Antibodies targeting integrin beta-2

Abstract

Antibodies that specifically target integrin beta-2, as well as compositions containing such antibodies for therapeutic and diagnostic uses, are provided herein. The antibody comprises a heavy chain variable region comprising an HCDR1 sequence containing ISYYYM, an HCDR2 sequence containing SISSSSGYTY; and an HCDR3 sequence containing GAM; and a light chain variable region comprising an LCDR1 sequence containing SVSSA, an LCDR2 sequence containing SASSLYS; and an LCDR3 sequence containing FSSGSWAPI, comprising an integrin beta-2 binding domain.

Description

Technical Field

[0001] The embodiments disclosed herein relate to antibody therapeutics, particularly antibodies that specifically target integrin beta-2.

Background Art

[0002] It has proven difficult to develop safe and effective immunotherapies due to the lack of characterized cancer-specific surface markers. Given the abnormalities in tumor signaling, metabolism, or cell-microenvironment communication (all of which are highly involved in membrane proteins), it is hypothesized that cancer-specific surface protein conformations can actually span a wide range. In particular, integrin beta-2 may be a promising immunotherapy target that is widely expressed across cell lines and patient tumors.

[0003] Therefore, there is a need for novel antibodies that specifically target integrin beta-2 for the further development of targeted cancer immunotherapies and diagnostic methods.

Summary of the Invention

[0004] Using phage display selection, anti-integrin beta-2 antibodies that can be used for diagnostic and therapeutic purposes were identified.

[0005] In some embodiments, anti-integrin beta-2 antibodies having a KD of less than about 10 nM are provided.

[0006] In some embodiments, the antibody comprises an anti-integrin beta-2 binding domain having at least 1, at least 2, or 3 CDRs of the variable domain sequences of SEQ ID NO: 2 or SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR3 of SEQ ID NO: 2 and LCDR3 of SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2 and LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3.

[0007] In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 2. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 2. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 2.

[0008] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 3.

[0009] In some embodiments, the anti-integrin beta-2 binding domain comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region array of SEQ ID NO: 2 or SEQ ID NO: 3. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 2 or SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 2 and has a heavy chain variable region having at least 95% identity to SEQ ID NO: 2; and the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 3 and has a light chain variable region having at least 95% identity to SEQ ID NO: 3.

[0010] Other aspects and features will become apparent to those skilled in the art upon review of the following description of some exemplary embodiments.

[0011] The drawings included herein are for the purpose of illustrating various examples of the articles, methods, and apparatuses herein. In the figures, as follows.

Brief Description of the Drawings

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DETAILED DESCRIPTION OF THE INVENTION

[0018] To provide examples of each claimed embodiment, various compositions of matter are described below. None of the embodiments described below limit the claimed embodiments, and the claimed embodiments may encompass compositions different from those described below. The claimed embodiments are not limited to compositions having all the features of any one of the compositions described below, or to features common to a plurality or all of the compositions described below.

[0019] The term "about," as used herein, refers to the normal error range of each value readily known to those of ordinary skill in the art in this technical field. For example, for KD and IC50 values, ±20%, ±10%, or ±5% is within the intended meaning of the recited values.

[0020] The term integrin beta-2, also known as CD18, LAD, LCAMB, LFA-1, MAC-1, MF17, MFI7 or integrin subunit beta2, as used herein, refers to a polypeptide that is cytogenetically located on human chromosome 21q22.3 by the ITGB2 gene (chr21:44,885,949-44,931,989 (GRCH38 / hg38), HGNC, Entrez Gene, and Ensembl (genomic coordinates (GRCh38 / hg38 assembly December 2013:)) and plays a role in cell adhesion, cell surface-mediated sequencing, and immune response. The exemplary human integrin beta-2 protein sequence encoded by the human ITGB2 gene, P05107-1, is available under Uniprot number P05107 and is provided as SEQ ID NO:1. Integrin beta-2 can bind to several alpha chains and can thus be derived from multiple heterodimers, but also exists in a soluble ligand-binding form. Deficiency in Itgb2 expression results in adhesion defects in human circulating leukocytes and may reduce the ability of the immune system to fight foreign invaders. Exemplary integrin beta-2 heterodimers include, for example, integrin ITGAL / ITGB2, which is a receptor for ICAM1, ICAM2, ICAM3 and ICAM4 and also a receptor for the secreted form of the ubiquitin-like protein ISG15; integrin ITGAM / ITGB2 and ITGAX / ITGB2, which are receptors for the iC3b fragment of the third complement component and fibrinogen; integrin ITGAX / ITGB2, which recognizes the sequence G-P-R in the fibrinogen alpha chain, integrin ITGAM / ITGB2, which recognizes the P1 and P2 peptides of the fibrinogen gamma chain and is also a receptor for factor X; and integrin ITGAD / ITGB2, which is a receptor for ICAM3 and VCAM1.

