded to the extracellular domain (ECD) and intracellular domain (ICD) of the transmembrane polypeptide. The transmembrane domain may be homologous (naturally related) to either or both of the extracellular and / or intracellular domains, or may be heterologous (not naturally related). In some embodiments, where the receptor is a chimeric receptor that includes an intracellular domain derived from a first parental receptor and a second extracellular domain that is derived from a second different parental receptor, the transmembrane domain of the chimeric receptor is typically the transmembrane domain associated with either the ICD or ECD of the parental receptor from which the chimeric receptor originated.

[0087] Treat: The terms "treat," "treating," "treatment," and the like refer to a course of action (e.g., contacting the subject with a pharmaceutical composition comprising an IL27Rα-binding sdAb alone or in combination with adjuvants) undertaken against a subject in response to a diagnosis that the subject is suffering from a disease, disorder, or condition, or a symptom thereof, where the course of action is undertaken to temporarily or permanently eliminate, reduce, inhibit, alleviate, or ameliorate at least one of (a) the underlying cause of such disease, disorder, or condition afflicting the subject; and / or (b) at least one of the symptoms associated with such disease, disorder, or condition. In some embodiments, treating includes a course of action taken against a subject suffering from a disease, where the course of action inhibits the disease in the subject (e.g., arrests the development of, or ameliorates, one or more symptoms associated with, the disease, disorder, or condition).

[0088] Treg cell or regulatory T cell: The term "regulatory T cells," "Treg cells," or "Treg" refers to effector T cells (T eff ) that can suppress the responses of other T cells, including but not limited to CD4 + They are used interchangeably herein to refer to a type of T cell. Treg cells are typically characterized by expression of CD4 (CD4+), the CD25 subunit of the IL2 receptor (CD25+), and the transcription factor forkhead box P3 (FOXP3+) (Sakaguchi, Annu Rev Immunol 22, 531-62 (2004). In some cases, the term "conventional CD4+ T cells" refers to non-Treg CD4 + T cells are CD4 + Used to distinguish from Treg.

[0089] Variant: The terms "variant", "protein variant", or "variant protein", or "variant polypeptide" are used interchangeably herein to refer to a polypeptide that differs from a parent polypeptide by at least one amino acid modification, substitution, or deletion. The parent polypeptide may be a native or wild-type (WT) polypeptide, or a modified version of the WT polypeptide. The term variant polypeptide may refer to the polypeptide itself, a composition comprising the polypeptide, or a nucleic acid sequence encoding it. In some embodiments, the variant polypeptide comprises from about 1 to about 10, or from about 1 to about 8, or from about 1 to about 7, or from about 1 to about 5, or from about 1 to about 4, or from about 1 to about 3, or 1 to 2 amino acid modifications, substitutions, or deletions, or 1 amino acid modification, substitution, or deletion compared to the parent polypeptide. The variant may be at least about 99% identical, or at least about 98% identical, or at least about 97% identical, or at least about 95% identical, or at least about 90% identical to the parent polypeptide from which it is derived.

[0090] Wild type: As used herein, "wild-type" or "WT" or "native" means a naturally occurring amino acid sequence or nucleotide sequence, including allelic variations. Wild-type proteins, polypeptides, antibodies, immunoglobulins, IgG, etc. have an amino acid sequence or nucleotide sequence that has not been modified by human hand.

[0091] IL27Rα The IL27Rα binding molecule of the present disclosure specifically binds to the extracellular domain of IL27Rα.

[0092] Human IL27Rα In one aspect, it specifically binds to the extracellular domain of the human IL27Rα receptor subunit (hIL27Rα). hIL27Rα contains a 32 - amino - acid N - terminal signal sequence, and is expressed as a 636 - amino - acid precursor where the N - terminal signal sequence is cleaved after translation to become a 604 - amino - acid mature protein. The classical full - length hIL27Rα precursor (including the signal peptide) is a 636 - amino - acid polypeptide with the amino acid sequence: TIFF2025094022000022.tif62134.

[0093] For the purposes of the present disclosure, the numbering of the amino acid residues of the human IL27Rα polypeptide as described herein follows the numbering of this classical sequence (UniProt reference number Q6UWB1, SEQ ID NO:1). Amino acids 1 - 32 of SEQ ID NO:1 are identified as the signal peptide of hIL27Rα, amino acids 33 - 516 of SEQ ID NO:1 are identified as the extracellular domain, amino acids 517 - 537 of SEQ ID NO:1 are identified as the transmembrane domain, and amino acids 538 - 636 of SEQ ID NO:1 are identified as the intracellular domain.

[0094] For the purpose of generating an antibody that binds to the ECD of IL27Rα, immunization can be performed using the extracellular domain of hIL27Rα. The extracellular domain of hIL27Rα is a 484 - amino - acid polypeptide with the sequence: TIFF2025094022000023.tif49134.

[0095] Mouse IL27Rα In one aspect, it specifically binds to the extracellular domain of the mouse or rat IL27Rα receptor subunit (mIL27Rα). mIL27Rα contains a 24 - amino - acid N - terminal signal sequence, and is expressed as a 623 - amino - acid precursor where the N - terminal signal sequence is cleaved after translation to become a 599 - amino - acid mature protein. The classical full - length mIL27Rα precursor (including the 24 - signal peptide) has the amino acid sequence: It is a 623 - amino acid polypeptide having TIFF2025094022000024.tif62134.

[0096] For the purposes of the present disclosure, the numbering of the amino acid residues of the mIL27Rα polypeptide as described herein follows the numbering of this classical sequence (UniProt reference number O70394, SEQ ID NO:193). Amino acids 1 - 24 of SEQ ID NO:193 have been determined to be the signal peptide of mIL27Rα, amino acids 23 - 510 of SEQ ID NO:193 have been determined to be the extracellular domain, amino acids 511 - 531 of SEQ ID NO:193 have been determined to be the transmembrane domain, and amino acids 532 - 623 of SEQ ID NO:193 have been determined to be the intracellular domain.

[0097] For the purpose of generating an antibody that binds to the ECD of IL27Rα, immunization can be performed using the extracellular domain of mIL27Rα. The extracellular domain of the mIL27Rα receptor has the sequence: It is a 486 - amino acid polypeptide of TIFF2025094022000025.tif49133.

[0098] IL27Rα - binding molecules and single - domain antibodies In some embodiments, the IL27Rα - binding molecules of the present disclosure are single - domain antibodies (sdAbs). The present disclosure relates to IL27Rα - binding molecules comprising single - domain antibodies (sdAbs) that specifically bind to the extracellular domain of the human IL27Rα isoform (hIL27Rα) found on all IL27Rα - expressing cells.

[0099] A single-domain antibody (sdAb) is an antibody that contains one monomeric variable antibody domain. Similar to a full-length antibody, an sdAb can specifically bind to an epitope. The hIL27Rα-binding VHH single-domain antibody can be engineered from heavy-chain antibodies isolated from camelid mammals (e.g., camels, llamas, alpacas, and guanacos) immunized with the extracellular domain of hIL27Rα or an immunologically active fragment thereof. Descriptions of sdAbs and VHHs can be found, for example, in De Greve et al., (2019) Curr Opin Biotechnol. 61:96-101; Ciccarese, et al., (2019) Front Genet. 10:997: Chanier and Chames (2019) Antibodies (Basel) 8(1); and De Vlieger, et al. (2018) Antibodies (Basel) 8(1). Alternatively, the hIL27Rα single-domain antibody may be engineered from a heavy-chain antibody isolated from an IgNAR heavy-chain antibody isolated from cartilaginous fish immunized with the extracellular domain of hIL27Rα or an immunologically active fragment thereof. The hIL27Rα-binding sdAb may also be obtained by splitting the dimeric variable domain derived from an immunoglobulin G (IgG) isotype from other mammalian species including humans, rats, and rabbits immunized with the extracellular domain of hIL27Rα or an immunologically active fragment thereof. Currently, most sdAb research is based on the heavy-chain variable domain, but sdAbs derived from the light chain have also been shown to specifically bind to a target protein containing an antigenic immunizing sequence. Moller et al., J Biol Chem. 285(49):38348-38361, 2010.

[0100] In some embodiments, the sdAb is a VHH. A VHH is a type of sdAb that has a single monomeric heavy chain variable antibody domain. Similar to conventional antibodies, VHHs can specifically bind to specific antigens. Exemplary VHHs have a molecular weight of approximately 12 - 15 kDa, which is considerably smaller than that of conventional mammalian antibodies (150 - 160 kDa) composed of two heavy chains and two light chains. VHHs may be found in and produced from camelid mammals (e.g., camels, llamas, vicuñas, alpacas, and guanacos) that are naturally lacking light chains.

[0101] The present disclosure provides an IL27Rα binding molecule comprising a polypeptide having at least 75%, or 80%, or 90%, or 95%, or 98%, or 99% or 100% identity to any one of the polypeptides of SEQ ID NO: 2 - 25.

[0102] The present disclosure provides an IL27Rα binding molecule comprising a polypeptide having at least 75%, or 80%, or 90%, or 95%, or 98%, or 99%, or 100% identity to any one of the polypeptides of SEQ ID NO: 61 - 74.

[0103] The present disclosure provides an IL27Rα binding molecule comprising CDR1, CDR2, and CDR3 as described in the rows of Table 1 provided herein. In some embodiments, CDR1, CDR2, and CDR3 each independently may have at least 90% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to the sequences described in the rows of Table 1 provided herein and may have 0, 1, 2, or 3 amino acid changes, optionally conservative amino acid changes.

[0104] The present disclosure provides an IL27Rα binding molecule comprising CDR1, CDR2, and CDR3 as described in the rows of Table 3 provided herein. In some embodiments, CDR1, CDR2, and CDR3 each independently may comprise at least 90% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to the sequences described in the rows of Table 3 provided herein, and may have 0, 1, 2, or 3 amino acid changes, optionally conservative amino acid changes.

[0105] Experiment The single-domain antibodies of the present disclosure were obtained from camels by immunizing with the extracellular domain of the IL27Rα receptor. In accordance generally with the disclosure of the examples, the IL27Rα VHH molecules of the present disclosure were prepared. Briefly, using the ECDs of human IL27Rα and mouse IL27Rα, camels were continuously immunized over several weeks with a subcutaneous adjuvant-containing composition containing a fusion protein recombinantly produced and comprising the extracellular domain of IL27Rα, the human IgG1 hinge domain, and the human IgG1 heavy chain Fc. After immunization, RNA extracted from blood samples of the appropriate-sized VHH-hinge-CH2-CH3 species was transcribed to generate a DNA sequence, digested, and a ~400 bp fragment containing the nucleic acid sequence encoding the VHH domain was identified and isolated. To facilitate insertion into a phagemid vector in-frame with the sequence encoding the his-tag, the isolated sequence was digested with a restriction endonuclease and introduced into Escherichia coli (E. coli) by transformation to generate a phage library. Multiple rounds of phage library biopanning were performed to identify VHHs that bound to the ECD of IL27Rα (human or mouse as appropriate). Individual phage clones were isolated for periplasmic extract ELISA (PE-ELISA) in 96-well plate format and for selective binding confirmed by colorimetric measurement. IL27Rα-binding molecules that showed specific binding to the IL27Rα antigen were isolated, sequenced, and sequence analyzed to identify the VHH sequences, CDRs, and unique VHH clonotypes. As used herein, the term "clonotype" refers to a collection of binding molecules arising from the same B-cell progenitor cell, within a particular collection of antigen-binding molecules belonging to the same germline family, having the same CDR3 length and having at least 70% homology in the CDR3 sequence. The VHH molecules (anti-human IL27Rα VHHs) that showed specific binding to the hIL27RαECD antigen and the CDRs isolated from such VHHs are shown in Table 1. The VHH molecules (anti-mouse IL27Rα VHHs) that showed specific binding to the mIL27RαECD antigen and the CDRs isolated from such VHHs are shown in Table 3.The nucleic acid sequences encoding the VHHs of Table 1 and Table 3 are shown in Table 2 and Table 4, respectively.

[0106] To more fully characterize the binding properties of the VHH molecules prepared according to the foregoing and to evaluate their binding affinity, representative examples of each human VHH chronotype were subjected to surface plasmon resonance analysis generally in accordance with the disclosure of Example 5 herein. The results of these SPR studies are summarized in Table 6 below.

[0107] (Table 6) Binding of anti-hIL27Ra mono-Fc VHH (ligand) to hIL27Ra-his (Antigen: Origene, catalog number TP307012) TIFF2025094022000026.tif54168 * Both the association rate constant and the dissociation rate constant may be suppressed at Rmax > 100. If this effect is present, it is likely to be offset by the reaction rate ratio, i.e., the affinity constant.

[0108] As demonstrated by the data shown in Table 6 above, the IL27Rα VHH binding molecules showed specific binding to the antigen and exhibited a range of affinities for the IL27Rα antigen.

[0109] In some cases, due to sequence similarity or structural similarity between the extracellular domains of the IL27Rα receptor derived from various mammalian species, immunization with an antigen derived from the IL27Rα of a first mammalian species (e.g., hIL27Rα-ECD) may result in antibodies that specifically bind to the IL27Rα receptor of one or more additional mammalian species. Such antibodies are called "cross-reactive". For example, immunization of a camelid with a human-derived antigen (e.g., hIL27Rα-ECD) may result in antibodies that are cross-reactive with the mouse and human receptors. Evaluation of the cross-reactivity of antibodies against receptors from other mammalian species can be readily ascertained by those skilled in the art using methods related to the evaluation of binding affinity and / or specific binding described elsewhere in this specification, such as flow cytometry or SPR. As a result, the use of the terms "human IL27Rα VHH" or "hIL27Rα VHH" only indicates that the species of the IL27Rα antigen used for immunization of the camelid from which the VHH originated was human IL27Rα (e.g., hIL27Rα, ECD, SEQ ID NO:192), and should not be understood as a limitation regarding the specific binding affinity of the VHH for hIL27Rα molecules of other mammalian species. Similarly, the use of the terms "mouse IL27Rα VHH" or "mIL27Rα" only indicates that the species of the IL27Rα antigen used for immunization of the camelid from which the VHH originated was mouse IL27Rα (e.g., mIL27Rα ECD, SEQ ID NO:194), and should not be understood as a limitation regarding the specific binding affinity of the VHH for IL27Rα molecules of other mammalian species.

[0110] Modified form of single domain antibody CDR-grafted sdAb In some embodiments, the IL27Rα-binding sdAbs of the present disclosure are CDR-grafted IL27Rα-binding sdAbs. To generate CDR-grafted sdAbs, CDRs obtained from antibodies, heavy chain antibodies, and sdAbs derived therefrom may be grafted into a different framework as described in Saerens, et al. (2005) J. Mol Biol 352:597-607. In some embodiments, the present disclosure provides an IL27Rα-binding molecule comprising a CDR-grafted IL27Rα-binding sdAb, wherein the CDR-grafted IL27Rα-binding sdAb comprises a set of CDR1, 2, and 3 as shown in the rows of Table 3 above.

[0111] Chimeric sdAbs and humanized sdAbs Any framework region can be used together with the CDRs as described herein. In some embodiments, the IL27Rα-binding sdAb is a chimeric sdAb, in which the CDRs are derived from one species (e.g., camel), and the framework and / or constant region are derived from another species (e.g., human or mouse). In certain embodiments, the framework region is a human or humanized sequence. Accordingly, humanized IL27Rα-binding sdAbs derived from hIL27Rα-binding VHHs are considered to be within the scope of the present disclosure. Techniques for humanizing single-domain antibodies from camelids are well known in the art. See, for example, Vincke, et al. (2009) General Strategy to Humanize a Camelid Single-domain Antibody and Identification of a Universal Humanized Nanobody Scaffold J. Biol. Chem. 284(5)3273-3284.

[0112] In some embodiments, the V H H can be humanized to contain a human framework region. Humanized V HExamples of human germlines that can be used to generate H include, but are not limited to, VH3-23 (e.g., UniProt ID: P01764), VH3-74 (e.g., UniProt ID: A0A0B4J1X5), VH3-66 (e.g., UniProt ID: A0A0C4DH42), VH3-30 (e.g., UniProt ID: P01768), VH3-11 (e.g., UniProt ID: P01762), and VH3-9 (e.g., UniProt ID: P01782).

[0113] IL27Rα binding molecule comprising additional agent In some embodiments, the IL27Rα binding molecules of the disclosure comprise an IL27Rα single domain antibody (sdAb) conjugated to one or more additional biologically active agents including, but not limited to, a therapeutic agent, a chemically active agent, an optically active agent, or a radioactive substance, which may be a combination of a therapeutic agent, a chemically active agent, an optically active agent, or a radioactive substance. Conjugation of at least one such biologically active agent, chemically active agent, optically active agent, or radioactive substance imparts additional biological or chemical properties to the IL27Rα binding sdAb, and this combination results in an IL27Rα binding molecule having additional utility or a different utility.

[0114] For example, the additional agent may be a molecule selected from one or more of the following: an immunomodulatory substance (e.g., an immunogen); a molecule that improves water solubility (e.g., a water-soluble polymer and a hydrophilic molecule, such as a sugar); a carrier molecule that extends the in vivo half-life (e.g., PEGylation, Fc fusion, or acylation); for use in a detection assay (e.g., an epitope tag), to enhance ease of purification (e.g., a chelating peptide, such as a polyHis tag), for antibody production; a targeting domain that selectively targets the IL27Rα binding molecule to a specific cell or tissue type; a therapeutic agent (e.g., a therapeutic agent including a small molecule or polypeptide agent); an agent that makes it visible to an optical sensor or an electromagnetic sensor (e.g., a radionuclide or a fluorescent substance). In some embodiments, the linker is a cleavable linker or a non-cleavable linker. As contemplated herein, the use of a cleavable linker in the IL27Rα binding molecule facilitates the release of the therapeutic agent into the intracellular cytoplasm upon internalization of the IL27Rα binding molecule. The use of a non-cleavable linker would allow for release upon digestion of the IL27Rα binding molecule. Alternatively, the non-cleavable linker could be used with an agent that does not require release from the antibody (e.g., an imaging agent).