[0021] The terms "anti-integrin beta-2 antibody", "integrin beta-2 specific antibody", "integrin beta-2 antibody", and "anti-integrin beta-2" are used interchangeably herein to refer to an antibody that specifically binds to integrin beta-2. An exemplary human integrin beta-2 sequence is provided in SEQ ID NO: 1.

[0022] "Anti-integrin beta-2 binding domain", as used herein, refers to the antigen-binding domain of the anti-integrin beta-2 antibody described herein that contains the V H region and the V L region, and this antigen-binding domain binds to integrin beta-2.

[0023] The anti-integrin beta-2 antibodies of the present disclosure bind to integrin beta-2. The active state of integrin beta-2 is an extended-open conformation (see, e.g., Nishida et al, Immunity 25:583-94, 2006; Li et al, EMBO J. 36:629-45, 2017). The active conformation (extended-open) has a 4,000-fold increased ligand affinity compared to the other two states (bent-closed, inactive; and extended-closed (intermediate)) (Li et al, 2017, supra). Integrin activation occurs upon cell stimulation via various cell surface receptors. Cell stimulation induces an inverted signaling pathway that ultimately recruits cytoplasmic factors such as talin and kindlin to the NPxY motif in the cytoplasmic tail of the beta chain of integrin, thereby separating the cytoplasmic tails of the integrin subunits and switching the integrin to the active (extended-open) conformation.

[0024] As used herein, the term "antibody" refers to a polypeptide that specifically binds to and recognizes an antigen, e.g., integrin beta-2, and that contains a framework region encoded by an immunoglobulin gene or a fragment thereof. Typically, the "variable region" contains the antigen-binding region of the antibody (or a functional equivalent thereof) and is important in binding specificity and affinity. Accordingly, the term "antibody" as used herein includes antigen-binding fragments, e.g., antigen-binding domains, or other antigen-binding fragments. Antigen-binding fragments may be produced by modification of a whole antibody or may be produced using recombinant DNA methods (e.g., single-chain Fv format).

[0025] Exemplary immunoglobulin (antibody) structural units include tetramers. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having one "light" chain (about 25 kD) and one "heavy" chain (about 50 - 70 kD). The N-terminus of each chain defines a variable region of about 100 - 110 or more amino acids that is primarily responsible for antigen recognition. The variable light chain (V L ) and variable heavy chain (V H ) terms refer to these light and heavy chains, respectively.

[0026] As used herein, the term "V region" refers to an antibody, e.g., an antibody variable domain, that includes segments of framework 1, CDR1, framework 2, CDR2, and framework 3 that include CDR3 and framework 4, and these segments are added to the V segments as a result of rearrangement of V region genes during B cell differentiation.

[0027] As used herein, the term "complementary determining region (CDR)" refers to three hypervariable regions flanked by four "framework" regions of the variable domain. CDRs are the major contributing factors to binding to an epitope of an antigen. CDRs are called CDR1, CDR2, and CDR3 and are numbered in order starting from the N-terminus.

[0028] The amino acid sequences of the CDR and framework regions can be determined using various definitions well-known in the art, such as Kabat, Chothia, the international ImMunoGeneTics database (IMGT), and AbM (e.g., Johnson et al., supra; Chothia & Lesk, 1987, Canonical structures for the hypervariable regions of immunoglobulins. J. Mol. Biol. 196, 901-917; Chothia C. et al., 1989, Conformations of immunoglobulin hypervariable regions. Nature 342, 877-883; Chothia C. et al., 1992, structural repertoire of the human VH segments J. Mol. Biol. 227, 799-817; Al-Lazikani et al., J. Mol. Biol 1997, 273(4)).The definition of the antigen-binding site is also described below: Ruiz et al., IMGT, the international ImMunoGeneTics database. Nucleic Acids Res., 28, 219-221 (2000); and Lefranc, M.-P. IMGT, the international ImMunoGeneTics database. Nucleic Acids Res. Jan 1; 29(1): 207-9 (2001); MacCallum et al, Antibody-antigen interactions: Contact analysis and binding site topography, J. Mol. Biol., 262(5), 732-745 (1996); and Martin et al, Proc. Natl Acad. Sci. USA, 86, 9268-9272 (1989); Martin, et al, Methods Enzymol., 203, 121-153, (1991); Pedersen et al, Immunomethods, 1, 126, (1992); and Rees et al, In Sternberg M.J.E. (ed.), Protein Structure Prediction. Oxford University Press, Oxford, 141-172 1996). References to CDRs determined by Kabat numbering are based on, for example, Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institute of Health, Bethesda, MD (1991)). CDRs according to Chothia are determined as defined by Chothia (see, for example, Chothia and Lesk J. Mol. Biol. 196: 901-917 (1987)).

[0029] "Isotype", as used herein, is a classification of antibodies defined by the heavy chain constant region. The antibodies described herein can be of any isotype of the isotype classification. Immunoglobulin genes include kappa, lambda, alpha, gamma, delta, epsilon, and mu constant region genes. Light chains are classified into either kappa or lambda. Heavy chains are classified as gamma, mu, alpha, delta, or epsilon, which in turn define the isotype classifications IgG, IgM, IgA, IgD, and IgE, respectively. In some embodiments, IgG is IgG1, IgG2, IgG3, or IgG4.

[0030] Antibodies can exist as intact immunoglobulins or as any of several well-characterized fragments that include specific antigen-binding activity. Such fragments can be generated by digestion with various peptidases. Pepsin digests the antibody under the disulfide linkage of the hinge region to produce F(ab)'2, which is a dimer of Fab with the light chain joined to V H -C H 1 by a disulfide bond. F(ab)'2 can be reduced under mild conditions to break the disulfide linkage in the hinge region, thereby converting the F(ab)'2 dimer to Fab' monomers. Fab' monomers essentially have a portion of the hinge region (see Fundamental Immunology (Paul ed., 3d ed. 1993)). Although various antibody fragments are defined with respect to the digestion of intact antibodies, one of ordinary skill in the art will understand that such fragments can be synthesized de novo by chemical or recombinant DNA methodologies.

[0031] The antibody or antigen-binding molecule of the present invention is further included one or more immunoglobulin chains which are chemically conjugated to a fusion protein with another protein or expressed as a fusion protein. Bispecific antibodies are also included. A bispecific antibody or bifunctional antibody is an artificial hybrid antibody having two different heavy chain / light chain pairs and two different binding sites. Other antigen-binding fragments or antibody portions of the present invention include single-chain variable fragments (scFv), bivalent scFv (diabody), bispecific scFv antibodies in which the antibody molecule recognizes two different epitopes, single binding domains (dAb), and minibodies. The term "antibody" further encompasses bispecific and multispecific antibodies, as well as any other monovalent, bivalent or multivalent antibody format.

[0032] The various antibodies or antigen-binding fragments described herein may be generated by enzymatic or chemical modification of intact antibodies, or may be synthesized de novo using recombinant DNA methodologies (e.g., single-chain Fv), or may be identified using yeast or phage display libraries (see, e.g., McCafferty et al., Nature 348:552-554, 1990; Boder, et al (2000) Proc. Natl. Acad. Sci. U S A. 97:10701). For example, minibodies can be generated using methods described in the art, e.g., Vaughan and Sollazzo, Comb Chem High Throughput Screen. 4:417-30 2001. Bispecific antibodies can be generated by a variety of methods including fusion of hybridomas or ligation of Fab’ fragments. See, e.g., Songsivilai & Lachmann, Clin. Exp. Immunol. 79:315-321 (1990); Kostelny et al., J. Immunol. 148, 1547-1553 (1992). Single-chain antibodies can be identified using phage display libraries, yeast display or ribosome display libraries, gene shuffling libraries. Such libraries can be constructed from synthetic, semi-synthetic or natural and immunocompetent sources.

[0033] "Monoclonal antibody" refers to a clonal preparation of an antibody having a single binding specificity and affinity for a given epitope on an antigen.

[0034] A "chimeric antibody" is an antibody molecule in which (a) the antigen-binding site (variable region, CDR, or a portion thereof) is such that the constant region or a portion thereof has been altered, replaced, or exchanged so that it is linked to a constant region of a different or modified class, effector function, and / or species, or to a wholly different molecule (e.g., an enzyme, toxin, hormone, growth factor, drug, etc.) that confers new properties to the chimeric antibody; or (b) the variable region or a portion thereof has been altered, replaced, or exchanged with a variable region having a different or modified antigen specificity (e.g., CDRs and framework regions from different species).

[0035] A "humanized" antibody is an antibody that retains the reactivity of a non-human antibody but has low immunogenicity in humans. This can be achieved, for example, by retaining the non-human CDR regions and replacing the rest of the antibody with their human counterparts. In one embodiment, some, most, or all of the amino acids outside the CDR domains are replaced with amino acids corresponding to the human immunoglobulin germline, while the amino acids within one or more CDR regions are unchanged. In some embodiments, one or more CDR residues can be altered, for example, to provide a sequence closer to the germline or to replace residues that may interfere with production.