[0115] In some embodiments, the IL27Rα binding molecule comprises an IL27Rα binding sdAb stably bound to a further agent, linked via a linker. The linker is a covalent bond between two elements of the IL27Rα binding molecule (e.g., the hIL27Rα binding VHH and the PEG polymer). The linker may be a covalent bond, a chemical linker, or a peptide linker. Suitable linkers generally include a "flexible linker" of sufficient length to move somewhat between the IL27Rα binding sdAb and the linked agent. Examples of chemical linkers include arylacetylenes, ethylene glycol oligomers containing 2 - 10 monomer units, diamines, dibasic acids, amino acids, or combinations thereof. In some embodiments, the linker is a peptide linker. Suitable peptide linkers can be readily selected and can be peptide linkers of any suitable length, e.g., a peptide linker of 1 amino acid (e.g., Gly), 2, 3, 4, 5, 6, 7, 8, 9, 10, 10 - 20, 20 - 30, 30 - 50, or more than 50 amino acids. Suitable peptide linkers are known in the art and include, for example, peptide linkers containing flexible amino acid residues such as glycine and serine. Examples of flexible linkers include glycine polymers (G) n , glycine - serine polymers, glycine - alanine polymers, alanine - serine polymers, and other flexible linkers. Glycine and glycine - serine polymers are relatively conformationally disordered and thus may serve as neutral tethers between components. Further examples of flexible linkers include glycine polymers (G) n, including glycine-alanine polymers, alanine-serine polymers, and glycine-serine polymers. Glycine and glycine-serine polymers are relatively structurally indeterminate and can thus serve as neutral tethers between components. To provide a flexible linker that can be used to conjugate heterologous amino acid sequences to the IL27Rα-binding sdAbs disclosed herein, multimers of such linker sequences (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 10-20, 20-30, or 30-50) may be linked together. In some embodiments, the linker has the formula (GGGS)n (SEQ ID NO:197), (GGGSG)n (SEQ ID NO:198), (GGGGS)n (SEQ ID NO:199), (GGS)nG (SEQ ID NO:200), or (GGSG)n (SEQ ID NO:201), where n is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0116] Immunomodulatory agent In some embodiments, the IL27Rα-binding molecules of the disclosure are operably linked to an immunomodulatory agent (immunoconjugate). Immunomodulatory agents that can be conjugated to the hIL27Rα-binding sdAbs of the disclosure include, but are not limited to, inactivated virus particles, inactivated bacterial toxins such as toxoids from diphtheria, tetanus, cholera, or leukotoxin molecules, inactivated bacteria, and dendritic cells. Such immunoconjugates are useful in promoting an immune response against IL27Rα or cells expressing IL27Rα.

[0117] Flag tag In some embodiments, the IL27Rα binding molecules of the disclosure are operably linked to an antigenic tag such as a FLAG sequence. The FLAG sequence is recognized by biotinylated, highly specific anti-FLAG antibodies as described herein (see, e.g., Blanar et al. (1992) Science 256:1014 and LeClair, et al. (1992) PNAS-USA 89:8145). In some embodiments, the IL27Rα binding sdAb polypeptide further comprises a C-terminal c-myc epitope tag.

[0118] Chelating peptide In one aspect, the IL27Rα binding molecule of the present disclosure is operably linked to one or more transition metal chelating polypeptide sequences. Incorporating such a transition metal chelating domain facilitates purification by immobilized metal affinity chromatography (IMAC), as described in U.S. Patent No. 4,569,794 to Smith et al., issued February 11, 1986. Examples of transition metal chelating polypeptides useful in the practice of the present IL27Rα binding molecule are described in Smith et al., supra and in U.S. Patent No. 5,320,663 to Dobeli et al., issued May 10, 1995. The entire disclosures of these are incorporated herein by reference. A particular transition metal chelating polypeptide useful in the practice of the present IL27Rα binding molecule is a polypeptide containing 3 to 6 contiguous histidine residues (SEQ ID NO: 202), for example, the 6 histidine (His)6 peptide (SEQ ID NO: 195), which is often referred to in the art as a "His tag". In general accordance with the disclosures of Anderson et al. (U.S. Patent No. 5,439,829, issued August 8, 1995) and Hale, J.E (1996) Analytical Biochemistry 231(1):46-49, conjugation of the hIL27Rα binding molecule to such a chelating peptide, in addition to providing a purification "handle" for recombinant proteins, or for the purpose of facilitating immobilization on an SPR sensor chip, facilitates the targeted delivery of transition metal ions to IL27Rα-expressing cells as kinetically inert, or kinetically labile complexes. The transition metal ions are reporter molecules, such as fluorescent compounds or radiological imaging agents, and include radioactive substances or therapeutic agents.

[0119] Carrier molecule In some embodiments, the IL27Rα-binding sdAb of the present disclosure may be conjugated to one or more carrier molecules. Carrier molecules are typically large, slowly metabolized macromolecules that confer in vivo stabilization and / or a long duration of action, distinguishing such molecules from conventional carrier molecules used in the preparation of pharmaceutical formulations as described below. Examples of in vivo carriers that can be incorporated into IL27Rα-binding molecules include proteins (including but not limited to human serum albumin); fatty acids (acylation); polysaccharides ((N-linked and O-linked) sugars, sepharose, agarose, cellulose, or cellulose-containing, but not limited to); polypeptide amino acid copolymers; acylation, or polycyallylation, polyethylene glycol (PEG) polymers, but not limited to.

[0120] Water-soluble polymer In some embodiments, the IL27Rα-binding sdAb is conjugated to one or more water-soluble polymers. Examples of water-soluble polymers useful in the practice of this IL27Rα-binding molecule include polyethylene glycol (PEG), polypropylene glycol (PPG), polysaccharides (polyvinylpyrrolidone, copolymers of ethylene glycol and propylene glycol, poly(oxyethylated polyol), polyolefin alcohol, polysaccharide, polyα-hydroxy acid, polyvinyl alcohol (PVA), polyphosphazen, polyoxazoline (POZ), poly(N-acryloylmorpholine), or combinations thereof.

[0121] Polyethylene glycol In one aspect, the carrier molecule is a polyethylene glycol (「PEG」) polymer. Conjugation of a PEG polymer to a protein (PEGylation) is a well-established method for extending the serum half-life of a biological agent. Further, PEGylated polypeptides may be referred to as monoPEGylated, diPEGylated, triPEGylated (etc.) to indicate a polypeptide to which 1, 2, 3 (or more) PEG moieties are each attached to the polypeptide. In some aspects, PEG may be attached directly, covalently, to the sdAb (e.g., via a lysine side chain, a cysteine sulfhydryl group, or an N-terminal amine), and optionally, a linker is used between the PEG and the sdAb. In some aspects, the IL27Rα binding molecule comprises multiple PEG molecules, each of the PEG molecules being attached to a different amino acid residue. In some aspects, the sdAb may be modified by incorporating unnatural amino acids having unnatural amino acid side chains to facilitate site-specific PEGylation. In other aspects, one or more cysteine residues at one or more positions in the sdAb may be substituted to facilitate site-specific PEGylation via a cysteine sulfhydryl side chain.

[0122] Optionally, the IL27Rα binding molecule of the present disclosure has an N-terminal glutamine (「1Q」) residue. The N-terminal glutamine residue has been observed to cyclize spontaneously to form pyroglutamate (pE) under or near physiological conditions (see, e.g., Liu, et al (2011) J. Biol. Chem. 286(13): 11211-11217). In some aspects, when pyroglutamate forms, N-terminal PEG conjugation is complicated, particularly when aldehyde chemistry is used for N-terminal PEGylation. As a result, when PEGylating the IL27Rα binding molecule of the present disclosure, particularly when aldehyde chemistry is used, position 1 of the IL27Rα binding molecule having an amino acid (e.g., 1Q) at position 1 is substituted with an alternative amino acid or position 1 is deleted (e.g., des-1Q). In some aspects, the IL27Rα binding molecule of the present disclosure comprises an amino acid substitution selected from the group of Q1E and Q1D.

[0123] PEGs suitable for conjugation to polypeptide sequences are generally soluble in water at room temperature and have the general formula R(O-CH2-CH2) n O-R wherein R is hydrogen or a protecting group, such as an alkyl group or an alkanol group, and n is an integer from 1 to 1000. When R is a protecting group, it generally has from 1 to 8 carbons. The PEG can be linear or branched. Branched PEG derivatives, "star PEGs", and multi-armed PEGs are contemplated by the present disclosure.

[0124] The molecular weight of the PEG used in the IL27Rα binding molecule is not limited to any particular range. The molecular weight of the PEG component of the IL27Rα binding molecule may be greater than about 5 kDa, greater than about 10 kDa, greater than about 15 kDa, greater than about 20 kDa, greater than about 30 kDa, greater than about 40 kDa, or greater than about 50 kDa. In some embodiments, the molecular weight is from about 5 kDa to about 10 kDa, from about 5 kDa to about 15 kDa, from about 5 kDa to about 20 kDa, from about 10 kDa to about 15 kDa, from about 10 kDa to about 20 kDa, from about 10 kDa to about 25 kDa, or from about 10 kDa to about 30 kDa. A linear or branched PEG molecule having a molecular weight of from about 2,000 to about 80,000 daltons, or from about 2,000 to about 70,000 daltons, or from about 5,000 to about 50,000 daltons, or from about 10,000 to about 50,000 daltons, or from about 20,000 to about 50,000 daltons, or from about 30,000 to about 50,000 daltons, or from about 20,000 to about 40,000 daltons, or from about 30,000 to about 40,000 daltons. In one embodiment of the IL27Rα binding molecule, the PEG is a 40kD branched PEG containing two 20kD arms.

[0125] The present disclosure also contemplates IL27Rα binding molecules that include multiple PEG moieties, where the PEGs have different size values and thus various different PEGs are present in specific ratios. For example, in the preparation of PEGylated IL27Rα binding molecules, some compositions include a mixture of mono-PEGylated, di-PEGylated, tri-PEGylated, and quadra-PEGylated sdAb conjugates. In some compositions, the percentage of mono-PEGylated species is 18-25%, the percentage of di-PEGylated species is 50-66%, the percentage of tri-PEGylated species is 12-16%, and the percentage of quadra-PEGylated species is up to 5%. Such composite compositions can be generated by reaction conditions and purification methods known in the art. Chromatography may be used to separate conjugate fractions, and then, for example, fractions containing conjugates with the desired number of PEGs attached are identified and purified from unmodified protein sequences and conjugates with other numbers of PEGs attached.

[0126] PEGylation is most frequently performed at the α-amino group at the N-terminus of the polypeptide, the ε-amino group in the side chain of lysine residues, and the imidazole group in the side chain of histidine residues. Since most recombinant polypeptides have one α-amino group and multiple ε-amino groups and imidazole groups, a very large number of positional isomers can be generated depending on the linker chemistry.

[0127] Two widely used first-generation activated monomethoxy PEG (mPEG) are succinimidyl carbonate PEG (SC-PEG; see, e.g., Zalipsky, et al. (1992) Biotehnol. Appl. Biochem 15:100-114) and benzotriazole carbonate PEG (BTC-PEG; see, e.g., Dolence et al., U.S. Patent No. 5,650,234), which preferentially react with lysine residues to form carbamate bonds, but are also known to react with histidine and tyrosine residues. The use of PEG-aldehyde linkers targets one site at the N-terminus of the polypeptide via reductive amination.

[0128] PEG can be attached to the IL27Rα binding molecules of the present disclosure via a terminal reactive group ("spacer") that mediates the bond between one or more free amino or carboxyl groups of the polypeptide sequence and polyethylene glycol. PEGs having a spacer capable of attaching to a free amino group include N-hydroxysuccinimide polyethylene glycol, which can be prepared by activating the succinate ester of polyethylene glycol with N-hydroxysuccinimide.

[0129] In some embodiments, PEGylation of the sdAb is facilitated by incorporating unnatural amino acids having unique side chains to promote site-specific PEGylation. To achieve site-specific PEGylation of such polypeptides, it is known in the art to incorporate unnatural amino acids into the polypeptide to provide a functional moiety. See, e.g., Ptacin, et al., PCT International Application No. PCT / US2018 / 045257, filed August 3, 2018 and published February 7, 2019 as International Publication No. WO2019 / 028419Al.

[0130] The PEG moiety of the PEGylated IL27Rα binding molecule may be linear or branched. Branched PEG derivatives, "star PEGs", and multi-armed PEGs are contemplated by the present disclosure.Certain embodiments of PEG useful in the practice of the present disclosure include 10 kDa linear PEG-aldehyde (e.g., Sunbright® ME-100AL, NOF America Corporation, One North Broadway, White Plains, NY 10601 USA), 10 kDa linear PEG-NHS ester (e.g., Sunbright® ME-100CS, Sunbright® ME-100AS, Sunbright® ME-100GS, Sunbright® ME-100HS, NOF), 20 kDa linear PEG-aldehyde (e.g., Sunbright® ME-200AL, NOF, 20 kDa linear PEG-NHS ester (e.g., Sunbright® ME-200CS, Sunbright® ME-200AS, Sunbright® ME-200GS, Sunbright® ME-200HS, NOF), 20 kDa 2-arm branched PEG-aldehyde, 20 kDa PEG-aldehyde containing two 10 kDa linear PEG molecules (e.g., Sunbright® GL2-200AL3, NOF), 20 kDa 2-arm branched PEG-NHS ester, 20 kDa PEG-NHS ester containing two 10 kDa linear PEG molecules (e.g., Sunbright® GL2-200TS, Sunbright® GL200GS2, NOF), 40 kDa 2-arm branched PEG-aldehyde, 40 kDa PEG-aldehyde containing two 20 kDa linear PEG molecules (e.g., Sunbright® GL2-400AL3), 40 kDa 2-arm branched PEG-NHS ester, 40 kDa PEG-NHS ester containing two 20 kDa linear PEG molecules (e.g., Sunbright® GL2-400AL3, Sunbright® GL2-400GS2, NOF), linear 30 kDa PEG-aldehyde (e.g., Sunbright® ME-300AL), and linear 30 kDa PEG-NHS ester are included.

[0131] Fc fusion In some embodiments, the carrier molecule is an Fc molecule or a monomeric subunit thereof. In some embodiments, the dimeric Fc molecule may be engineered to have a "knob-into-hole modification". The knob-into-hole modification is further described in Ridgway, et al. (1996) Protein Engineering 9(7):617-621, as well as U.S. Patent No. 5,731,168, issued March 24, 1998, U.S. Patent No. 7,642,228, issued January 5, 2010, U.S. Patent No. 7,695,936, issued April 13, 2010, or U.S. Patent No. 8,216,805, issued July 10, 2012. The knob-into-hole modification refers to a modification at the interface between two immunoglobulin heavy chains in the CH3 domain, where i) an amino acid residue in the CH3 domain of the first heavy chain is replaced with an amino acid residue having a larger side chain (e.g., tyrosine or tryptophan), creating a protrusion ("knob") from the surface, and ii) an amino acid residue in the CH3 domain of the second heavy chain is replaced with an amino acid residue having a smaller side chain (e.g., alanine or threonine), thereby creating a cavity ("hole") within the interface of the second CH3 domain, and the protruding side chain ("knob") of the first CH3 domain is accommodated within the cavity in the second CH3 domain. In one embodiment, the "knob-into-hole modification" includes the amino acid substitution T366W, and optionally, the amino acid substitution S354C is in one antibody heavy chain, and the amino acid substitutions T366S, L368A, Y407V, and optionally Y349C are in the other antibody heavy chain. Further, the Fc domain may be modified by introducing a cysteine residue at position S354 of one chain and Y349 of the other chain, resulting in a stabilizing disulfide bond between the two antibody heavy chains in the Fc region (Carter, et al. (2001) Immunol Methods 248, 7-15).The knob-into-hole format is used to facilitate the expression of a first polypeptide (e.g., an IL27Rα-binding sdAb) on a first Fc monomer having a "knob" modification and a second polypeptide on a second Fc monomer having a "hole" modification to facilitate the expression of a heterodimeric polypeptide conjugate.

[0132] Targeting domain In some embodiments, the IL27Rα-binding molecule is provided as a component of a multivalent (e.g., bivalent) fusion protein having such a targeting domain, optionally with a linker incorporated between the IL27Rα-binding sdAb sequence and the sequence of the targeting domain of the fusion protein, to facilitate selective binding to a particular cell type or tissue that expresses a cell surface molecule that specifically binds to a polypeptide sequence (the "targeting domain").

[0133] In some embodiments of the IL27Rα-binding molecule, the IL27Rα-binding molecule can be targeted to a particular cell type cell by incorporating the targeting domain into the structure of the IL27Rα-binding molecule. As used herein, the term targeting domain refers to a portion that specifically binds to a molecule expressed on the target cell surface. The targeting domain can be any portion that specifically binds to one or more cell surface molecules (e.g., T cell receptor) expressed on the target cell surface. In some embodiments, the target cell is a T cell. In some embodiments, the target cell is an IL27Rα+ T cell.

[0134] In some embodiments, the targeting domain is a ligand for a receptor. In some embodiments, the targeting domain is a ligand for a receptor expressed on the surface of a T cell. In some embodiments, the ligand is a cytokine. In some embodiments, the cytokine includes, but is not limited to, the group consisting of interleukins, interferons, and functional derivatives thereof. In some embodiments, the cytokine includes, but is not limited to, the group consisting of IL2, IL3, IL4, IL7, IL9, IL12, IL15, IL18, IL21, IL22, IL23, IL27, IL28, IL34, and modified versions or fragments thereof that bind to cognate ligands expressed on the surface of T cells. In some embodiments, the cytokine includes, but is not limited to, the group consisting of interferon α, interferon a2b, interferon γ, or interferon λ, and modified versions or fragments thereof that bind to cognate ligands expressed on the surface of T cells.