[0036] The terms "specifically binds" or "specifically targets" refer to a molecule (e.g., an antibody or antibody fragment) that binds to a target with an affinity that is at least 2-fold greater, e.g., at least 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 25-fold, 50-fold, or 100-fold greater, than to a non-target compound. For example, an antibody that specifically binds to integrin beta-2 typically binds to integrin beta-2 with an affinity that is at least 2-fold higher than to a non-integrin beta-2 target, or in the case of an antibody that specifically binds to the active form of integrin beta-2, binds to the inactive form of integrin beta-2. In some embodiments, the antibody binds to the active integrin beta-2 with a K D that is at least 100-fold greater than to its affinity-inactive integrin beta-2.

[0037] "Epitope" or "antigenic determinant" refers to the site on an antigen to which an antibody binds. An epitope can be formed from both contiguous amino acids or non-contiguous amino acids juxtaposed by the tertiary folding of a protein. Epitopes formed from adjacent amino acids are typically retained upon exposure to a denaturing solvent, while epitopes formed by tertiary folding are typically lost upon treatment with a denaturing solvent. An epitope typically contains at least 3, more commonly at least 5 or 8 - 10 amino acids in its native spatial conformation. Methods for determining the spatial conformation of an epitope include, for example, x-ray crystallography and two-dimensional nuclear magnetic resonance. See, for example, Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, Glenn E. Morris, Ed (1996).

[0038] The term "valence", as used herein, refers to the number of different binding sites of an antibody to an antigen. A monovalent antibody contains one binding site to an antigen. A polyvalent antibody contains multiple binding sites.

[0039] The terms "protein", "peptide", and "polypeptide" are used interchangeably to denote an amino acid polymer or a set of two or more interacting or linked amino acid polymers. This term applies to amino acid polymers in which one or more amino acid residues are artificial chemical mimics of the corresponding naturally occurring amino acids, as well as to naturally occurring amino acid polymers, those containing modified residues, and amino acid polymers that do not occur naturally.

[0040] "Flexible linker", as used herein, refers to an amino acid sequence that joins domains and provides some degree of movement or interaction. Such linkers generally consist of small non-polar (e.g., Gly) or polar (e.g., Ser or Thr) amino acids, but may also include polar amino acids, such as Lys and Glu, for example, to improve solubility. These small-sized amino acids provide mobility and allow the mobility of the connected functional domains. Incorporation of Ser or Thr can maintain the stability of the linker in an aqueous solution by forming hydrogen bonds with water molecules, thus reducing unfavorable interactions between the linker and the protein moiety. In some embodiments, the flexible linker consists mainly of a stretch of Gly and Ser residues ("GS linker"). The most widely used example of a flexible linker has the sequence (Gly-Gly-Gly-Gly-Ser)n. By adjusting the copy number "n", the length of this GS linker can be adjusted to achieve appropriate separation of the functional domains and / or maintain the necessary domain-domain interactions. In addition to the GS linker, many other flexible linkers have been designed for recombinant protein expression.

[0041] The term "amino acid" refers to naturally occurring amino acids and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to naturally occurring amino acids. Naturally occurring amino acids include those encoded by the genetic code, as well as amino acids that are subsequently modified, such as hydroxyproline, y-carboxyglutamate, and O-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as naturally occurring amino acids, such as a carbon atom bonded to hydrogen, a carboxyl group, an amino group, and an R group, such as homoserine, norleucine, methionine sulfoxide, and methionine methyl sulfonium. Such analogs may have a modified R group (e.g., norleucine) or a modified peptide backbone, but retain the same basic chemical structure as naturally occurring amino acids. Amino acid mimetics refer to chemical compounds that have a structure different from the general chemical structure of amino acids, but function in a manner similar to naturally occurring amino acids.

[0042] Amino acids may be referred to herein by either their generally known three-letter symbols or the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Similarly, nucleotides may be referenced by their generally accepted one-letter codes.

[0043] "Conservatively modified variants" applies to both amino acid and nucleic acid sequences. With respect to a particular nucleic acid sequence, conservatively modified variants refers to nucleic acids that encode the identical or essentially identical amino acid sequence, or, when the nucleic acid does not encode an amino acid sequence, to essentially identical or related sequences, such as those that are naturally adjacent. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode most proteins. For example, the codons GCA, GCC, GCG, and GCU all encode the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be altered to another of the corresponding codons described without changing the encoded polypeptide. Such nucleic acid variations are "silent mutations" which are one species of conservatively modified mutations. Every nucleic acid sequence herein that encodes a polypeptide also describes every possible silent variation of the nucleic acid. One of skill will recognize that, in certain circumstances, it is possible to modify each codon of a nucleic acid (except AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan) in order to obtain a functionally identical molecule. Thus, silent mutations of a nucleic acid that encodes a polypeptide are included in the described sequence with respect to the expressed product, but not with respect to an actual probe sequence.