[0135] In another aspect, the present disclosure provides a multivalent binding molecule comprising (a) an IL27Rα binding molecule and (b) a second binding molecule that specifically binds to the extracellular domain of a second cell surface molecule, wherein the IL27Rα binding molecule and the second binding molecule are functionally linked, optionally functionally linked via a chemical linker or a polypeptide linker. In some embodiments, the IL27Rα binding molecule of the present disclosure is useful in the preparation of the multivalent binding molecules described in Gonzalez, et al. PCT / US2018 / 021301, published as WO2018 / 182935A1 on October 4, 2018. In some aspects, the second binding molecule specifically binds to (i) a component of a cytokine receptor that activates the JAK / STAT pathway of a cell; (ii) a receptor tyrosine kinase; or (iii) the extracellular domain of a TNFR superfamily member. In some embodiments, the second surface molecule is a tyrosine kinase selected from EGFR, ErbB2, ErbB3, ErbB4, InsR, IGF1R, InsRR, PDGFRα, PDGFRβ, CSF1R / Fms, cKit, Flt-3 / Flk2, VEGFR1, VEGFR2, VEGFR3, FGFR1, FGFR2, FGFR3, FGFR4, PTK7 / CCK4, TrkA, TrkB, TrkC, Ror1, Ror2, MuSK, Met, Ron, Axl, Mer, Tyro3, Tie1, Tie2, EphA1-8, EphA10, EphB1-4, EphB6, Ret, Ryk, DDR1, DDR2, Ros, LMR1, LMR2, LMR3, ALK, LTK, SuRTK106 / STYK1.In some embodiments, the second surface molecule is TNFR1 (TNFRSF1A), TNFR2 (TNFRSF1B; TNFRSF2), 41-BB (TNFRSF9); AITR (TNFRSF18); BCMA (TNFRSF17), CD27 (TNFRSF7), CD30 (TNFRSF8), CD40 (TNFRSF5), decoy receptor 1 (TNFRSF10C), decoy receptor-3 (TNFRSF25), decoy receptor 4 (TNFRSF10A), decoy receptor 5 (TNFRSF10B), decoy receptor-6 (TNFRSF21), decoy receptor-3 (TNFRSF6B), decoy receptor 2 (TNFRSF10D), EDAR, Fas (TNFRSF6), HVEM (TNFRSF14),. LTBR (TNFRSF3), OX40 (TNFRSF4), RANK (TNFRSF11A), TACI (TNFRSF13B), Troy (TNFRSF19), XEDAR (TNFRSF27), osteoprotegerin (TNFRSF11B), TWEAK receptor (TNFRSF12A), BAFF receptor (TNFRSF13C), NGF receptor (TNFRSF16), which is a TNFR superfamily member selected from the group consisting of.

[0136] In some embodiments, the targeting domain is a polypeptide that specifically binds to a cell surface molecule associated with tumor cells (e.g., the cognate ligand of a tumor cell receptor) selected from the group consisting of GD2, BCMA, CD19, CD33, CD38, CD70, GD2, IL3Ra2, CD19, mesothelin, Her2, EpCam, Muc1, ROR1, CD133, CEA, EGRFRVIII, PSCA, GPC3, Pan-ErbB, and FAP.

[0137] In one aspect, the targeting domain of the IL27Rα binding molecule is an antibody (molecule, including molecules such as VHH, scFv, etc. as defined above). Examples of antibodies that can be incorporated as the targeting domain of the IL27Rα binding molecule include anti-GD2 antibody, anti-BCMA antibody, anti-CD19 antibody, anti-CD33 antibody, anti-CD38 antibody, anti-CD70 antibody, anti-GD2 antibody and IL3Ra2 antibody, anti-CD19 antibody, anti-mesothelin antibody, anti-Her2 antibody, anti-EpCam antibody, anti-Muc1 antibody, anti-ROR1 antibody, anti-CD133 antibody, anti-CEA antibody, anti-PSMA antibody, anti-EGRFRVIII antibody, anti-PSCA antibody, anti-GPC3 antibody, anti-Pan-ErbB antibody, and anti-FAP antibody, but are not limited thereto.

[0138] The antibody or its antigen-binding fragment can be linked to another antibody to form, for example, a bispecific antibody or a multispecific antibody.

[0139] Label In some aspects, the IL27Rα binding molecules of the present disclosure are operably linked to a label. In some aspects, the label is incorporated to facilitate use as an imaging agent, a diagnostic agent, or for use in cell sorting procedures. The term label includes, but is not limited to, fluorescent labels, biologically active enzyme labels, radioisotopes (e.g., radioactive ions), nuclear magnetic resonance active labels, luminescence labels, or magnetic compounds. In one aspect, an IL27Rα binding sdAb (e.g., an IL27Rα binding VHH) molecule that is stably associated (e.g., covalently, coordinately covalently) with an imaging label. The term imaging label is used to describe any of a variety of compounds that are signatures that facilitate the identification, tracking, and / or location measurement of an IL27Rα binding sdAb (or its metabolites) using diagnostic procedures. Examples of imaging labels include, but are not limited to, fluorescent compounds, radioactive compounds, and compounds that do not pass through imaging methods (e.g., X-rays, ultrasound). Examples of radioactive compounds useful as imaging labels include technetium-99m ( 99m Tc), indium-111 ( 111 In), iodine-131 ( 131I), Iodine-123( 123 I), Iodine-125( 125 I), Gallium-67( 67 Ga), and Lutetium-177( 177 Lu), Phosphorus( 32 P), Carbon( 14 C), Tritium( 3 H), Yttrium( 90 Y), Actinium( 225 Ac), Astatine( 211 At), Rhenium( 186 Re), Bismuth( 212 Bi or 213 Bi), and Rhodium( 188 Rh) are included, but not limited to this.

[0140] Therapeutic agent In some embodiments, the IL27Rα binding molecule of the present disclosure is functionally linked to a therapeutic agent. Examples of therapeutic agents include antibodies, cytotoxic or cytostatic compounds, radioisotopes, plant-derived, fungal-derived, or bacterial-derived molecules, or biological proteins (e.g., protein toxins) or particles (e.g., nanoparticles or recombinant virus particles, e.g., recombinant virus particles via viral coat proteins), therapeutic antibodies, therapeutic small molecules (e.g., chemotherapeutic agents) or biotherapeutic agents, as more fully described herein.

[0141] In some embodiments, the therapeutic agent that can be incorporated into the IL27Rα binding molecule of the present disclosure is, for example, a short-range radiation emitter including a short-range, high-energy a-emitter. Examples of such radioisotopes include α-emitters, β-emitters, γ-emitters, or β / γ-emitters. Radioisotopes useful as therapeutic agents include Yttrium 90( 90 Y), Lutetium-177( 177 Lu), Actinium-225( 225 Ac), Astatine-211( 211 At), Rhenium-186( 186 Re), Bismuth-212( 212 Bi), Bismuth-213( 213Bi), and rhodium-188( 188 Rh) is included.

[0142] In some embodiments, the IL27Rα binding molecule of the present disclosure is functionally linked to a cytotoxic agent (or a derivative thereof), such as maytansinol or a DM1 maytansinoid), a taxane, or calicheamicin, Pseudomonas exotoxin A, debuganin, ricin toxin, diphtheria toxin, amatoxin, e.g., a-amanitin, saporin, maytansine, maytansinoid, auristatin, anthracycline, calicheamicin, irinotecan, SN-38, duocarmycin, pyrrolobenzodiazepine, pyrrolobenzodiazepine dimer, indolinobenzodiazepine, and indolinobenzodiazepine dimer, or a variant thereof).

[0143] Synthesis of the IL27Rα binding molecule: In some embodiments, the IL27Rα binding molecule of the present disclosure is a polypeptide. However, in some embodiments, only a portion of the IL27Rα binding molecule is a polypeptide, e.g., the IL27Rα binding molecule includes a non-peptidyl domain (e.g., a PEG IL27Rα binding sdAb conjugate, a radionucleotide IL27Rα binding sdAb conjugate, or a small molecule IL27Rα binding sdAb conjugate). The following provides guidance for enabling solid-phase synthesis and recombinant synthesis of the polypeptide portion (domain) of the IL27Rα binding molecule of the present disclosure. In embodiments where only a portion of the IL27Rα binding molecule is a polypeptide, the peptidyl domain of the IL27Rα binding molecule will be understood to be an intermediate in the process that may be subject to further processing to complete the synthesis of the desired IL27Rα binding molecule. The polypeptide domain of the IL27Rα binding molecule may be produced by conventional methodologies for polypeptide construction, including recombinant synthesis or solid-phase synthesis, as described in more detail below.

[0144] Chemical synthesis In addition to generating mutant polypeptides via the expression of nucleic acid molecules altered by recombinant molecular biological techniques, the polypeptide domains of the IL27Rα binding molecules can be chemically synthesized. Chemically synthesized polypeptides are routinely produced by those skilled in the art. Chemical synthesis includes direct peptide synthesis by chemical means of the polypeptide domains of the IL27Rα binding molecules that exhibit the described characteristics. This method can incorporate natural and unnatural amino acids at desired positions that facilitate the ligation of specific molecules (e.g., PEG).

[0145] In some embodiments, the polypeptide domains of the IL27Rα binding molecules of the present disclosure can be produced by chemical synthesis. Chemical synthesis of the polypeptide domains of the IL27Rα binding molecules may proceed via a liquid phase or via a solid phase. Use of solid-phase peptide synthesis (SPPS) allows for the incorporation of unnatural amino acids and / or backbone modifications of peptides / proteins. A variety of types of SPPS can be utilized to synthesize the polypeptide domains of the IL27Rα binding molecules of the present disclosure and are known in the art (e.g., Ganesan A. (2006) Mini Rev. Med. Chem. 6:3-10; and Camarero J.A. et al., (2005) Protein Pept Lett. 12:723-8). Alpha-functional groups and any reactive side chains may be protected during chemical synthesis by acid-labile or base-labile groups that are stable under the conditions for ligating amide bonds but can be readily cleaved without damaging the formed peptide chain.

[0146] In solid-phase synthesis, either the N-terminal amino acid or the C-terminal amino acid can be attached to a suitable support material. A suitable support material is inert to the reagents and reaction conditions for the stepwise condensation and cleavage reactions of the synthesis process and does not dissolve in the reaction medium being used. Examples of commercially available support materials include styrene / divinylbenzene copolymers modified with reactive groups and / or polyethylene glycol; chloromethylated styrene / divinylbenzene copolymers; hydroxymethylated or aminomethylated styrene / divinylbenzene copolymers, and the like. The sequential coupling of protected amino acids can be carried out according to conventional methods in peptide synthesis, typically in an automated peptide synthesizer.

[0147] At the end of solid-phase synthesis, the peptide is cleaved from the support material while the side-chain protecting groups are simultaneously cleaved. The resulting peptide can be purified by various chromatography methods including, but not limited to, hydrophobic adsorption chromatography, ion-exchange chromatography, distribution chromatography, high-pressure liquid chromatography (HPLC), and reverse-phase HPLC.

[0148] Recombinant production Alternatively, the polypeptide domain of the IL27Rα binding molecule of the present disclosure may be produced by recombinant DNA technology. In a typical practice of recombinant production of a polypeptide, a nucleic acid sequence encoding the desired polypeptide is incorporated into an expression vector suitable for the host cell in which expression is to occur, and this nucleic acid sequence is operably linked to one or more expression control sequences encoded by the vector and functions in the target host cell. The recombinant protein may be recovered by disrupting the host cell or, if a secretion leader sequence (signal peptide) is incorporated into the polypeptide, from the cell culture medium. The recombinant protein may be purified and concentrated for further use including incorporation.

[0149] Synthesis of the nucleic acid sequence encoding the IL27Rα binding molecule In some embodiments, the polypeptide domain of the IL27Rα binding molecule is produced by a recombinant method using a nucleic acid sequence encoding the polypeptide domain of the IL27Rα binding molecule (or a fusion protein comprising the polypeptide domain of the IL27Rα binding molecule). The nucleic acid sequence encoding the desired polypeptide domain of the IL27Rα binding molecule can be synthesized by chemical means using an oligonucleotide synthesizer.

[0150] The nucleic acid molecule is not limited to a sequence encoding a polypeptide. It may also include some or all of the non-coding sequences upstream or downstream from the coding sequence (e.g., the coding sequence of the polypeptide domain of the IL27Rα binding molecule). Those skilled in the art of molecular biology are familiar with routine procedures for isolating nucleic acid molecules. For example, a nucleic acid molecule can be prepared by treating genomic DNA with a restriction endonuclease or by performing polymerase chain reaction (PCR). When the nucleic acid molecule is ribonucleic acid (RNA), the molecule can be produced, for example, by in vitro transcription.

[0151] The nucleic acid molecule encoding the polypeptide domain of the IL27Rα binding molecule (and its fusions) may contain a native sequence and, although different from that which occurs in nature, may contain a sequence encoding the same polypeptide due to genetic code degeneracy. These nucleic acid molecules may consist of RNA or DNA (e.g., genomic DNA, cDNA, or synthetic DNA, such as that generated by phosphoramidite-based synthesis), or combinations or modifications of nucleotides within these types of nucleic acids. Furthermore, the nucleic acid molecule may be double-stranded or single-stranded (i.e., either the sense strand or the antisense strand).

[0152] Nucleic acid sequences encoding the polypeptide domains of the IL27Rα binding molecule may be obtained from various commercial suppliers that provide custom synthesis of nucleic acid sequences. Amino acid sequence variants of the human IL27Rα binding molecule of the present disclosure are prepared by introducing appropriate nucleotide changes into the coding sequence based on the genetic code well known in the art. Such variants are insertions, substitutions, and / or specified deletions of residues as mentioned. Any combination of insertions, substitutions, and / or specified deletions can be added to reach the final construct, provided that the final construct has the desired biological activity as defined herein.

[0153] Methods for constructing DNA sequences encoding the polypeptide domains of the IL27Rα binding molecule, and methods for expressing these sequences in a suitably transformed host, include, but are not limited to, the use of mutagenesis methods assisted by PCR. Mutations consisting of deletions or additions of amino acid residues to the polypeptide domain of the IL27Rα binding molecule can also be added using standard recombinant methods. In the case of deletions or additions, optionally, a nucleic acid molecule encoding the polypeptide domain of the IL27Rα binding molecule that is digested by an appropriate restriction endonuclease. The resulting fragment may be expressed directly or may be further manipulated, for example, by ligation to a second fragment. Ligation may be facilitated if the two ends of the nucleic acid molecule contain complementary nucleotides that overlap with each other, but blunt-ended fragments can also be ligated. Nucleic acids produced by PCR can also be used to generate various mutant sequences.

[0154] The polypeptide domains of the IL27Rα binding molecules of the present disclosure can be produced directly by recombination, or can be produced as fusion polypeptides together with heterologous polypeptides, such as signal sequences, or other polypeptides having specific cleavage sites at the N-terminus or C-terminus of the mature IL27Rα binding molecule. Generally, the signal sequence can be a component of the vector or part of the coding sequence inserted into the vector. The heterologous signal sequence selected is preferably one that is recognized and processed by the host cell (i.e., cleaved by signal peptidase). In some embodiments, the signal sequence is a signal sequence that is naturally associated with the IL27Rα binding molecule (i.e., the human IL27Rα signal sequence). Incorporation of the signal sequence depends on whether it is desirable to secrete the IL27Rα binding molecule from the recombinant cell in which the IL27Rα binding molecule is made. If the selected cell is prokaryotic, generally, it is preferred that the DNA sequence does not encode a signal sequence. If the selected cell is eukaryotic, generally, a signal sequence is encoded, and most preferably, a wild-type IL-2 signal sequence is preferably used. Alternatively, signal sequences derived from secreted polypeptides of the same or related species, as well as heterologous mammalian signal sequences such as viral secretion leaders, e.g., herpes simplex gD signal, may be suitable. When the recombinant host cell is a yeast cell such as Saccharomyces cerevisiae, an α mating factor secretion signal sequence may be used to secrete the IL27Rα binding molecule extracellularly into the culture medium, as described in Singh, U.S. Patent No. 7,198,919 B1.

[0155] When the polypeptide domain of the IL27Rα binding molecule to be expressed is expressed as a chimera (e.g., a fusion protein comprising an IL27Rα binding molecule and a heterologous polypeptide sequence), the chimeric protein may be encoded by a hybrid nucleic acid molecule comprising a first sequence encoding all or part of the polypeptide domain of the IL27Rα binding molecule and a second sequence encoding all or part of the heterologous polypeptide. For example, the polypeptide domain of the IL27Rα binding molecule described herein may be fused with a hexahistidine tag (SEQ ID NO:195) to facilitate purification of the protein expressed by bacteria, or may be fused with hexahistidine (SEQ ID NO:195), hemagglutinin, or an Fc tag to facilitate purification of the protein expressed in eukaryotic cells. The first and second should not be understood as limitations on the orientation of the elements of the fusion protein, and the heterologous polypeptide can be linked to either the N-terminus and / or the C-terminus of the polypeptide domain of the IL27Rα binding molecule. For example, the N-terminus may be linked to a targeting domain, and the C-terminus may be linked to a hexahistidine tag (SEQ ID NO:195) purification handle.

[0156] Using the complete amino acid sequence of the polypeptide domain of the IL27Rα binding molecule (or fusion / chimera) to be expressed, a gene can be constructed by reverse translation. A DNA oligomer containing a nucleotide sequence encoding the polypeptide domain of the IL27Rα binding molecule can be synthesized. For example, several small oligonucleotides encoding a portion of the desired polypeptide can be synthesized and then ligated. The individual oligonucleotides typically contain 5' overhangs or 3' overhangs for complementary assembly.

[0157] In some embodiments, the nucleic acid sequence encoding the polypeptide domain of the IL27Rα binding molecule may be "codon optimized" to facilitate expression in a particular host cell type. Techniques for codon optimization in a variety of expression systems, including mammalian host cells, yeast host cells, and bacterial host cells, are well known in the art, and there are online tools for providing codon-optimized sequences for expression in various host cell types. See, for example, Hawash, et al., (2017) 9:46-53, and Mauro and Chappell in Recombinant Protein Expression in Mammalian Cells: Methods and Protocols , edited by David Hacker (Human Press New York). In addition, there are various web-based online software packages freely available to assist in the preparation of codon-optimized nucleic acid sequences.

[0158] Expression vector Once assembled (by synthesis, site-directed mutagenesis, or another method), the nucleic acid sequence encoding the polypeptide domain of the IL27Rα binding molecule is inserted into an expression vector. A variety of expression vectors are available for use in a variety of host cells and are typically based on the host cell to be expressed. Expression vectors typically include, but are not limited to, one or more of the following: an origin of replication, one or more marker genes, enhancer elements, a promoter, and a transcription termination sequence. Vectors include viral vectors, plasmid vectors, integrative vectors, and the like. A plasmid is an example of a non-viral vector. To facilitate efficient expression of the recombinant polypeptide, the nucleic acid sequence encoding the polypeptide sequence to be expressed is operably linked to transcriptional and translational regulatory control sequences that function in the selected expression host.

[0159] Expression vectors typically contain a selectable gene, also called a selectable marker. This gene encodes a protein necessary for the survival or growth of transformed host cells grown in a selective culture medium. Host cells not transformed with a vector containing the selectable gene do not survive in the culture medium. Representative selectable genes encode (a) proteins that confer resistance to antibiotics or other toxins, such as ampicillin, neomycin, methotrexate, or tetracycline, (b) proteins that complement auxotrophic deficiencies, or (c) proteins that supply essential nutrients not available from complex media.

[0160] An expression vector for the polypeptide domain of the IL27Rα binding molecule of the present disclosure contains regulatory sequences recognized by the host organism and operably linked to a nucleic acid sequence encoding the polypeptide domain of the IL27Rα binding molecule. The terms "regulatory control sequence", "regulatory sequence", or "expression control sequence" are used interchangeably herein to refer to promoters, enhancers, and other expression control elements (e.g., polyadenylation signals). See, for example, Goeddel (1990) in Gene Expression Technology: Methods in Enzymology 185 (Academic Press, San Diego CA USA). Regulatory sequences include regulatory sequences that induce constitutive expression of nucleotide sequences in many types of host cells, and regulatory sequences that induce expression of nucleotide sequences only in certain host cells (e.g., tissue-specific regulatory sequences). It will be understood by those skilled in the art that the design of an expression vector may be influenced by factors such as the choice of host cell to be transformed and the desired level of expression of the protein. Various factors understood by those skilled in the art must be considered in the selection of expression control sequences. These include, for example, the relative strength of the sequence, its controllability, and in particular, its compatibility with the actual DNA sequence encoding the present IL27Rα binding molecule with respect to potential secondary structure.

[0161] In one aspect, the regulatory sequence is a promoter, and the promoter is selected, for example, based on the cell type in which expression is desired. A promoter is an untranslated sequence that is located upstream (5') of the start codon of a structural gene (generally within about 100 - 1000 bp) and controls the transcription and translation of a functionally linked specific nucleic acid sequence. Such promoters are typically divided into two classes, inducible promoters and constitutive promoters. An inducible promoter is a promoter that initiates high-level transcription from DNA under its control in response to some change in culture conditions, such as the presence or absence of nutrients or a change in temperature. A number of promoters recognized by various potential host cells are well known.

[0162] The T7 promoter can be used in bacteria, the polyhedrin promoter can be used in insect cells, and the cytomegalovirus or metallothionein promoter can be used in mammalian cells. Similarly, in the case of higher eukaryotes, tissue-specific promoters and cell-type-specific promoters can be widely utilized. These promoters are so named because of their ability to induce the expression of nucleic acid molecules in a particular tissue or cell type in the body. Those skilled in the art are well aware of a very large number of promoters and other regulatory elements that can be used to induce nucleic acid expression.

[0163] Transcription from a vector in a mammalian host cell may be controlled by such promoters as are obtained from the genomes of viruses such as polyoma virus, fowlpox virus, adenovirus (e.g., human adenovirus serotype 5), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, retroviruses (e.g., murine stem cell virus), hepatitis B virus, most preferably the promoter obtained from the genome of simian virus 40 (SV40), heterologous mammalian promoters such as the actin promoter, PGK (phosphoglycerate kinase), or immunoglobulin promoter, or the promoter obtained from a heat shock promoter, provided such promoter is compatible with the host cell system. Conveniently, the early and late promoters of the SV40 virus are obtained as an SV40 restriction fragment which also contains the SV40 origin of replication.

[0164] Transcription by higher eukaryotes is often increased by inserting enhancer sequences into vectors. Enhancers are typically cis-acting DNA elements of about 10 - 300 bp that act on promoters to increase transcription. Enhancers are relatively independent of direction and position and have been found to be 5' and 3' to the transcription unit, within introns, and even within the coding sequences themselves. Now, many enhancer sequences derived from mammalian genes (globin, elastase, albumin, α-fetoprotein, and insulin) are known. However, enhancers derived from eukaryotic viruses are usually used. Examples include the SV40 enhancer located downstream of the origin of replication, the cytomegalovirus immediate-early promoter enhancer, the polyoma enhancer located downstream of the origin of replication, and the adenovirus enhancer. The enhancer may be spliced and inserted 5' or 3' to the position of the coding sequence of the expression vector, preferably located 5' to the site from the promoter. Expression vectors used in eukaryotic host cells also contain sequences necessary for transcription termination and mRNA stabilization. Such sequences can generally be obtained from the 5' untranslated regions, and in some cases the 3' untranslated regions, of eukaryotic or viral DNA or cDNA. Standard techniques are used in the construction of appropriate vectors containing one or more of the components listed above.

[0165] In addition to the sequences that facilitate transcription of the inserted nucleic acid molecule, the vector may contain an origin of replication and other genes encoding selectable markers. For example, the neomycin resistance (neoR) gene confers G418 resistance to cells in which the neomycin resistance (neoR) gene is expressed, thus enabling phenotypic selection of transfected cells. Further examples of marker or reporter genes include β-lactamase, chloramphenicol acetyltransferase (CAT), adenosine deaminase (ADA), dihydrofolate reductase (DHFR), hygromycin-B-phosphotransferase (HPH), thymidine kinase (TK), lacZ (encoding β-galactosidase), and xanthine-guanine phosphoribosyltransferase (XGPRT). One of ordinary skill in the art can readily determine whether a particular regulatory element or selectable marker is suitable for use in a particular experimental situation. The correct assembly of the expression vector can be confirmed by nucleotide sequencing, restriction enzyme mapping, and the expression of a biologically active polypeptide in an appropriate host.

[0166] Host cell Furthermore, the present disclosure provides a prokaryotic or eukaryotic cell that contains and expresses a nucleic acid molecule encoding a polypeptide domain of an IL27Rα binding molecule. The cells of the present disclosure are transfected cells, i.e., cells into which a nucleic acid molecule, e.g., a nucleic acid molecule encoding a polypeptide domain of an IL27Rα binding molecule, has been introduced by recombinant DNA methods. The progeny of such cells are also considered to be within the scope of the present disclosure.

[0167] Host cells are typically selected according to their compatibility with the selected expression vector, the toxicity of the product encoded by the DNA sequence of this IL27Rα binding molecule, secretion characteristics, the ability to correctly fold the polypeptide, the requirements for fermentation or culture, and the ease of purification of the product encoded by the DNA sequence. Suitable host cells for cloning or expressing DNA in vectors herein are prokaryotes, yeasts, or higher eukaryotic cells.

[0168] In some embodiments, the recombinant polypeptide domain of the IL27Rα binding molecule or a biologically active variant thereof can also be produced in eukaryotes such as yeast or human cells. Suitable eukaryotic host cells include insect cells (examples of baculovirus vectors available for protein expression in cultured insect cells (e.g., Sf9 cells) include the pAc series (Smith et al. (1983) Mol. Cell Biol. 3:2156-2165) and the pVL series (Lucklow and Summers (1989) Virology 170:31-39)); yeast cells (examples of vectors for expression in yeast S. cerevisiae include pYepSecl (Baldari et al. (1987) EMBO J. 6:229-234), pMFa (Kurjan and Herskowitz (1982) Cell 30:933-943), pJRY88 (Schultz et al. (1987) Gene 54:113-123), pYES2 (Invitrogen Corporation, San Diego, Calif.), and pPicZ (Invitrogen Corporation, San Diego, Calif.)); or mammalian cells (mammalian expression vectors include pCDM8 (Seed (1987) Nature 329:840) and pMT2PC (Kaufman et al. (1987) EMBO J. 6:187:195)).

[0169] Examples of useful mammalian host cell lines are mouse L cells (L-M [TK-], ATCC#CRL-2648), monkey kidney CV1 line transformed by SV40 (COS-7, ATCC CRL 1651); human fetal kidney cells (HEK293 cells or HEK293 cells subcloned for growth in suspension culture; baby hamster kidney cells (BHK, ATCC CCL 10); Chinese hamster ovary cells / -DHFR (CHO); mouse Sertoli cells (TM4); monkey kidney cells (CV1 ATCC CCL 70); African green monkey kidney cells (VERO-76, ATCC CRL-1 587); human cervical carcinoma cells (HELA, ATCC CCL 2); dog kidney cells (MDCK, ATCC CCL 34); buffalo rat liver cells (BRL 3A, ATCC CRL 1442); human lung cells (W138, ATCC CCL 75); human liver cells (Hep G2, HB 8065); mouse mammary tumor (MMT 060562, ATCC CCL51); TRI cells; MRC5 cells; FS4 cells; and human hepatoma cell line (HepG2). In mammalian cells, the regulatory functions of the expression vector are often provided by viral regulatory elements. For example, commonly used promoters are derived from polyoma, adenovirus 2, cytomegalovirus, and simian virus 40.

[0170] The polypeptide domain of the IL27Rα binding molecule may be produced in a prokaryotic host such as the bacterium Escherichia coli, or in a eukaryotic host such as an insect cell (e.g., Sf21 cell) or a mammalian cell (e.g., COS cell, NIH3T3 cell, or HeLa cell). These cells are available from many suppliers, including the American Type Culture Collection (Manassas, Va.). One of ordinary skill in the art can make such a determination. Further, if guidance is needed in selecting an expression system, one of ordinary skill in the art can consult Ausubel et al. (Current Protocols in Molecular Biology, John Wiley and Sons, New York, N.Y., 1993) and Pouwels et al. (Cloning Vectors: A Laboratory Manual, 1985 Suppl. 1987).

[0171] In some embodiments, the recombinant polypeptide domain of the IL27Rα binding molecule may or may not be glycosylated depending on the host organism used to produce the IL27Rα binding molecule. When bacteria are selected as the host, the polypeptide domain of the IL27Rα binding sdAb may contain glycosylation motifs, particularly N-linked glycosylation motifs of the sequence Asn-X-Ser (N-X-S) or Asn-X-Thr (N-X-T). In the formula, X is any amino acid except proline. In such cases, it is desirable to eliminate the N-linked glycosylation motif by modifying the sequence of such N-linked glycosylation motifs so as to prevent glycosylation. In some embodiments, elimination of the Asn-X-Ser (N-X-S) N-linked glycosylation motif can be accomplished by incorporating conservative amino acid substitutions of the Asn (N) residue and / or the Ser (S) residue of the Asn-X-Ser (N-X-S) N-linked glycosylation motif. In some embodiments, elimination of the Asn-X-Thr (N-X-T) N-linked glycosylation motif can be accomplished by incorporating conservative amino acid substitutions of the Asn (N) residue and / or the Thr (T) residue of the Asn-X-Thr (N-X-T) N-linked glycosylation motif. In some embodiments, when producing recombinant Criss recombinant IL27Rα binding sdAb using a prokaryotic expression system, since the prokaryotic host cell does not provide a glycosylation mechanism for the recombinant protein, modification of the sequence to eliminate N-linked glycosylation sites may not be necessary.

[0172] For additional expression systems for both prokaryotic and eukaryotic cells, see Chapters 16 and 17 of Sambrook et al. (1989) Molecular Cloning: A Laboratory Manual (2nd ed., Cold Spring Harbor Laboratory Press, Plainview, N.Y.). See also Goeddel (1990) in Gene Expression Technology: Methods in Enzymology 185 (Academic Press, San Diego, Calif.).

[0173] Transfection An expression construct can be introduced into a host cell to produce a recombinant polypeptide domain of an IL27Rα binding molecule disclosed herein or a biologically active mutein thereof. Vector DNA can be introduced into prokaryotic or eukaryotic cells by conventional transformation or transfection methods. Suitable methods for transforming or transfecting host cells can be found in Sambrook et al. (1989) Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring Harbor Laboratory Press, Plainview, N.Y.) and other standard molecular biology laboratory manuals.

[0174] To facilitate transfection of target cells, the target cells may be directly exposed to a non-viral vector under conditions that facilitate uptake of the non-viral vector. Examples of conditions that facilitate uptake of foreign nucleic acids by mammalian cells are well known in the art and include, but are not limited to, chemical means (e.g., Lipofectamine®, Thermo-Fisher Scientific), high salt, and magnetic fields (electroporation).

[0175] Cell culture The cells may be cultured in conventional nutrient media that are appropriately modified for induction of a promoter, selection of transformants, or amplification of a gene encoding a desired sequence. Mammalian host cells can be cultured in a variety of media. Suitable media for culturing host cells include commercially available media such as Ham's F10 (Sigma), Minimal Essential Medium ((MEM), Sigma), RPMI 1640 (Sigma), and Dulbecco's Modified Eagle Medium ((DMEM), Sigma). To any of these media, hormones and / or other growth factors (such as insulin, transferrin, or epidermal growth factor), salts (such as sodium chloride, calcium, magnesium, and phosphate), buffers (such as HEPES), nucleosides (such as adenosine and thymidine), antibiotics, trace elements, and glucose or an equivalent energy source can be added as needed. Other optional necessary supplements can also be included at appropriate concentrations known to those skilled in the art. Culture conditions, such as temperature, pH, etc., are those previously used with the host cells selected for expression and will be apparent to those skilled in the art.

[0176] Recovery of Recombinant Protein If a secretion leader sequence is used, the recombinantly produced IL27Rα-binding polypeptide can be recovered from the culture medium as a secreted polypeptide. Alternatively, the IL27Rα-binding polypeptide can also be recovered from host cell lysates. Protease inhibitors such as phenylmethylsulfonyl fluoride (PMSF) may be used during the recovery from cell lysates to inhibit proteolysis, and antibiotics may be included to prevent the growth of exogenous contaminants.

[0177] Purification Various purification processes such as affinity chromatography are known and utilized in the art. In affinity chromatography, typically, highly specific binding sites present in biological macromolecules are utilized to separate molecules that have the ability to bind to a specific ligand. The ligand is presented explicitly to the protein sample, and thereby, the ligand is attached to an insoluble porous support medium by covalent bonding in such a way that a second species is separated and purified from the mixture using the natural specific binding of a certain molecular species. Antibodies are commonly used in affinity chromatography. A size selection step may also be used, and for example, gel filtration chromatography (also known as size exclusion chromatography or molecular sieve chromatography) is used to separate proteins according to size. In gel filtration, a protein solution filled with a semi-permeable porous resin is passed through a column. The semi-permeable resin has a certain range of pore sizes that determine the size of the proteins that can be separated by the column.

[0178] The recombinant polypeptide domain of the IL27Rα binding molecule produced by the transformed host can be purified according to any suitable method. The IL27Rα binding molecule may be isolated from inclusion bodies formed in E. coli using cation exchange, gel filtration, and / or reverse phase liquid chromatography, or may be isolated from the conditioned medium derived from either mammalian or yeast cultures that produce a particular IL27Rα binding molecule.

[0179] The recombinant polypeptide in a substantially purified form can be used as a therapeutic agent, for example, as described herein.

[0180] The biological activity of the recombinant polypeptide domain of the IL27Rα-binding molecule produced according to the foregoing can be confirmed by IL27Rα binding using procedures well known in the art, including, but not limited to, competitive ELISA, radioactive ligand binding assays (e.g., saturation binding, Scatchard plots, non-linear curve fitting programs, and competitive binding assays); non-radioactive ligand binding assays (e.g., fluorescence polarization (FP), fluorescence resonance energy transfer (FRET), and surface plasmon resonance assays (see, e.g., Drescher et al., Methods Mol Biol 493:323-343 (2009)), and measuring devices commercially available from GE Healthcare Bio-Sciences, such as Biacore 8+, Biacore S200, Biacore T200 (GE Healthcare Bio-Sciences, 100 Results Way, Marlborough MA 01752)); liquid phase ligand binding assays (e.g., real-time polymerase chain reaction (RT-qPCR), and immunoprecipitation); and solid phase ligand binding assays (e.g., multiwell plate assays, on-bead ligand binding assays, on-column ligand binding assays, and filter assays).

[0181] Method of Use Inhibition of IL27Rα Activity In one aspect, the disclosure provides a method of modulating the activity of IL27Rα-expressing cells by administering to a subject an IL27Rα-binding molecule in an amount sufficient to interfere with the activity of a receptor comprising IL27Rα. Further, the disclosure provides a method of modulating the activity of IL27Rα-expressing cells in a mixed cell population, the method comprising contacting the cell population, in vivo and / or ex vivo, with an IL27Rα-binding molecule or complex of the disclosure in an amount sufficient to interfere with the activity of a receptor comprising IL27Rα.

[0182] Identification, Isolation, Enrichment, or Depletion of IL27Rα+ Cells In one aspect, the present disclosure provides a method of using the IL27Rα binding molecule of the present disclosure useful in a process for isolating, enriching, or depleting IL27Rα+ cells from a biological sample containing IL27Rα+ cells. The biological sample may include T cells, B cells, blood-derived cells such as PBMCs, cells of cell culture origin, or cells of tissue origin such as brain or bone marrow. Processes suitable for isolation, enrichment, or depletion of IL27Rα+ cells include centrifugation, filtration, magnetic cell sorting, and fluorescence-activated cell sorting by techniques well known in the art. Further, the present disclosure provides a method for treating a subject suffering from a disease, disorder, or condition by administering a therapeutically effective amount of a cell product in which IL27Rα+ cells have been enriched or depleted by use of an IL27Rα binding molecule as described herein.

[0183] In one aspect, in a sorting procedure, an IL27Rα binding molecule comprising a fluorescent label is used for FACS isolation or depletion of IL27Rα+ cells from a sample. The fluorescent label may be attached directly to the sdAb of the IL27Rα binding molecule (e.g., by chemical conjugation optionally using a linker), or indirectly (e.g., by biotinylation of the sdAb and binding of the biotinylated antibody to a streptavidin-fluorophore conjugate). Such fluorescently labeled IL27Rα+ cells can be separated from a mixed cell population using conventional FACS techniques.

[0184] In another aspect, in the selection procedure, for use in a magnetic cell separation procedure, the IL27Rα-binding molecules of the present disclosure (e.g., IL27Rα-binding VHHs) conjugated to magnetic particles that effect magnetic labeling of IL27Rα+ cells are used. In one aspect, the method comprises: (a) conjugating one or more IL27Rα-binding molecules of the present disclosure (e.g., IL27Rα-binding VHHs) to magnetic particles; (b) creating a mixture by contacting a biological sample with an amount of magnetic particles conjugated to an IL27Rα-binding molecule; (c) subjecting the mixture to a magnetic field such that magnetically labeled IL27Rα+ cells are retained; (d) removing non-magnetically labeled cells from the mixture; and (e) removing the magnetic field to isolate the IL27Rα+ cells.