[0044] "Substitution", as used herein, refers to the substitution of an amino acid such that the charge, hydrophobicity, and / or size of the side chain group chain is maintained. Exemplary sets of amino acids that can be substituted for one another include: (i) the positively charged amino acids Lys, Arg, and His; (ii) the negatively charged amino acids Glu and Asp; (iii) the aromatic amino acids Phe, Tyr, and Trp; (iv) the nitrogen-ring amino acids His and Trp; (v) the large aliphatic non-polar amino acids Val, Leu, and Ile; (vi) the slightly polar amino acids Met and Cys; (vii) the small side-chain amino acids Ser, Thr, Asp, Asn, Gly, Ala, Glu, Gln, and Pro; (viii) the aliphatic amino acids Val, Leu, Ile, Met, and Cys; and (ix) the small hydroxyl amino acids Ser and Thr. References to the charge of amino acids in this paragraph refer to the charge at physiological pH.

[0045] The terms "nucleic acid" and "polynucleotide" are used interchangeably and, as used herein, refer to RNA, cDNA, both the sense and antisense strands of genomic DNA, as well as synthetic forms and hybrid polymers thereof. In certain embodiments, nucleotides refer to ribonucleotides, deoxynucleotides, or modified forms of either type of nucleotide, and combinations thereof. The term also includes, but is not limited to, single-stranded and double-stranded forms of DNA. Further, polynucleotides, such as cDNA or mRNA, can include either or both naturally occurring nucleotides and modified nucleotides linked together by naturally occurring nucleotide linkages and / or non-naturally occurring nucleotide linkages. Nucleic acid molecules, such as oligonucleotide probes or primers, may be chemically or biochemically modified or contain non-natural or derivatized nucleotide bases, as will be readily recognized by those of skill in the art. Such modifications include, for example, labeling, methylation, substitution by one or more analogs of a naturally occurring nucleotide, nucleotide modifications such as those by non-charged linkages (e.g., methylphosphonate, phosphotriester, phosphoramidate, carbamate, etc.), charged linkages (e.g., phosphorothioate, phosphorodithioate, etc.), those containing pendant moieties (e.g., polypeptides), those containing intercalators (e.g., acridine, psoralen, etc.), those containing chelators, those containing alkylating agents, and those containing modified linkages (e.g., alpha anomeric nucleic acids, etc.). The above terms are also intended to include any topological conformation, including single-stranded, double-stranded, partially double-stranded, triple-stranded, hairpin-type, circular, and padlock conformations. References to nucleic acid sequences include their complements unless otherwise specified. Thus, it should be understood that references to nucleic acid molecules having a particular sequence include the complementary strand, along with its complementary sequence. The term also includes codon-optimized nucleic acids encoding the same polypeptide sequence.

[0046] As used herein, the term "vector" refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is ligated. This term includes vectors as self-replicating nucleic acid structures and vectors integrated into the genome of a host cell into which it has been introduced. "Vector" as used herein refers to a recombinant construct into which a nucleic acid sequence of interest has been inserted. Certain vectors are capable of directing the expression of nucleic acids to which they are operably linked. Such vectors are referred to herein as "expression vectors".

[0047] The terms "identical" or "percent identity" in the context of two or more nucleic acids or polypeptides refer to sequences or subsequences that are the same or have a specified percentage of nucleotides or amino acids, i.e., when measured using the BLAST or BLAST 2.0 sequence comparison algorithms with default parameters, or by manual alignment and visual inspection, are the same (i.e., when compared and aligned with maximum correspondence over a comparison window or specified region, having about 60% identity, preferably 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher identity over the specified region) of two or more sequences. See, e.g., the NCBI web site at ncbi.nlm.nih.gov / BLAST. In such cases, such sequences are said to be "substantially identical". This definition also refers to or can apply to the complement of a nucleotide test sequence. This definition includes sequences having deletions and / or additions, as well as sequences having substitutions. As described below, the algorithms can take into account gaps, etc. Typically, identity exists over a region that includes an antibody epitope, or a sequence that is at least about 25 amino acids or nucleotides in length, or over a region that is 50 - 100 amino acids or nucleotides in length, or over the full length of a reference sequence.