[0185] By a cell selection procedure (e.g., FACS or magnetic separation), two products are obtained: (a) a cell population depleted of IL27Rα+ cells and (b) a cell population enriched in IL27Rα+ cells. Each of these populations can be further processed by conventional procedures that identify specific IL27Rα+ cell subsets or IL27Rα− cell subsets that may be useful in research, diagnostic, or clinical applications. For example, isolation of specific IL27Rα+ T cell subsets that also express one or more of CD4, CD8, CD19, CD25, and CD62L, by FACS or magnetic field separation by techniques well known in the art, and further iterations of the use of one or more antibodies that specifically bind to the CD4, CD8, CD19, CD25, and CD62L antigens, respectively.

[0186] In one aspect, the IL27Rα binding molecule can be used to deplete IL27Rα-expressing cells from a biological sample, such as peripheral blood or lymphoid tissue, which can optionally be further processed for the further isolation of IL27Rα+ naive T cell subsets, the isolation of human IL27Rα+ memory T cells from a population of CD4+ or CD8+ cells, or the isolation of human IL27RαRA+ naive T cells from preselected CD4+ or CD8+ cells by depletion of IL27Rα+ cells. In one aspect, the IL27Rα binding molecule is a method of generating a population of cells enriched for naive Tregs from a biological sample, comprising the step of depleting IL27Rα+ cells using the IL27Rα binding molecule of the present disclosure as described above, and optionally further comprising the step of depleting CD8+ and / or CD19+ cells. The IL27Rα+ depleted cell population can optionally be further expanded in vitro for the preparation of a cell product containing a therapeutically effective amount of an IL27Rα+ depleted cell product for administration to a subject suffering from a disease, disorder, or condition. The IL27Rα+ enriched cell population can optionally be further expanded in vitro for the preparation of a cell product containing a therapeutically effective amount of IL27Rα+ cells.

[0187] Kit The present disclosure also contemplates kits comprising pharmaceutical compositions of IL27Rα binding molecules. Kits generally take the form of a physical structure that houses various components, as described below, and can be utilized, for example, in the practice of the above methods. The kits of the present disclosure can be designed for the conditions (e.g., refrigeration or freezing) necessary to properly maintain the components housed therein. The kit may further comprise a label or package insert that describes the identifying information of the components within the kit, and instructions for use. Each component of the kit may be enclosed in a separate container, or the various containers may all be within one packaging container. The label or package insert may contain manufacturer information such as lot number and expiration date. The label or package insert may be integrated, for example, on the surface of the physical structure that houses the components, separately enclosed within the physical structure, or affixed to a component of the kit (e.g., an ampule, syringe, or vial). The label or package insert may be provided in physical form or as a computer-readable medium. In some embodiments, no actual instructions are present in the kit, and the kit provides means for obtaining instructions from a remote information source, such as via an Internet site, including obtaining the instructions by secure access provided by a password (or a scannable code such as a barcode or QR code on the container of the IL27Rα binding molecule or on the surface of the kit containing it), in accordance with government regulations (e.g., HIPAA).

Example

[0188] The following examples are provided to fully disclose and describe to those skilled in the art how to make and use the present IL27Rα binding molecule, and are not intended to limit the scope of what the inventors regard as the IL27Rα binding molecule, nor are they intended to represent that the following experiments were all the experiments that could be carried out or that all the experiments that could be carried out. The exemplary descriptions written in the present tense are not necessarily carried out, but this description should be understood to be capable of being carried out in order to obtain the data etc. described in the description. Efforts have been made to ensure accuracy for the numerical values used (for example, amounts, temperatures, etc.), but some experimental errors and deviations should be expected. Variations of the procedures described in detail that are used may be apparent to those skilled in the art, and those skilled in the art are expected to be able to use such variations as appropriate. Accordingly, the IL27Rα binding molecule may be practiced in ways other than those specifically described herein, and the present invention is intended to cover all modifications and equivalents of the subject matter described in the claims appended hereto as permitted by applicable law.

[0189] Unless otherwise specified, parts are parts by weight, molecular weights are weight average molecular weights, temperatures are in degrees Celsius (°C), and pressures are at atmospheric pressure or near atmospheric pressure. The following standard abbreviations are used: bp = base pair(s); kb = kilobase(s); pl = picoliter; s or sec = second; min = minute; h or hr = hour; aa = amino acid(s); kb = kilobase(s); nt = nucleotide(s); pg = picogram; ng = nanogram; μg = microgram; mg = milligram; g = gram; kg = kilogram; dl or dL = deciliter; μl or μL = microliter; ml or mL = milliliter; l or L = liter; μM = micromolar concentration; mM = millimolar concentration; M = molar concentration; kDa = kilodalton; i.m. = intramuscular (into muscle); i.p. = intraperitoneal (into the peritoneal cavity); SC or SQ = subcutaneous (subcutaneously); QD = once daily; BID = twice daily; QW = once weekly; QM = once monthly; HPLC = high performance liquid chromatography; BW = body weight; U = unit; ns = not statistically significant; PBS = phosphate buffered saline; PCR = polymerase chain reaction; NHS = N-hydroxysuccinimide; HSA = human serum albumin; MSA = mouse serum albumin; DMEM = Dulbecco's modified Eagle's medium; GC = genomic copy; EDTA = ethylenediaminetetraacetic acid; PBMC = peripheral blood mononuclear cells; FBS = fetal bovine serum; FCS = fetal calf serum; HEPES = 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; LPS = lipopolysaccharide; ATCC = American Type Culture Collection.

[0190] Example 1. Immunization protocol The process for isolating anti-hIL27Rα VHH was initiated by immunizing camels with a polypeptide corresponding to amino acids 33 - 516 of hIL27Rα (UNIPROT reference number Q6UWB1). The process for isolating anti-mIL27Rα VHH was initiated by immunizing camels with the 201 amino acid extracellular domain of mIL27Rα, amino acids 25 - 510 of the mIL27Rα precursor (UNIPROT reference number O70394). For each antigen, VHHs were identified and isolated using the following methodology.

[0191] The synthetic DNA sequence encoding the antigen was inserted into the pFUSE_hIgG1_Fc2 vector (Generay Biotechnology) and introduced into HEK293F mammalian cell host cells by transfection for expression. The antigen was expressed as an Fc fusion protein purified using protein A chromatography. The antigen was diluted with 1×PBS (about 1 mg of antigen in total). Quality was evaluated by SDS-PAGE to ensure that the purity was sufficient for immunization (>80%). Camels were acclimatized at the facility for at least 7 days prior to immunization. Immunization with the antigen was performed using weekly antigen administration over 7 weeks. For the first immunization, the immunogen was prepared as follows. 10 mL of complete Freund's adjuvant (CFA) was added to a mortar, and then 10 mL of antigen dissolved in 1×PBS was slowly added to the mortar while grinding with a pestle until the antigen was emulsified and became milky white and difficult to disperse. The sample was then ground. Thereafter, the immunogen was prepared as above except that incomplete Freund's adjuvant (IFA) was used instead of CFA for immunization (weeks 2 - 7) six times in the immunization protocol. Approximately 2 mL of emulsified antigen, for a total of about 10 mL per camel, was subcutaneously injected at at least 6 sites on the camel. When injecting the antigen, the needle was maintained in the subcutaneous space for about 10 - 15 seconds after each injection to avoid leakage of the emulsion.

[0192] Example 2. Phage library construction In the immunization protocol, blood samples were collected from camels 3 days after the last injection. RNA was extracted from the blood and transcribed into cDNA. An approximately 900 bp reverse transcription sequence encoding a VH-CH1-hinge-CH2-CH3 construct was isolated from an approximately 700 bp fragment encoding the desired VHH-hinge-CH2-CH3 species. The purified approximately 700 bp fragment was amplified by nested PCR. The amplified sequence was digested with Pst1 and Not1. An approximately 400 bp PST1 / Not1 digested fragment was inserted into the Pst1 / Not1 digested pMECS phagemid vector such that the VHH coding sequence was in-frame with the DNA sequence encoding the HA / His sequence. The sequence generated by PCR and the pMECS phagemid vector were digested with PstI and NotI and then ligated into the pMECS / Nb recombination. After ligation, the product was introduced into Escherichia coli (E. coli) TG1 cells by transformation via electroporation. After concentrating the transformants in the growth medium, they were transferred to 2YT + 2% glucose agar plates.

[0193] Example 3: Isolation of antigen-specific VHH Biopanning of the phage library was performed to identify VHHs that bind to IL27Rα. A 96-well plate was coated with IL27Rα and the phage library was incubated in each well such that phage-expressed IL27Rα-reactive VHHs could bind to the IL27Rα present on the plate. Non-specifically bound phages were washed away and specifically bound phages were isolated. After selection, the enriched phage library expressing IL27Rα-reactive VHHs was amplified in TG1 cells. The above biopanning process was repeated 2 - 3 times to enrich the library for IL27Rα-reactive VHHs.

[0194] Example 4: Identification of antibodies that specifically bind to IFNgR1: After the biopanning of Example 3 was completed, periplasmic extract ELISA (PE-ELISA) was performed on an IL27Rα-coated plate to identify positive VHH binders that selectively bind to IFNgR1. Three 96-well plates of individual phage clones were isolated. The 96-well plates were coated with IL27Rα and PBS under the same conditions. Next, the wells were blocked at 37 °C for 1 hour. Then, 100 μl of the extracted antibody was added to each well and incubated for 1 hour. Thereafter, 100 μl of the HRP-conjugated anti-tag polyclonal antibody was added to each well and incubated at 37 °C for 1 hour. The plate was developed with a TMB substrate. The reaction was stopped by adding H2SO4. The absorbance at 450 nm was read with a microtiter plate reader. Antibodies with an absorbance in the antigen-coated wells at least three times that of the PBS-coated control were defined as showing specific binding to IL27Rα. The positive clones were sequenced and the sequences were analyzed to identify unique chronotypes.

[0195] Example 5. Evaluation of binding affinity via surface plasmon resonance To evaluate the binding via SPR as follows, representative examples from each hIL27Rα VHH clone prepared according to Examples 1 to 3 were selected. The binding affinity of the hIL27Rα binding molecules corresponding to SEQ ID NOs: 2 - 27 was evaluated using surface plasmon resonance (SPR) generally according to the following procedure. All experiments were performed on a Biacore T200 instrument equipped with a Protein A-derivatized sensor chip (Cytiva) in 10 mM Hepes, 150 mM NaCl, 0.05% (v / v) polysorbate 20 (PS20), and 3 mM EDTA (HBS-EP+ buffer). The mono-Fc VHH ligand was flowed at 5 μl / min for a variable time of 18 - 300 seconds to reach the capture load listed in the following table. After ligand capture, a two-fold dilution series of the extracellular domain of the IL27Rα receptor modified to incorporate a C-terminal poly-His sequence, typically containing at least five concentrations from 1 μM to 1 nM, was injected in high-performance mode or single-cycle kinetics mode. Surface regeneration was performed by flowing 10 mM glycine-HCl, pH 1.5 (60 seconds, 50 μL / min). The sensogram minus buffer was processed with Biacore T200 Evaluation Software and globally fit using a 1:1 Langmuir binding model (with bulk shift set to 0) to extract the rate constants and affinity constants (k a k d , K D ). R MAX <100 RU indicates a surface density compatible with kinetic analysis. R max The calculated value was generated using the formula: Rmax = load (RU) x valence of the ligand x (molecular weight of the analyte / molecular weight of the ligand). Surface activity was defined as the ratio of the experimental value of Rmax to the calculated value. The results of these binding affinity experiments are shown in Table 6.

[0196] The examples and aspects described in this specification are for illustrative purposes only, and various modifications or changes may be suggested to those skilled in the art in view of the examples and aspects described in this specification, and it is understood that they are included within the spirit and scope of this application and the appended claims. All publications, sequence accession numbers, patents, and patent applications cited herein are hereby incorporated by reference in their entirety for all purposes.

[0197] Sequence information SEQUENCE LISTING <110> SYNTHEKINE, INC. <120> IL27RALPHA BINDING MOLECULES AND METHODS OF USE <150> US 63 / 135,884 <151> 2021-01-11 <150> US 63 / 078,745 <151> 2020-09-15 <150> US 63 / 061,562 <151> 2020-08-05 <160> 202 <170> PatentIn version 3.5 <210> 1 <211> 636 <212> PRT <213> Homo sapiens <400> 1 Met Arg Gly Gly Arg Gly Ala Pro Phe Trp Leu Trp Pro Leu Pro Lys 1 5 10 15 Leu Ala Leu Leu Pro Leu Leu Trp Val Leu Phe Gln Arg Thr Arg Pro 20 25 30 Gln Gly Ser Ala Gly Pro Leu Gln Cys Tyr Gly Val Gly Pro Leu Gly 35 40 45 Asp Leu Asn Cys Ser Trp Glu Pro Leu Gly Asp Leu Gly Ala Pro Ser 50 55 60 Glu Leu His Leu Gln Ser Gln Lys Tyr Arg Ser Asn Lys Thr Gln Thr 65 70 75 80 Val Ala Val Ala Ala Gly Arg Ser Trp Val Ala Ile Pro Arg Glu Gln 85 90 95 Leu Thr Met Ser Asp Lys Leu Leu Val Trp Gly Thr Lys Ala Gly Gln 100 105 110 Pro Leu Trp Pro Pro Val Phe Val Asn Leu Glu Thr Gln Met Lys Pro 115 120 125 Asn Ala Pro Arg Leu Gly Pro Asp Val Asp Phe Ser Glu Asp Asp Pro 130 135 140 Leu Glu Ala Thr Val His Trp Ala Pro Pro Thr Trp Pro Ser His Lys 145 150 155 160 Val Leu Ile Cys Gln Phe His Tyr Arg Arg Cys Gln Glu Ala Ala Trp 165 170 175 Thr Leu Leu Glu Pro Glu Leu Lys Thr Ile Pro Leu Thr Pro Val Glu 180 185 190 Ile Gln Asp Leu Glu Leu Ala Thr Gly Tyr Lys Val Tyr Gly Arg Cys 195 200 205 Arg Met Glu Lys Glu Glu Asp Leu Trp Gly Glu Trp Ser Pro Ile Leu 210 215 220 Ser Phe Gln Thr Pro Pro Ser Ala Pro Lys Asp Val Trp Val Ser Gly 225 230 235 240 Asn Leu Cys Gly Thr Pro Gly Gly Glu Glu Pro Leu Leu Leu Trp Lys 245 250 255 Ala Pro Gly Pro Cys Val Gln Val Ser Tyr Lys Val Trp Phe Trp Val 260 265 270 Gly Gly Arg Glu Leu Ser Pro Glu Gly Ile Thr Cys Cys Cys Ser Leu 275 280 285 Ile Pro Ser Gly Ala Glu Trp Ala Arg Val Ser Ala Val Asn Ala Thr 290 295 300 Ser Trp Glu Pro Leu Thr Asn Leu Ser Leu Val Cys Leu Asp Ser Ala 305 310 315 320 Ser Ala Pro Arg Ser Val Ala Val Ser Ser Ile Ala Gly Ser Thr Glu 325 330 335 Leu Leu Val Thr Trp Gln Pro Gly Pro Gly Glu Pro Leu Glu His Val 340 345 350 Val Asp Trp Ala Arg Asp Gly Asp Pro Leu Glu Lys Leu Asn Trp Val 355 360 365 Arg Leu Pro Pro Gly Asn Leu Ser Ala Leu Leu Pro Gly Asn Phe Thr 370 375 380 Val Gly Val Pro Tyr Arg Ile Thr Val Thr Ala Val Ser Ala Ser Gly 385 390 395 400 Leu Ala Ser Ala Ser Ser Val Trp Gly Phe Arg Glu Glu Leu Ala Pro 405 410 415 Leu Val Gly Pro Thr Leu Trp Arg Leu Gln Asp Ala Pro Pro Gly Thr 420 425 430 Pro Ala Ile Ala Trp Gly Glu Val Pro Arg His Gln Leu Arg Gly His 435 440 445 Leu Thr His Tyr Thr Leu Cys Ala Gln Ser Gly Thr Ser Pro Ser Val 450 455 460 Cys Met Asn Val Ser Gly Asn Thr Gln Ser Val Thr Leu Pro Asp Leu 465 470 475 480 Pro Trp Gly Pro Cys Glu Leu Trp Val Thr Ala Ser Thr Ile Ala Gly 485 490 495 Gln Gly Pro Pro Gly Pro Ile Leu Arg Leu His Leu Pro Asp Asn Thr 500 505 510 Leu Arg Trp Lys Val Leu Pro Gly Ile Leu Phe Leu Trp Gly Leu Phe 515 520 525 Leu Leu Gly Cys Gly Leu Ser Leu Ala Thr Ser Gly Arg Cys Tyr His 530 535 540 Leu Arg His Lys Val Leu Pro Arg Trp Val Trp Glu Lys Val Pro Asp 545 550 555 560 Pro Ala Asn Ser Ser Ser Gly Gln Pro His Met Glu Gln Val Pro Glu 565 570 575 Ala Gln Pro Leu Gly Asp Leu Pro Ile Leu Glu Val Glu Glu Met Glu 580 585 590 Pro Pro Pro Val Met Glu Ser Ser Gln Pro Ala Gln Ala Thr Ala Pro 595 600 605 Leu Asp Ser Gly Tyr Glu Lys His Phe Leu Pro Thr Pro Glu Glu Leu 610 615 620 Gly Leu Leu Gly Pro Pro Arg Pro Gln Val Leu Ala 625 630 635 <210> 2 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 2 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Pro Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Ile 35 40 45 Ser Thr Ile Ser Ala Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ser Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Ala Ala Ile Tyr Tyr Cys 85 90 95 Ala Lys Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Arg Arg Asn Tyr 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 3 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 3 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Ser 20 25 30 Gly Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ile Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 100 105 110 Gly Ser Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 4 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 4 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Tyr Val Ser Cys Asp Tyr 20 25 30 Phe Leu Pro Ser Trp Tyr Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe 35 40 45 Val Ser Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val 50 55 60 Lys Gly Arg Phe Thr Ala Ser Glu Asp Lys Gly Lys Asn Ile Ala Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Lys Ala Ser Cys Val Arg Gly Arg Ala Val Ser Glu Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 5 <211> 124 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 5 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Glu 1 5 10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Asn Val Ala Tyr Gly Ile Thr Ser Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Thr Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Ile Tyr Tyr Cys 85 90 95 Val Lys His Ser Gly Thr Thr Ile Pro Arg Gly Phe Ile Ser Tyr Thr 100 105 110 Lys Arg Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 6 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 6 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Tyr Val Ser Cys Asp Tyr 20 25 30 Phe Leu Pro Ser Trp Tyr Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe 35 40 45 Val Ser Val Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val 50 55 60 Lys Gly Arg Phe Thr Ala Ser Gln Asp Lys Gly Lys Asn Ile Ala Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Lys Ala Ser Cys Val Arg Gly Arg Ala Ile Ser Glu Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 7 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 7 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Ser Phe Ser Ser Tyr 20 25 30 Ala Met Lys Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Thr Ile Ser Ser Gly Gly Ser Ser Thr Asn Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Ile Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Lys Ala Ile Val Pro Thr Gly Ala Thr Met Glu Arg Gly Gln Gly 100 105 110 Thr Gln Val Thr Val Ser Ser 115 <210> 8 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 8 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Pro Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Ile 35 40 45 Ser Thr Ile Ser Ala Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ser Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Ile Tyr Tyr Cys 85 90 95 Ala Lys Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Arg Arg Asn Tyr 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 9 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 9 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Val Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Pro Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Ile 35 40 45 Ser Thr Ile Ser Ala Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ser Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Ile Tyr Tyr Cys 85 90 95 Ala Lys Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Arg Arg Asn Tyr 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 10 <211> 128 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 10 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ser Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Tyr Ser Thr Ser 20 25 30 Asn Ser Trp Met Ala Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu 35 40 45 Gly Val Ala Ala Ile Tyr Thr Val Gly Gly Ser Ile Phe Tyr Ala Asp 50 55 60 Ser Val Arg Gly Arg Phe Thr Ile Ser Gln Asp Ala Thr Lys Asn Met 65 70 75 80 Phe Tyr Leu Gln Met Asn Thr Leu Lys Pro Glu Asp Thr Ala Met Tyr 85 90 95 Tyr Cys Ala Ala Ala Ser Gly Arg Leu Arg Gly Lys Trp Phe Trp Pro 100 105 110 Tyr Glu Tyr Asn Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 125 <210> 11 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 11 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Gly Ser Thr Tyr Ser Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Ile Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp Gln Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Asp Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 12 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 12 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Gly Ser Thr Tyr Ser Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Ser Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp His Ala Lys Asn Thr Val Thr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Gly Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 13 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 13 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Glu 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Gly Ser Thr Tyr Ser Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Ile Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp Gln Ala Lys Asn Thr Val Tyr 65 70 75 80 Leu Glu Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Thr Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Asp Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 14 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 14 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Tyr Val Ser Cys Asp Tyr 20 25 30 Phe Leu Pro Ser Trp Tyr Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe 35 40 45 Val Ser Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val 50 55 60 Lys Gly Arg Phe Thr Ala Ser Gln Asp Arg Gly Lys Asn Ile Ala Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Lys Ala Ser Cys Val Arg Gly Arg Thr Ile Ser Glu Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 15 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 15 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Tyr Val Ser Cys Asp Tyr 20 25 30 Phe Leu Pro Ser Trp Tyr Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe 35 40 45 Val Ser Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val 50 55 60 Lys Gly Arg Phe Thr Ala Ser Gln Asp Lys Gly Lys Asn Ile Ala Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Lys Ala Ser Cys Val Arg Gly Arg Ala Ile Ser Glu Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 16 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 16 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Tyr Val Ser Cys Asp Tyr 20 25 30 Phe Leu Pro Ser Trp Tyr Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe 35 40 45 Val Ser Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val 50 55 60 Lys Gly Arg Phe Thr Ala Ser Gln Asp Lys Gly Lys Asn Ile Ala Tyr 65 70 75 80 Leu Gln Met Asn Thr Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Lys Ala Ser Cys Val Arg Gly Arg Ala Ile Ser Glu Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 17 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 17 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Gly Ser Thr Tyr Ser Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Ile Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp Gln Ala Lys Asn Thr Val Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Glu Ser Ala Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Asp Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 18 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 18 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Ser 20 25 30 Gly Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Met Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 100 105 110 Gly Ser Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 19 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 19 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Tyr Val Ser Cys Asp Tyr 20 25 30 Phe Leu Pro Ser Trp Tyr Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe 35 40 45 Val Ser Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val 50 55 60 Lys Gly Arg Phe Thr Ala Ser Gln Asp Lys Gly Lys Asn Ile Ala Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Lys Ala Ser Cys Val Arg Gly Arg Gly Ile Ser Glu Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 20 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 20 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Gly Ser Thr Tyr Ser Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Ile Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp Gln Ala Lys Asn Thr Val Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Asp Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 21 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 21 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Pro Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Thr Ile Ser Ser Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ser Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Lys Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Lys Arg Ser Tyr 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 22 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 22 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Ser 20 25 30 Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Met Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 100 105 110 Gly Ser Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 23 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 23 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Gly Ser Thr Tyr Ser Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Thr Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp Gln Ala Lys Asn Thr Val Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Asp Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 24 <211> 129 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 24 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Arg Ala Ser Arg Ser Pro Tyr Gly Asn Tyr 20 25 30 Cys Leu Gly Trp Phe Arg Gln Ser Thr Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Gln Asp His Ala Lys Asn Thr Val Thr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg 100 105 110 Pro Asp Arg Val Pro Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser 115 120 125 Ser <210> 25 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 25 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser His Ser 20 25 30 Gly Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Thr Ile Asn Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Met Leu Tyr 65 70 75 80 Leu Gln Leu Asn Ser Leu Lys Thr Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 100 105 110 Gly Ser Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 26 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 26 Phe Thr Phe Ser Ser Tyr Pro Met Ser 1 5 <210> 27 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 27 Thr Ile Ser Ala Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 28 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 28 Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Arg Arg Asn Tyr 1 5 10 <210> 29 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 29 Phe Thr Phe Ser Leu Ser Gly Met Ser 1 5 <210> 30 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 30 Ala Ile Ser Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val Lys 1 5 10 15 Gly <210> 31 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 31 Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 1 5 10 <210> 32 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 32 Tyr Val Ser Cys Asp Tyr Phe Leu Pro Ser 1 5 10 <210> 33 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 33 Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val Lys Gly 1 5 10 15 <210> 34 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 34 Ser Cys Val Arg Gly Arg Ala Val Ser Glu Tyr 1 5 10 <210> 35 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 35 Phe Thr Phe Ser Asn Tyr Ala Met Ser 1 5 <210> 36 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 36 Gly Ile Asn Val Ala Tyr Gly Ile Thr Ser Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 37 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 37 His Ser Gly Thr Thr Ile Pro Arg Gly Phe Ile Ser Tyr Thr Lys 1 5 10 15 <210> 38 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 38 Tyr Val Ser Cys Asp Tyr Phe Leu Pro Ser 1 5 10 <210> 39 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 39 Val Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val Lys Gly 1 5 10 15 <210> 40 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 40 Ser Cys Val Arg Gly Arg Ala Ile Ser Glu Tyr 1 5 10 <210> 41 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 41 Phe Ser Phe Ser Ser Tyr Ala Met Lys 1 5 <210> 42 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 42 Thr Ile Ser Ser Gly Gly Ser Ser Thr Asn Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 43 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 43 Ala Ile Val Pro Thr Gly Ala Thr Met Glu 1 5 10 <210> 44 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 44 Phe Thr Phe Ser Ser Tyr Pro Met Ser 1 5 <210> 45 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 45 Thr Ile Ser Ala Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 46 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 46 Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Arg Arg Asn Tyr 1 5 10 <210> 47 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 47 Phe Thr Phe Ser Ser Tyr Pro Met Ser 1 5 <210> 48 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 48 Thr Ile Ser Ala Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 49 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 49 Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Arg Arg Asn Tyr 1 5 10 <210> 50 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 50 Phe Thr Tyr Ser Thr Ser Asn Ser Trp Met Ala 1 5 10 <210> 51 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 51 Ala Ile Tyr Thr Val Gly Gly Ser Ile Phe Tyr Ala Asp Ser Val Arg 1 5 10 15 Gly <210> 52 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 52 Ala Ser Gly Arg Leu Arg Gly Lys Trp Phe Trp Pro Tyr Glu Tyr Asn 1 5 10 15 Tyr <210> 53 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 53 Ser Thr Tyr Ser Asn Tyr Cys Leu Gly 1 5 <210> 54 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 54 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 55 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 55 Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Asp 1 5 10 15 Arg Val Pro Tyr 20 <210> 56 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 56 Ser Thr Tyr Ser Asn Tyr Cys Leu Gly 1 5 <210> 57 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 57 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 58 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 58 Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Gly 1 5 10 15 Arg Val Pro Tyr 20 <210> 59 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 59 Ser Thr Tyr Ser Asn Tyr Cys Leu Gly 1 5 <210> 60 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 60 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 61 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 61 Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Asp 1 5 10 15 Arg Val Pro Tyr 20 <210> 62 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 62 Tyr Val Ser Cys Asp Tyr Phe Leu Pro Ser 1 5 10 <210> 63 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 63 Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val Lys Gly 1 5 10 15 <210> 64 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 64 Ser Cys Val Arg Gly Arg Thr Ile Ser Glu Tyr 1 5 10 <210> 65 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 65 Tyr Val Ser Cys Asp Tyr Phe Leu Pro Ser 1 5 10 <210> 66 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 66 Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val Lys Gly 1 5 10 15 <210> 67 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 67 Ser Cys Val Arg Gly Arg Ala Ile Ser Glu Tyr 1 5 10 <210> 68 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 68 Tyr Val Ser Cys Asp Tyr Phe Leu Pro Ser 1 5 10 <210> 69 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 69 Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val Lys Gly 1 5 10 15 <210> 70 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 70 Ser Cys Val Arg Gly Arg Ala Ile Ser Glu Tyr 1 5 10 <210> 71 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 71 Ser Thr Tyr Ser Asn Tyr Cys Leu Gly 1 5 <210> 72 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 72 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 73 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 73 Glu Ser Ala Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Asp 1 5 10 15 Arg Val Pro Tyr 20 <210> 74 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 74 Phe Thr Phe Ser Leu Ser Gly Met Ser 1 5 <210> 75 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 75 Ala Ile Ser Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val Lys 1 5 10 15 Gly <210> 76 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 76 Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 1 5 10 <210> 77 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 77 Tyr Val Ser Cys Asp Tyr Phe Leu Pro Ser 1 5 10 <210> 78 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 78 Ile Ile Asp Gly Thr Gly Ser Thr Ser Tyr Ala Ala Ser Val Lys Gly 1 5 10 15 <210> 79 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 79 Ser Cys Val Arg Gly Arg Gly Ile Ser Glu Tyr 1 5 10 <210> 80 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 80 Ser Thr Tyr Ser Asn Tyr Cys Leu Gly 1 5 <210> 81 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 81 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 82 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 82 Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Asp 1 5 10 15 Arg Val Pro Tyr 20 <210> 83 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 83 Phe Thr Phe Ser Ser Tyr Pro Met Ser 1 5 <210> 84 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 84 Thr Ile Ser Ser Gly Gly Asp Thr Thr Leu Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 85 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 85 Arg Ile Asp Cys Asn Ser Gly Tyr Cys Tyr Lys Arg Ser Tyr 1 5 10 <210> 86 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 86 Phe Thr Phe Ser Leu Ser Ser Met Ser 1 5 <210> 87 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 87 Ala Ile Ser Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val Lys 1 5 10 15 Gly <210> 88 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 88 Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 1 5 10 <210> 89 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 89 Ser Thr Tyr Ser Asn Tyr Cys Leu Gly 1 5 <210> 90 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 90 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 91 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 91 Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Asp 1 5 10 15 Arg Val Pro Tyr 20 <210> 92 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 92 Ser Pro Tyr Gly Asn Tyr Cys Leu Gly 1 5 <210> 93 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 93 Val Ile Asn Trp Val Gly Gly Met Leu Tyr Phe Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 94 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 94 Glu Ser Val Ser Ser Phe Ser Cys Gly Gly Trp Leu Thr Arg Pro Asp 1 5 10 15 Arg Val Pro Tyr 20 <210> 95 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 95 Phe Thr Phe Ser His Ser Gly Met Ser 1 5 <210> 96 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 96 Thr Ile Asn Ser Gly Gly Ala Ser Thr Tyr Tyr Thr Asp Ser Val Lys 1 5 10 15 Gly <210> 97 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 97 Gly Gly Ser Gly Tyr Gly Asp Ala Ser Arg Met Thr Ser Pro 1 5 10 <210> 98 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 98 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc agctacccca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg gatcagcacc atcagcgccg gcggcgacac caccctgtac 180 gccgacagcg tgaagggcag gttcaccagc agcagggaca acgccaagaa caccctgtac 240 ctgcagctga acagcctgaa gaccgaggac gccgccatct actactgcgc caagaggatc 300 gactgcaaca gcggctactg ctacaggagg aactactggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 99 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 99 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc ctgagcggca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcgcc atcagcagcg gcggcgccag cacctactac 180 accgacagcg tgaagggcag gttcaccatc agcagggaca acgccaagaa catcctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatgt actactgcgc caagggcggc 300 agcggctacg gcgacgccag caggatgacc agccccggca gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 100 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 100 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgtgg ccagcggcta cgtgagctgc gactacttcc tgcccagctg gtacaggcag 120 gcccccggca aggagaggga gttcgtgagc atcatcgacg gcaccggcag caccagctac 180 gccgccagcg tgaagggcag gttcaccgcc agcgaggaca agggcaagaa catcgcctac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcaa ggccagctgc 300 gtgaggggca gggccgtgag cgagtactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 101 <211> 372 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 101 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcgagag cctgaggctg 60 agctgcaccg ccagcggctt caccttcagc aactacgcca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcggc atcaacgtgg cctacggcat caccagctac 180 gccgacagcg tgaagggcag gttcaccatc agcagggaca acaccaagaa caccctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatct actactgcgt gaagcacagc 300 ggcaccacca tccccagggg cttcatcagc tacaccaaga ggggccaggg cacccaggtg 360 accgtgagca gc 372 <210> 102 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 102 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaccg ccagcggcta cgtgagctgc gactacttcc tgcccagctg gtacaggcag 120 gcccccggca aggagaggga gttcgtgagc gtgatcgacg gcaccggcag caccagctac 180 gccgccagcg tgaagggcag gttcaccgcc agccaggaca agggcaagaa catcgcctac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcaa ggccagctgc 300 gtgaggggca gggccatcag cgagtactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 103 <211> 357 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 103 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt cagcttcagc agctacgcca tgaagtgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcacc atcagcagcg gcggcagcag caccaactac 180 gccgacagcg tgaagggcag gttcaccatc agcagggaca acgccaagaa caccctgtac 240 ctgcagctga acagcctgaa gatcgaggac accgccatgt actactgcgc caaggccatc 300 gtgcccaccg gcgccaccat ggagaggggc cagggcaccc aggtgaccgt gagcagc 357 <210> 104 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 104 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc agctacccca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg gatcagcacc atcagcgccg gcggcgacac caccctgtac 180 gccgacagcg tgaagggcag gttcaccagc agcagggaca acgccaagaa caccctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatct actactgcgc caagaggatc 300 gactgcaaca gcggctactg ctacaggagg aactactggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 105 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 105 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg tgggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc agctacccca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg gatcagcacc atcagcgccg gcggcgacac caccctgtac 180 gccgacagcg tgaagggcag gttcaccagc agcagggaca acgccaagaa caccctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatct actactgcgc caagaggatc 300 gactgcaaca gcggctactg ctacaggagg aactactggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 106 <211> 384 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 106 caggtgcagc tgcaggagag cggcggcggc agcgtgcaga gcggcggcag cctgaggctg 60 agctgcgccg ccagcggctt cacctacagc accagcaaca gctggatggc ctggttcagg 120 caggcccccg gcaaggagag ggagggcgtg gccgccatct acaccgtggg cggcagcatc 180 ttctacgccg acagcgtgag gggcaggttc accatcagcc aggacgccac caagaacatg 240 ttctacctgc agatgaacac cctgaagccc gaggacaccg ccatgtacta ctgcgccgcc 300 gccagcggca ggctgagggg caagtggttc tggccctacg agtacaacta ctggggccag 360 ggcacccagg tgaccgtgag cagc 384 <210> 107 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 107 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaggg ccagcggcag cacctacagc aactactgcc tgggctggtt caggcagatc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc aggccaagaa caccctgtac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgc cgccgagagc 300 gtgagcagct tcagctgcgg cggctggctg accaggcccg acagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 108 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 108 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaggg ccagcggcag cacctacagc aactactgcc tgggctggtt caggcagagc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc acgccaagaa caccgtgacc 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgc cgccgagagc 300 gtgagcagct tcagctgcgg cggctggctg accaggcccg gcagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 109 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 109 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcgagag cctgaggctg 60 agctgcaggg ccagcggcag cacctacagc aactactgcc tgggctggtt caggcagatc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc aggccaagaa caccgtgtac 240 ctggagatga acagcctgaa gcccgaggac accgccatgt actactgcgc caccgagagc 300 gtgagcagct tcagctgcgg cggctggctg accaggcccg acagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 110 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 110 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgtgg ccagcggcta cgtgagctgc gactacttcc tgcccagctg gtacaggcag 120 gcccccggca aggagaggga gttcgtgagc atcatcgacg gcaccggcag caccagctac 180 gccgccagcg tgaagggcag gttcaccgcc agccaggaca ggggcaagaa catcgcctac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcaa ggccagctgc 300 gtgaggggca ggaccatcag cgagtactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 111 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 111 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgtgg ccagcggcta cgtgagctgc gactacttcc tgcccagctg gtacaggcag 120 gcccccggca aggagaggga gttcgtgagc atcatcgacg gcaccggcag caccagctac 180 gccgccagcg tgaagggcag gttcaccgcc agccaggaca agggcaagaa catcgcctac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcaa ggccagctgc 300 gtgaggggca gggccatcag cgagtactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 112 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 112 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgtgg ccagcggcta cgtgagctgc gactacttcc tgcccagctg gtacaggcag 120 gcccccggca aggagaggga gttcgtgagc atcatcgacg gcaccggcag caccagctac 180 gccgccagcg tgaagggcag gttcaccgcc agccaggaca agggcaagaa catcgcctac 240 ctgcagatga acaccctgaa gcccgaggac accgccatgt actactgcaa ggccagctgc 300 gtgaggggca gggccatcag cgagtactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 113 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 113 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaggg ccagcggcag cacctacagc aactactgcc tgggctggtt caggcagatc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc aggccaagaa caccgtgtac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgc cgccgagagc 300 gccagcagct tcagctgcgg cggctggctg accaggcccg acagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 114 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 114 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc ctgagcggca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcgcc atcagcagcg gcggcgccag cacctactac 180 accgacagcg tgaagggcag gttcaccatc agcagggaca acgccaagaa catgctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatgt actactgcgc caagggcggc 300 agcggctacg gcgacgccag caggatgacc agccccggca gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 115 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 115 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgtgg ccagcggcta cgtgagctgc gactacttcc tgcccagctg gtacaggcag 120 gcccccggca aggagaggga gttcgtgagc