[0048] The terms "corresponding to", "determined with reference to", or "numbered with reference to", when used in the context of a particular amino acid residue in a polypeptide sequence, refer to the position of the residue of a particular reference sequence when a given amino acid sequence is optimally aligned and compared to the reference sequence. Thus, for example, an amino acid residue in a variable domain polypeptide "corresponds to" an amino acid in SEQ ID NO:1 if the residue aligns optimally with SEQ ID NO:1 and matches the amino acid in SEQ ID NO:1. A polypeptide aligned to a reference sequence need not be the same length as the reference sequence.

[0049] The term "recombinant", when used with respect to, for example, a cell, or a nucleic acid, protein, or vector, indicates that the cell, nucleic acid, protein, or vector has been modified by the introduction of a heterologous nucleic acid or protein or the alteration of a native nucleic acid or protein, or that the cell is derived from a cell so modified. Thus, for example, a recombinant cell expresses a gene not found within the natural (non-recombinant) form of the cell or expresses a native gene that is otherwise abnormally expressed, under-expressed, or not expressed at all.

[0050] The term "heterologous", when used with respect to a portion of a nucleic acid, indicates that the nucleic acid comprises two or more subsequences that are not found in essentially the same relationship to each other in nature. For example, a nucleic acid is typically produced recombinantly and has two or more sequences from unrelated genes arranged to create a new functional nucleic acid, such as a promoter from one source and a coding region from another source. Similarly, a heterologous protein indicates that the protein comprises two or more subsequences that are not found in the same relationship to each other in nature (e.g., a fusion protein).

[0051] The term "isolated," when applied to a nucleic acid or protein, indicates that the nucleic acid or protein is substantially free of other cellular components with which it is associated in nature. It is preferably in a homogeneous state. This can be either in a dry solution or an aqueous solution. Purity and homogeneity are typically determined using analytical chemistry techniques such as polyacrylamide gel electrophoresis or high performance liquid chromatography. The protein, which is the major species present in the preparation, is substantially purified. In particular, an isolated gene is separated from open reading frames adjacent to the gene and encoding proteins other than the gene of interest. The term "purified" means that the nucleic acid or protein gives rise to essentially one band in an electrophoretic gel. In particular, this means that the nucleic acid or protein is at least 85% pure, more preferably at least 95% pure, and most preferably at least 99% pure.

[0052] The terms "subject," "patient," "individual," etc. are used interchangeably and, unless indicated otherwise, refer to mammals such as humans and non-human primates, as well as rabbits, rats, mice, goats, pigs, and other mammalian species. This term does not necessarily indicate that the subject has been diagnosed with a particular disease, but typically refers to an individual under medical supervision. A patient can be an individual seeking treatment, monitoring, adjustment, or modification of an existing treatment regimen.

[0053] Terms such as "cancer", "tumor", "transformed" include pre-cancerous, neoplastic, transformed, and cancerous cells and can refer to solid tumors or non-solid cancers. Cancer includes both benign neoplasms and malignant neoplasms (abnormal growths). Thus, the term "cancer" can refer to carcinomas, sarcomas, adenocarcinomas, lymphomas, leukemias, solid and lymphoid cancers, etc. Examples of various types of cancer include, but are not limited to, lung cancer (e.g., non-small cell lung cancer or NSCLC), ovarian cancer, prostate cancer, colorectal cancer, liver cancer (i.e., hepatocellular carcinoma), kidney cancer (i.e., renal cell carcinoma), bladder cancer, breast cancer, thyroid cancer, pleural cancer, pancreatic cancer, uterine cancer, cervical cancer, testicular cancer, anal cancer, pancreatic cancer, cholangiocarcinoma, gastrointestinal carcinoid tumor, esophageal cancer, gallbladder cancer, appendiceal cancer, small intestine cancer, stomach (stomach) (gastric) cancer, central nervous system cancer, skin cancer, choriocarcinoma; head and neck cancer, blood cancer, osteogenic sarcoma, fibrosarcoma, neuroblastoma, glioma, melanoma, B-cell lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, small cell lymphoma, large cell lymphoma, monocytic leukemia, myeloid leukemia, acute lymphocytic leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and multiple myeloma. In some embodiments, the antibody compositions and methods described herein can be used to treat cancer.