atcatcgacg gcaccggcag caccagctac 180 gccgccagcg tgaagggcag gttcaccgcc agccaggaca agggcaagaa catcgcctac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcaa ggccagctgc 300 gtgaggggca ggggcatcag cgagtactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 116 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 116 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaggg ccagcggcag cacctacagc aactactgcc tgggctggtt caggcagatc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc aggccaagaa caccgtgtac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgc cgccgagagc 300 gtgagcagct tcagctgcgg cggctggctg accaggcccg acagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 117 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 117 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc agctacccca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcacc atcagcagcg gcggcgacac caccctgtac 180 gccgacagcg tgaagggcag gttcaccagc agcagggaca acgccaagaa caccctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatgt actactgcgc caagaggatc 300 gactgcaaca gcggctactg ctacaagagg agctactggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 118 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 118 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc ctgagcagca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcgcc atcagcagcg gcggcgccag cacctactac 180 accgacagcg tgaagggcag gttcaccatc agcagggaca acgccaagaa catgctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatgt actactgcgc caagggcggc 300 agcggctacg gcgacgccag caggatgacc agccccggca gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 119 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 119 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaggg ccagcggcag cacctacagc aactactgcc tgggctggtt caggcagacc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc aggccaagaa caccgtgtac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgc cgccgagagc 300 gtgagcagct tcagctgcgg cggctggctg accaggcccg acagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 120 <211> 387 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 120 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaggg ccagcaggag cccctacggc aactactgcc tgggctggtt caggcagagc 120 accggcaagg agagggaggg cgtggccgtg atcaactggg tgggcggcat gctgtacttc 180 gccgacagcg tgaagggcag gttcaccgtg agccaggacc acgccaagaa caccgtgacc 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgc cgccgagagc 300 gtgagcagct tcagctgcgg cggctggctg accaggcccg acagggtgcc ctactggggc 360 cagggcaccc aggtgaccgt gagcagc 387 <210> 121 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 121 caggtgcagc tgcaggagag cggcggcggc ctggtgcagc ccggcggcag cctgaggctg 60 agctgcgccg ccagcggctt caccttcagc cacagcggca tgagctgggt gaggcaggcc 120 cccggcaagg gcctggagtg ggtgagcacc atcaacagcg gcggcgccag cacctactac 180 accgacagcg tgaagggcag gttcaccatc agcagggaca acgccaagaa catgctgtac 240 ctgcagctga acagcctgaa gaccgaggac accgccatgt actactgcgc caagggcggc 300 agcggctacg gcgacgccag caggatgacc agccccggca gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 122 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 122 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Lys Asn Ser Asn Phe Met Gly 20 25 30 Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val Ala Ala Met 35 40 45 Met Thr Lys Asn Asn Asn Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg 50 55 60 Phe Thr Ile Ser His Asp Asn Ala Lys Asn Thr Val Tyr Leu Gln Met 65 70 75 80 Asp Ser Leu Lys Pro Glu Asp Thr Ala Val Tyr Tyr Cys Ala Ala Val 85 90 95 Tyr Arg Thr Arg Arg Leu Arg Val Leu Glu Ala Ala Asn Phe Asp Tyr 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 123 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 123 Asn Ser Asn Phe Met Gly 1 5 <210> 124 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 124 Ala Met Met Thr Lys Asn Asn Asn Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 125 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 125 Val Tyr Arg Thr Arg Arg Leu Arg Val Leu Glu Ala Ala Asn Phe Asp 1 5 10 15 Tyr <210> 126 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 126 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Tyr Thr Ser Ser Arg Tyr 20 25 30 Cys Met Gly Trp Phe Arg Gln Thr Pro Gly Lys Lys Arg Glu Gly Val 35 40 45 Ala Ala Ile Tyr Thr Gly Gly Gly Thr Thr Phe Tyr His Gly Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Gln Asp Asn Thr Thr Asn Thr Val Tyr 65 70 75 80 Leu Gln Met His Asn Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Ala Gly Pro Val Thr Arg Ala Cys Asp Glu Tyr Asn Tyr Trp Gly 100 105 110 Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 127 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 127 Tyr Thr Ser Ser Arg Tyr Cys Met Gly 1 5 <210> 128 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 128 Ala Ile Tyr Thr Gly Gly Gly Thr Thr Phe Tyr His Gly Ser Val Lys 1 5 10 15 Gly <210> 129 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 129 Gly Pro Val Thr Arg Ala Cys Asp Glu Tyr Asn Tyr 1 5 10 <210> 130 <211> 126 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 130 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Tyr Ser Leu Ser Asn Tyr 20 25 30 Cys Met Gly Trp Phe Arg Gln Ala Pro Gly Gln Gly Arg Glu Gly Val 35 40 45 Ala Ser Leu Arg Phe Val Ser Gly Ala Thr Phe Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ala Gln Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Gly Ile Lys Ser Arg Gly Ile Cys Gly Gly Arg Leu Val Asp Val Asp 100 105 110 Phe Gly Asn Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 125 <210> 131 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 131 Tyr Ser Leu Ser Asn Tyr Cys Met Gly 1 5 <210> 132 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 132 Ser Leu Arg Phe Val Ser Gly Ala Thr Phe Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 133 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 133 Lys Ser Arg Gly Ile Cys Gly Gly Arg Leu Val Asp Val Asp Phe Gly 1 5 10 15 Asn <210> 134 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 134 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Ser Ile Asn Arg Met 20 25 30 Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Ser Ile Gly Gly Gly Gln Thr Tyr Tyr Ala Asp Ser Val Lys Gly 50 55 60 Arg Phe Thr Ile Ser Gln Asp Asn Ala Lys Asn Thr Val Asp Leu Gln 65 70 75 80 Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala 85 90 95 Gly Leu Val Tyr Gly Glu Ala Trp Leu Asp Ser Arg His Tyr Asn Lys 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 135 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 135 Tyr Ser Ile Asn Arg Met Gly 1 5 <210> 136 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 136 Ala Ile Ser Ile Gly Gly Gly Gln Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 137 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 137 Gly Leu Val Tyr Gly Glu Ala Trp Leu Asp Ser Arg His Tyr Asn Lys 1 5 10 15 <210> 138 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 138 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Ala Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg 50 55 60 Phe Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met 65 70 75 80 Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His 85 90 95 Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly 100 105 110 Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 139 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 139 Asp Ser Thr Tyr Ser Met Gly 1 5 <210> 140 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 140 Ala Ile Ala Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 141 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 141 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 142 <211> 122 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 142 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Thr Tyr Ser Ser Tyr 20 25 30 Cys Met Ala Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val 35 40 45 Ala Ala Ile Asp Ser Asp Gly Ser Thr Ser Tyr Ala Asp Ser Val Lys 50 55 60 Gly Arg Phe Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala 85 90 95 Ala Ala Ser Gly Arg Cys Leu Gly Pro Gly Ile Arg Ser Leu Ile Trp 100 105 110 Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 143 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 143 Tyr Thr Tyr Ser Ser Tyr Cys Met Ala 1 5 <210> 144 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 144 Ala Ile Asp Ser Asp Gly Ser Thr Ser Tyr Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 145 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 145 Ala Ser Gly Arg Cys Leu Gly Pro Gly Ile Arg Ser Leu Ile 1 5 10 <210> 146 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 146 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Thr Lys Asp Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg Phe 50 55 60 Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met Asn 65 70 75 80 Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His Arg 85 90 95 Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 147 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 147 Asp Ser Thr Tyr Ser Met Gly 1 5 <210> 148 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 148 Ala Ile Thr Lys Asp Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 149 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 149 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 150 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 150 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Pro Thr Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg 50 55 60 Phe Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met 65 70 75 80 Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His 85 90 95 Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly 100 105 110 Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 151 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 151 Asp Ser Thr Tyr Ser Met Gly 1 5 <210> 152 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 152 Ala Ile Pro Thr Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 153 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 153 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 154 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 154 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Ala Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg 50 55 60 Phe Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met 65 70 75 80 Ser Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His 85 90 95 Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly 100 105 110 Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 155 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 155 Asp Ser Thr Tyr Ser Met Gly 1 5 <210> 156 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 156 Ala Ile Ala Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 157 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 157 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 158 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 158 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Gly Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg 50 55 60 Phe Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met 65 70 75 80 Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His 85 90 95 Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly 100 105 110 Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 159 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 159 Asp Ser Thr Tyr Ser Met Gly 1 5 <210> 160 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 160 Ala Ile Gly Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 161 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 161 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 162 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 162 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Thr Lys Asp Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg Phe 50 55 60 Thr Ile Ser Lys Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met Asn 65 70 75 80 Ser Leu Arg Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His Arg 85 90 95 Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly Gln 100 105 110 Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 163 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 163 Asp Ser Thr Tyr Ser Met Gly 1 5 <210> 164 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 164 Ala Ile Thr Lys Asp Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 165 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 165 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 166 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 166 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Thr Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Ser Ile Asn Arg Met 20 25 30 Ala Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Ser Ile Gly Gly Asp Arg Thr Tyr Tyr Ala Asp Ser Val Lys Gly 50 55 60 Arg Phe Thr Ile Ser Gln Asp Asn Ala Lys His Thr Val Asp Leu Gln 65 70 75 80 Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala 85 90 95 Gly Leu Val Tyr Gly Glu Ala Trp Leu Asp Ser Arg His Tyr Asn Lys 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 167 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 167 Tyr Ser Ile Asn Arg Met Ala 1 5 <210> 168 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 168 Ala Ile Ser Ile Gly Gly Asp Arg Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 169 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 169 Gly Leu Val Tyr Gly Glu Ala Trp Leu Asp Ser Arg His Tyr Asn Lys 1 5 10 15 <210> 170 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 170 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Ser Ile Asn Arg Met 20 25 30 Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Ser Ile Gly Gly Gly Arg Thr Tyr Tyr Ala Asp Ser Val Lys Gly 50 55 60 Arg Phe Thr Ile Ser Gln Asp Asn Ala Lys Asn Thr Val Asp Leu Gln 65 70 75 80 Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala 85 90 95 Gly Leu Val Tyr Gly Glu Ala Trp Leu Asp Ser Arg His Tyr Asn Lys 100 105 110 Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 171 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 171 Tyr Ser Ile Asn Arg Met Gly 1 5 <210> 172 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 172 Ala Ile Ser Ile Gly Gly Gly Arg Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 173 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 173 Gly Leu Val Tyr Gly Glu Ala Trp Leu Asp Ser Arg His Tyr Asn Lys 1 5 10 15 <210> 174 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 174 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Val Ser Gly Asp Ser Thr Tyr Ser Met 20 25 30 Gly Trp Phe Arg Gln Pro Pro Gly Lys Glu Arg Glu Gly Val Ala Ala 35 40 45 Ile Thr Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly Arg 50 55 60 Phe Thr Ile Ser Gly Asp Asn Ala Lys Asn Thr Leu Tyr Leu Gln Met 65 70 75 80 Asn Asn Leu Lys Pro Glu Asp Thr Ala Met Tyr Tyr Cys Ala Ala His 85 90 95 Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr Trp Gly 100 105 110 Gln Gly Thr Gln Val Thr Val Ser Ser 115 120 <210> 175 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 175 Asp Ser Thr Tyr Ser Met 1 5 <210> 176 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 176 Ala Ile Thr Lys Asp Gly Ile Thr Ile His Ala Asp Ser Val Lys Gly 1 5 10 15 <210> 177 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 177 His Arg Pro Tyr Gly Pro Pro Leu Asn Pro Arg Trp Tyr Thr Tyr 1 5 10 15 <210> 178 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 178 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg ccagcaagaa cagcaacttc atgggctggt tcaggcaggc ccccggcaag 120 gagagggagg gcgtggccgc catgatgacc aagaacaaca acacctacta cgccgacagc 180 gtgaagggca ggttcaccat cagccacgac aacgccaaga acaccgtgta cctgcagatg 240 gacagcctga agcccgagga caccgccgtg tactactgcg ccgccgtgta caggaccagg 300 aggctgaggg tgctggaggc cgccaacttc gactactggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 179 <211> 363 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 179 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcaccg ccagcggcta caccagcagc aggtactgca tgggctggtt caggcagacc 120 cccggcaaga agagggaggg cgtggccgcc atctacaccg gcggcggcac caccttctac 180 cacggcagcg tgaagggcag gttcaccatc agccaggaca acaccaccaa caccgtgtac 240 ctgcagatgc acaacctgaa gcccgaggac accgccatgt actactgcgc cgccggcccc 300 gtgaccaggg cctgcgacga gtacaactac tggggccagg gcacccaggt gaccgtgagc 360 agc 363 <210> 180 <211> 378 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 180 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg gcagcggcta cagcctgagc aactactgca tgggctggtt caggcaggcc 120 cccggccagg gcagggaggg cgtggccagc ctgaggttcg tgagcggcgc caccttctac 180 gccgacagcg tgaagggcag gttcaccatc gcccaggaca acgccaagaa caccctgtac 240 ctgcagatga acagcctgaa gcccgaggac accgccatgt actactgcgg catcaagagc 300 aggggcatct gcggcggcag gctggtggac gtggacttcg gcaactgggg ccagggcacc 360 caggtgaccg tgagcagc 378 <210> 181 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 181 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg ccagcggcta cagcatcaac aggatgggct ggttcaggca ggcccccggc 120 aaggagaggg agggcgtggc cgccatcagc atcggcggcg gccagaccta ctacgccgac 180 agcgtgaagg gcaggttcac catcagccag gacaacgcca agaacaccgt ggacctgcag 240 atgaacagcc tgaagcccga ggacaccgcc atgtactact gcgccgccgg cctggtgtac 300 ggcgaggcct ggctggacag caggcactac aacaagtggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 182 <211> 363 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 182 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcgcc aaggacggca tcaccatcca cgccgacagc 180 gtgaagggca ggttcaccat cagcaaggac aacgccaaga acaccctgta cctgcagatg 240 aacagcctga agcccgagga caccgccatg tactactgcg ccgcccacag gccctacggc 300 ccccccctga accccaggtg gtacacctac tggggccagg gcacccaggt gaccgtgagc 360 agc 363 <210> 183 <211> 366 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 183 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg ccagcggcta cacctacagc agctactgca tggcctggtt caggcaggcc 120 cccggcaagg agagggaggg cgtggccgcc atcgacagcg acggcagcac cagctacgcc 180 gacagcgtga agggcaggtt caccatcagc aaggacaacg ccaagaacac cctgtacctg 240 cagatgaaca gcctgaagcc cgaggacacc gccatgtact actgcgccgc cgccagcggc 300 aggtgcctgg gccccggcat caggagcctg atctggggcc agggcaccca ggtgaccgtg 360 agcagc 366 <210> 184 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 184 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcacc aaggacatca ccatccacgc cgacagcgtg 180 aagggcaggt tcaccatcag caaggacaac gccaagaaca ccctgtacct gcagatgaac 240 agcctgaagc ccgaggacac cgccatgtac tactgcgccg cccacaggcc ctacggcccc 300 cccctgaacc ccaggtggta cacctactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 185 <211> 363 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 185 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcccc accgacggca tcaccatcca cgccgacagc 180 gtgaagggca ggttcaccat cagcaaggac aacgccaaga acaccctgta cctgcagatg 240 aacagcctga agcccgagga caccgccatg tactactgcg ccgcccacag gccctacggc 300 ccccccctga accccaggtg gtacacctac tggggccagg gcacccaggt gaccgtgagc 360 agc 363 <210> 186 <211> 363 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 186 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcgcc aaggacggca tcaccatcca cgccgacagc 180 gtgaagggca ggttcaccat cagcaaggac aacgccaaga acaccctgta cctgcagatg 240 agcagcctga agcccgagga caccgccatg tactactgcg ccgcccacag gccctacggc 300 ccccccctga accccaggtg gtacacctac tggggccagg gcacccaggt gaccgtgagc 360 agc 363 <210> 187 <211> 363 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 187 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcggc aaggacggca tcaccatcca cgccgacagc 180 gtgaagggca ggttcaccat cagcaaggac aacgccaaga acaccctgta cctgcagatg 240 aacagcctga agcccgagga caccgccatg tactactgcg ccgcccacag gccctacggc 300 ccccccctga accccaggtg gtacacctac tggggccagg gcacccaggt gaccgtgagc 360 agc 363 <210> 188 <211> 360 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 188 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcacc aaggacatca ccatccacgc cgacagcgtg 180 aagggcaggt tcaccatcag caaggacaac gccaagaaca ccctgtacct gcagatgaac 240 agcctgaggc ccgaggacac cgccatgtac tactgcgccg cccacaggcc ctacggcccc 300 cccctgaacc ccaggtggta cacctactgg ggccagggca cccaggtgac cgtgagcagc 360 <210> 189 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 189 caggtgcagc tgcaggagag cggcggcggc agcgtgcaga ccggcggcag cctgaggctg 60 agctgcgccg ccagcggcta cagcatcaac aggatggcct ggttcaggca ggcccccggc 120 aaggagaggg agggcgtggc cgccatcagc atcggcggcg acaggaccta ctacgccgac 180 agcgtgaagg gcaggttcac catcagccag gacaacgcca agcacaccgt ggacctgcag 240 atgaacagcc tgaagcccga ggacaccgcc atgtactact gcgccgccgg cctggtgtac 300 ggcgaggcct ggctggacag caggcactac aacaagtggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 190 <211> 369 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 190 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg ccagcggcta cagcatcaac aggatgggct ggttcaggca ggcccccggc 120 aaggagaggg agggcgtggc cgccatcagc atcggcggcg gcaggaccta ctacgccgac 180 agcgtgaagg gcaggttcac catcagccag gacaacgcca agaacaccgt ggacctgcag 240 atgaacagcc tgaagcccga ggacaccgcc atgtactact gcgccgccgg cctggtgtac 300 ggcgaggcct ggctggacag caggcactac aacaagtggg gccagggcac ccaggtgacc 360 gtgagcagc 369 <210> 191 <211> 363 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 191 caggtgcagc tgcaggagag cggcggcggc agcgtgcagg ccggcggcag cctgaggctg 60 agctgcgccg tgagcggcga cagcacctac agcatgggct ggttcaggca gccccccggc 120 aaggagaggg agggcgtggc cgccatcacc aaggacggca tcaccatcca cgccgacagc 180 gtgaagggca ggttcaccat cagcggcgac aacgccaaga acaccctgta cctgcagatg 240 aacaacctga agcccgagga caccgccatg tactactgcg ccgcccacag gccctacggc 300 ccccccctga accccaggtg gtacacctac tggggccagg gcacccaggt gaccgtgagc 360 agc 363 <210> 192 <211> 484 <212> PRT <213> Homo sapiens <400> 192 Gln Gly Ser Ala Gly Pro Leu Gln Cys Tyr Gly Val Gly Pro Leu Gly 1 5 10 15 Asp Leu Asn Cys Ser Trp Glu Pro Leu Gly Asp Leu Gly Ala Pro Ser 20 25 30 Glu Leu His Leu Gln Ser Gln Lys Tyr Arg Ser Asn Lys Thr Gln Thr 35 40 45 Val Ala Val Ala Ala Gly Arg Ser Trp Val Ala Ile Pro Arg Glu Gln 50 55 60 Leu Thr Met Ser Asp Lys Leu Leu Val Trp Gly Thr Lys Ala Gly Gln 65 70 75 80 Pro Leu Trp Pro Pro Val Phe Val Asn Leu Glu Thr Gln Met Lys Pro 85 90 95 Asn Ala Pro Arg Leu Gly Pro Asp Val Asp Phe Ser Glu Asp Asp Pro 100 105 110 Leu Glu Ala Thr Val His Trp Ala Pro Pro Thr Trp Pro Ser His Lys 115 120 125 Val Leu Ile Cys Gln Phe His Tyr Arg Arg Cys Gln Glu Ala Ala Trp 130 135 140 Thr Leu Leu Glu Pro Glu Leu Lys Thr Ile Pro Leu Thr Pro Val Glu 145 150 155 160 Ile Gln Asp Leu Glu Leu Ala Thr Gly Tyr Lys Val Tyr Gly Arg Cys 165 170 175 Arg Met Glu Lys Glu Glu Asp Leu Trp Gly Glu Trp Ser Pro Ile Leu 180 185 190 Ser Phe Gln Thr Pro Pro Ser Ala Pro Lys Asp Val Trp Val Ser Gly 195 200 205 Asn Leu Cys Gly Thr Pro Gly Gly Glu Glu Pro Leu Leu Leu Trp Lys 210 215 220 Ala Pro Gly Pro Cys Val Gln Val Ser Tyr Lys Val Trp Phe Trp Val 225 230 235 240 Gly Gly Arg Glu Leu Ser Pro Glu Gly Ile Thr Cys Cys Cys Ser Leu 245 250 255 Ile Pro Ser Gly Ala Glu Trp Ala Arg Val Ser Ala Val Asn Ala Thr 260 265 270 Ser Trp Glu Pro Leu Thr Asn Leu Ser Leu Val Cys Leu Asp Ser Ala 275 280 285 Ser Ala Pro Arg Ser Val Ala Val Ser Ser Ile Ala Gly Ser Thr Glu 290 295 300 Leu Leu Val Thr Trp Gln Pro Gly Pro Gly Glu Pro Leu Glu His Val 305 310 315 320 Val Asp Trp Ala Arg Asp Gly Asp Pro Leu Glu Lys Leu Asn Trp Val 325 330 335 Arg Leu Pro Pro Gly Asn Leu Ser Ala Leu Leu Pro Gly Asn Phe Thr 340 345 350 Val Gly Val Pro Tyr Arg Ile Thr Val Thr Ala Val Ser Ala Ser Gly 355 360 365 Leu Ala Ser Ala Ser Ser Val Trp Gly Phe Arg Glu Glu Leu Ala Pro 370 375 380 Leu Val Gly Pro Thr Leu Trp Arg Leu Gln Asp Ala Pro Pro Gly Thr 385 390 395 400 Pro Ala Ile Ala Trp Gly Glu Val Pro Arg His Gln Leu Arg Gly His 405 410 415 Leu Thr His Tyr Thr Leu Cys Ala Gln Ser Gly Thr Ser Pro Ser Val 420 425 430 Cys Met Asn Val Ser Gly Asn Thr Gln Ser Val Thr Leu Pro Asp Leu 435 440 445 Pro Trp Gly Pro Cys Glu Leu Trp Val Thr Ala Ser Thr Ile Ala Gly 450 455 460 Gln Gly Pro Pro Gly Pro Ile Leu Arg Leu His Leu Pro Asp Asn Thr 465 470 475 480 Leu Arg Trp Lys <210> 193 <211> 623 <212> PRT <213> Mus sp. <400> 193 Met Asn Arg Leu Arg Val Ala Arg Leu Thr Pro Leu Glu Leu Leu Leu 1 5 10 15 Ser Leu Met Ser Leu Leu Leu Gly Thr Arg Pro His Gly Ser Pro Gly 20 25 30 Pro Leu Gln Cys Tyr Ser Val Gly Pro Leu Gly Ile Leu Asn Cys Ser 35 40 45 Trp Glu Pro Leu Gly Asp Leu Glu Thr Pro Pro Val Leu Tyr His Gln 50 55 60 Ser Gln Lys Tyr His Pro Asn Arg Val Trp Glu Val Lys Val Pro Ser 65 70 75 80 Lys Gln Ser Trp Val Thr Ile Pro Arg Glu Gln Phe Thr Met Ala Asp 85 90 95 Lys Leu Leu Ile Trp Gly Thr Gln Lys Gly Arg Pro Leu Trp Ser Ser 100 105 110 Val Ser Val Asn Leu Glu Thr Gln Met Lys Pro Asp Thr Pro Gln Ile 115 120 125 Phe Ser Gln Val Asp Ile Ser Glu Glu Ala Thr Leu Glu Ala Thr Val 130 135 140 Gln Trp Ala Pro Pro Val Trp Pro Pro Gln Lys Val Leu Ile Cys Gln 145 150 155 160 Phe Arg Tyr Lys Glu Cys Gln Ala Glu Thr Trp Thr Arg Leu Glu Pro 165 170 175 Gln Leu Lys Thr Asp Gly Leu Thr Pro Val Glu Met Gln Asn Leu Glu 180 185 190 Pro Gly Thr Cys Tyr Gln Val Ser Gly Arg Cys Gln Val Glu Asn Gly 195 200 205 Tyr Pro Trp Gly Glu Trp Ser Ser Pro Leu Ser Phe Gln Thr Pro Phe 210 215 220 Leu Asp Pro Glu Asp Val Trp Val Ser Gly Thr Val Cys Glu Thr Ser 225 230 235 240 Gly Lys Arg Ala Ala Leu Leu Val Trp Lys Asp Pro Arg Pro Cys Val 245 250 255 Gln Val Thr Tyr Thr Val Trp Phe Gly Ala Gly Asp Ile Thr Thr Thr 260 265 270 Gln Glu Glu Val Pro Cys Cys Lys Ser Pro Val Pro Ala Trp Met Glu 275 280 285 Trp Ala Val Val Ser Pro Gly Asn Ser Thr Ser Trp Val Pro Pro Thr 290 295 300 Asn Leu Ser Leu Val Cys Leu Ala Pro Glu Ser Ala Pro Cys Asp Val 305 310 315 320 Gly Val Ser Ser Ala Asp Gly Ser Pro Gly Ile Lys Val Thr Trp Lys 325 330 335 Gln Gly Thr Arg Lys Pro Leu Glu Tyr Val Val Asp Trp Ala Gln Asp 340 345 350 Gly Asp Ser Leu Asp Lys Leu Asn Trp Thr Arg Leu Pro Pro Gly Asn 355 360 365 Leu Ser Thr Leu Leu Pro Gly Glu Phe Lys Gly Gly Val Pro Tyr Arg 370 375 380 Ile Thr Val Thr Ala Val Tyr Ser Gly Gly Leu Ala Ala Ala Pro Ser 385 390 395 400 Val Trp Gly Phe Arg Glu Glu Leu Val Pro Leu Ala Gly Pro Ala Val 405 410 415 Trp Arg Leu Pro Asp Asp Pro Pro Gly Thr Pro Val Val Ala Trp Gly 420 425 430 Glu Val Pro Arg His Gln Leu Arg Gly Gln Ala Thr His Tyr Thr Phe 435 440 445 Cys Ile Gln Ser Arg Gly Leu Ser Thr Val Cys Arg Asn Val Ser Ser 450 455 460 Gln Thr Gln Thr Ala Thr Leu Pro Asn Leu His Leu Gly Ser Phe Lys 465 470 475 480 Leu Trp Val Thr Val Ser Thr Val Ala Gly Gln Gly Pro Pro Gly Pro 485 490 495 Asn Leu Ser Leu His Leu Pro Asp Asn Arg Ile Arg Trp Lys Ala Leu 500 505 510 Pro Trp Phe Leu Ser Leu Trp Gly Leu Leu Leu Met Gly Cys Gly Leu 515 520 525 Ser Leu Ala Ser Thr Arg Cys Leu Gln Ala Arg Cys Leu His Trp Arg 530 535 540 His Lys Leu Leu Pro Gln Trp Ile Trp Glu Arg Val Pro Asp Pro Ala 545 550 555 560 Asn Ser Asn Ser Gly Gln Pro Tyr Ile Lys Glu Val Ser Leu Pro Gln 565 570 575 Pro Pro Lys Asp Gly Pro Ile Leu Glu Val Glu Glu Val Glu Leu Gln 580 585 590 Pro Val Val Glu Ser Pro Lys Ala Ser Ala Pro Ile Tyr Ser Gly Tyr 595 600 605 Glu Lys His Phe Leu Pro Thr Pro Glu Glu Leu Gly Leu Leu Val 610 615 620 <210> 194 <211> 486 <212> PRT <213> Mus sp. <400> 194 Thr Arg Pro His Gly Ser Pro Gly Pro Leu Gln Cys Tyr Ser Val Gly 1 5 10 15 Pro Leu Gly Ile Leu Asn Cys Ser Trp Glu Pro Leu Gly Asp Leu Glu 20 25 30 Thr Pro Pro Val Leu Tyr His Gln Ser Gln Lys Tyr His Pro Asn Arg 35 40 45 Val Trp Glu Val Lys Val Pro Ser Lys Gln Ser Trp Val Thr Ile Pro 50 55 60 Arg Glu Gln Phe Thr Met Ala Asp Lys Leu Leu Ile Trp Gly Thr Gln 65 70 75 80 Lys Gly Arg Pro Leu Trp Ser Ser Val Ser Val Asn Leu Glu Thr Gln 85 90 95 Met Lys Pro Asp Thr Pro Gln Ile Phe Ser Gln Val Asp Ile Ser Glu 100 105 110 Glu Ala Thr Leu Glu Ala Thr Val Gln Trp Ala Pro Pro Val Trp Pro 115 120 125 Pro Gln Lys Val Leu Ile Cys Gln Phe Arg Tyr Lys Glu Cys Gln Ala 130 135 140 Glu Thr Trp Thr Arg Leu Glu Pro Gln Leu Lys Thr Asp Gly Leu Thr 145 150 155 160 Pro Val Glu Met Gln Asn Leu Glu Pro Gly Thr Cys Tyr Gln Val Ser 165 170 175 Gly Arg Cys Gln Val Glu Asn Gly Tyr Pro Trp Gly Glu Trp Ser Ser 180 185 190 Pro Leu Ser Phe Gln Thr Pro Phe Leu Asp Pro Glu Asp Val Trp Val 195 200 205 Ser Gly Thr Val Cys Glu Thr Ser Gly Lys Arg Ala Ala Leu Leu Val 210 215 220 Trp Lys Asp Pro Arg Pro Cys Val Gln Val Thr Tyr Thr Val Trp Phe 225 230 235 240 Gly Ala Gly Asp Ile Thr Thr Thr Gln Glu Glu Val Pro Cys Cys Lys 245 250 255 Ser Pro Val Pro Ala Trp Met Glu Trp Ala Val Val Ser Pro Gly Asn 260 265 270 Ser Thr Ser Trp Val Pro Pro Thr Asn Leu Ser Leu Val Cys Leu Ala 275 280 285 Pro Glu Ser Ala Pro Cys Asp Val Gly Val Ser Ser Ala Asp Gly Ser 290 295 300 Pro Gly Ile Lys Val Thr Trp Lys Gln Gly Thr Arg Lys Pro Leu Glu 305 310 315 320 Tyr Val Val Asp Trp Ala Gln Asp Gly Asp Ser Leu Asp Lys Leu Asn 325 330 335 Trp Thr Arg Leu Pro Pro Gly Asn Leu Ser Thr Leu Leu Pro Gly Glu 340 345 350 Phe Lys Gly Gly Val Pro Tyr Arg Ile Thr Val Thr Ala Val Tyr Ser 355 360 365 Gly Gly Leu Ala Ala Ala Pro Ser Val Trp Gly Phe Arg Glu Glu Leu 370 375 380 Val Pro Leu Ala Gly Pro Ala Val Trp Arg Leu Pro Asp Asp Pro Pro 385 390 395 400 Gly Thr Pro Val Val Ala Trp Gly Glu Val Pro Arg His Gln Leu Arg 405 410 415 Gly Gln Ala Thr His Tyr Thr Phe Cys Ile Gln Ser Arg Gly Leu Ser 420 425 430 Thr Val Cys Arg Asn Val Ser Ser Gln Thr Gln Thr Ala Thr Leu Pro 435 440 445 Asn Leu His Leu Gly Ser Phe Lys Leu Trp Val Thr Val Ser Thr Val 450 455 460 Ala Gly Gln Gly Pro Pro Gly Pro Asn Leu Ser Leu His Leu Pro Asp 465 470 475 480 Asn Arg Ile Arg Trp Lys 485 <210> 195 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic 6xHis tag <400> 195 His His His His His His 1 5 <210> 196 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic 8xHis tag <400> 196 His His His His His His His His 1 5 <210> 197 <211> 40 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <220> <221> SITE <222> (1)..(40) <223> This sequence may encompass 1-10 "Gly Gly Gly Ser" repeating units <400> 197 Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser 1 5 10 15 Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser 20 25 30 Gly Gly Gly Ser Gly Gly Gly Ser 35 40 <210> 198 <211> 50 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <220> <221> SITE <222> (1)..(50) <223> This sequence may encompass 1-10 "Gly Gly Gly Ser Gly" repeating units <400> 198 Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly 1 5 10 15 Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly 20 25 30 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 35 40 45 Ser Gly 50 <210> 199 <211> 50 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <220> <221> SITE <222> (1)..(50) <223> This sequence may encompass 1-10 "Gly Gly Gly Gly Ser" repeating units <400> 199 Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly 1 5 10 15 Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly 20 25 30 Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly 35 40 45 Gly Ser 50 <210> 200 <211> 31 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <220> <221> SITE <222> (1)..(30) <223> This sequence may encompass 1-10 "Gly Gly Ser" repeating units <400> 200 Gly Gly Ser Gly Gly Ser Gly Gly Ser Gly Gly Ser Gly Gly Ser Gly 1 5 10 15 Gly Ser Gly Gly Ser Gly Gly Ser Gly Gly Ser Gly Gly Ser Gly 20 25 30 <210> 201 <211> 40 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <220> <221> SITE <222> (1)..(40) <223> This sequence may encompass 1-10 "Gly Gly Ser Gly" repeating units <400> 201 Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly 1 5 10 15 Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly 20 25 30 Gly Gly Ser Gly Gly Gly Ser Gly 35 40 <210> 202 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <220> <221> SITE <222> (1)..(6) <223> This sequence may encompass 3-6 residues <400> 202 His His His His His His 1 5

Claims

1. An IL27Rα-binding molecule that specifically binds to the extracellular domain of IL2Rb.

2. 2. The IL27Rα-binding molecule of claim 1, wherein the IL2Rb-binding molecule comprises a single domain antibody (sdAb).

3. The sdAb is shown in the following table: The IL27Rα binding molecule of claim 2, comprising complementarity determining region 1 (CDR1), CDR2, and CDR3 as shown in the row.

4. 4. The IL27Rα binding molecule of claim 2 or 3, wherein the sdAb has at least 80%, alternatively at least 85%, alternatively at least 90%, alternatively at least 95%, alternatively at least 98%, alternatively at least 99%, or alternatively 100% identity to the polypeptide sequence of any one of SEQ ID NOs:2-25.

5. The sdAb is shown in the following table: The IL27Rα binding molecule of claim 2, comprising complementarity determining region 1 (CDR1), CDR2, and CDR3 as shown in the row.

6. 3. The IL27Rα binding molecule of claim 2, wherein the sdAb has at least 80%, alternatively at least 85%, alternatively at least 90%, alternatively at least 95%, alternatively at least 98%, alternatively at least 99%, or alternatively 100% identity to the polypeptide sequence of any one of SEQ ID NOs: 122, 126, 130, 134, 138, 142, 146, 150, 154, 158, 162, 166, 170, and 174.

7. 6. The IL27Rα binding molecule of claim 3 or 5, wherein the sdAb is humanized or otherwise comprises CDRs grafted onto a heterologous framework.

8. The IL27Rα binding molecule of any one of claims 1 to 7, further comprising a labeling agent, an imaging agent, and / or a therapeutic agent.

9. 10. The IL27Rα binding molecule of any one of claims 1 to 8 for use in the isolation, depletion or enrichment of IL27Rα+ cells from a biological sample.

11. A nucleic acid sequence encoding an IL27Rα binding molecule according to any one of claims 1 to 8.

12. A recombinant viral or non-viral vector comprising the nucleic acid of claim 11.

13. A host cell comprising the nucleic acid of claim 11.

14. A kit comprising an IL2Rb binding molecule according to any one of claims 1 to 8.

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