[0054] As used interchangeably herein, the terms "chimeric antigen receptor" and "CAR" generally refer to an artificial multi-module molecule, although not exclusively, that includes an extracellular domain (e.g., ligand / antigen binding domain), a transmembrane domain, and one or more intracellular signaling domains capable of inducing or inhibiting the activation of immune cells. The term "CAR" is not specifically limited to CAR molecules and also includes CAR variants. CAR variants include split CARs in which the extracellular portion (e.g., ligand binding portion) and the intracellular portion (e.g., intracellular signaling portion) of the CAR are present on two separate molecules. CAR variants also include conditionally activatable CARs, such as on-switch CARs that include split CARs in which the conditional heterodimerization of the two parts of the split CAR is pharmacologically controlled. CAR variants also include bispecific CARs that include a secondary CAR binding domain capable of amplifying or inhibiting the activity of the primary CAR. CAR variants also include inhibitory chimeric antigen receptors (iCARs) that can be used, for example, as components of a bispecific CAR system in which the binding of the secondary CAR binding domain results in inhibition of primary CAR activation. CAR molecules and their derivatives (i.e., CAR variants) are described, for example, in PCT application number US2014 / 016527; Fedorov et al., Sci Transl Med (2013); 5(215):215ra172; Glienke et al., Front Pharmacol (2015) 6:21; Kakarla & Gottschalk, 52 Cancer J (2014) 20(2):151-5; Riddell et al., Cancer J (2014) 20(2):141-4; Pegram et al., Cancer J (2014) 20(2):127-33; Cheadle et al., Immunol Rev (2014) 257(1):91-106; Barrett et al., Annu Rev Med (2014) 65:333-47; Sadelain et al., Cancer Discov (2013) 3(4):388-98; Cartellieri et al., J Biomed Biotechnol (2010) 956304; the disclosures of which are incorporated herein by reference in their entirety.

[0055] As used herein, the term "immune cell" generally includes white blood cells (leukocytes) derived from hematopoietic stem cells (HSCs) generated in bone marrow "immune cells", including, for example, lymphocytes (T cells, B cells, natural killer (NK) cells) and bone marrow-derived cells (neutrophils, eosinophils, basophils, monocytes, macrophages, dendritic cells).

[0056] "T cells" include all types of immune cells expressing CD3, including helper T cells (CD4+ cells), cytotoxic T cells (CD8+ cells), regulatory T cells (Tregs) and gamma-delta T cells.

[0057] "Cytotoxic cells" include CD8+ T cells, natural killer (NK) cells and neutrophils, and these cells are capable of mediating cytotoxic responses. Selection of anti-integrin beta 2 antibodies by phage display

[0058] Referring to Figure 1, there is shown a diagram of a phage display selection strategy used to develop anti-integrin beta-2 antibodies. Using the previously described Fab-phage display platform (Persson, et al., J. Mol. Biol. 425:803-811, 2013) based on a fully human framework sequence, selection against recombinant integrin beta-2 containing integrin beta-2 / integrin alpha-M (R and D 4047-AM, antibody numbers 7061, 7062, 7063, 7064, 7065), integrin-beta2 / integrin alpha-L (R and D 3868-AV, antibody numbers 7060, 7341) and integrin-beta2 / integrin alpha-X (R and D 5755-AX, antibody numbers 7055, 7056, 7057) recombinant heterodimeric protein complexes was performed.

[0059] Briefly, the integrin beta-2 recombinant protein complex was immobilized on a Maxisorp Immuno Plate (ThermoFisher, 12-565-135), and positive binding selection was performed using a library phage pool initially exposed to wells coated with neutravidin to deplete non-specific binders. After four rounds of binding selection, clonal phage were prepared and evaluated by phage ELISA and sequencing as previously described (Persson, et al., supra) and summarized below.

[0060] From the diverse approximately 10 10 individual conjugates of the initial library, 10 initial Fab hits shown in FIGS. 2A-2D and 3 were identified against the integrin-beta2 recombinant heterodimer protein complex. Seven of the Fab hits (number 7055, number 7056, number 7057, number 7060, number 7061, number 7062, number 7063, number 7064, number 7065, number 7341) were further verified to have binding affinities in the low nM range for integrin beta-2 and no binding to irrelevant proteins using biolayer interferometry (BLI) (see FIGS. 5A-5R and 6) and non-specific ELISA (see FIGS. 4A-4H), respectively. These Fabs were cloned into the human IgG1 backbone and purified after recombinant expression in mammalian cells such as Expi93 human fetal kidney cells or Chinese hamster ovary cells.

[0061] Antibody production The anti-integrin beta-2 antibody was produced using the human Expi293 expression system (Thermofisher). Expi293 cells in a volume of 2 ml were transiently transfected with the construct DNA using the FectoPro transfection reagent (Polyplus Transfection, 101000014). After a 5-day expression period, the antibody was purified using rProteinA Sepharasoe (GE Healthcare) and stored in phosphate buffer (50 mM NaH2PO4, 75 mM Na2HPO4, 100 mM H3PO4, 154 mM NaCl). Biolayer interferometry (BLI) binding assay

[0062] The binding of the human integrin beta-2 antibody was tested against three different integrin beta-2 complexes, including integrin beta-2 / integrin alpha-M (R&D 4047-AM), integrin beta-2 / integrin alpha-X (R&D 5755-AX), and integrin beta-2 / integrin alpha-L (R&D 3868-AV). To determine the binding kinetic parameters of the antibody, BLI experiments were performed at 1000 rpm and 25 °C on an Octet HTX instrument (Sartorius). All proteins were diluted in assay buffer (PBS, 1% BSA, 0.05% Tween 20). The test antibody and the control antibody at a concentration of 2 μg / ml were first captured on the AHQ biosensor to achieve a binding signal of 0.8 - 1.3 nm. Subsequently, the unoccupied Fc binding sites on the antibody-coated sensor were quenched with 20 μg / mL of Fc protein. After equilibration with the assay buffer, the biosensor was then immersed in wells containing 5-fold serial dilutions of the integrin-beta2 complex for 600 seconds (association phase), followed by another 600 seconds back in the assay buffer (dissociation phase). Assay buffer alone was used as a negative control. The binding response data were baseline-subtracted and globally fitted to a 1:1 binding model using the ForteBio Octet Systems software 9.0. ELISA An ELISA protocol for evaluating the interaction between an antibody and a polymer unrelated to the antibody was adapted from Meirsch et al. (J Vis Exp. 2015 Jan 17;(95):51492). The antigens tested included integrin αL / β2 (50 μg / mL, R&D systems 3868-AV-050), integrin αX / β2 (50 μg / mL, R&D systems 5755-AX-050), integrin αM / β2 (50 μg / mL, R&D systems 4047-AM-050), histidine-tagged Sumo domain (100 μg / mL, recombinant), biotinylated Robo domain (100 μg / mL, recombinant), NeutrAvidin (100 μg / mL, Pierce TPPI31000) or all other integrin proteins (50 μg / mL R&D system). In addition, the binding of each antibody was also tested against empty wells (BSA-only control) and wells containing goat anti-human Fc antibody (positive control, 1 μg / ml, Jackson 109-005-098). The antigens were coated at 30 μL / well in a 384-well Maxisorp plate and incubated overnight at 4°C. The plate was blocked with 0.5% bovine serum albumin (BSA) for 1 hour at room temperature and washed with PBS + 0.05% Tween20. Phage-Fab was added and allowed to bind for 60 minutes at room temperature. The plate was washed with PBS + 0.05% Tween20, and the binding was detected with anti-M13 HRP antibody (1:5000, Sinobiological 11973-MM05T-H) and developed with TMB substrate (KPL (Mandel) KP-50-76-03).

[0063] Statistical analysis Unless otherwise specified, all statistical analyses were performed using GraphPad Prism. Data were represented as mean ±, and p < 0.05 was considered statistically significant.

[0064] Anti-integrin beta-2 antibody Provided herein are anti-integrin beta-2 antibodies that can be used for diagnostic and therapeutic purposes.

[0065] In some embodiments, the anti-integrin beta-2 antibodies of the present disclosure have a KD of less than about 10 nM.

[0066] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure has at least one, at least two, or three CDRs of the variable domain sequence of SEQ ID NO: 2 or SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCDR3 of SEQ ID NO: 2 and the LCDR3 of SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises the HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2 and the LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3.

[0067] In some embodiments, the anti-integrin beta-2 binding domain comprises the HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 2. In some embodiments, the anti-integrin beta-2 binding domain comprises the HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 2. In some embodiments, the anti-integrin beta-2 binding domain comprises the HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 2, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 2.

[0068] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 3, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 3.

[0069] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 2 or SEQ ID NO: 3. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 2 or SEQ ID NO: 3. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 2, a heavy chain variable region having at least 95% identity to SEQ ID NO: 2; and the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 3, a light chain variable region having at least 95% identity to SEQ ID NO: 3.

[0070] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure has at least one, at least two or three CDRs of the variable domain sequences of SEQ ID NO: 4 or SEQ ID NO: 5. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure includes HCDR3 of SEQ ID NO: 4 and LCDR3 of SEQ ID NO: 5. In some embodiments, the anti-integrin beta-2 binding domain includes HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 4 and LCDR1, LCDR2 and LCDR3 of SEQ ID NO: 5.

[0071] In some embodiments, the anti-integrin beta-2 binding domain includes HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 4, wherein one of the CDRs includes a substitution relative to the corresponding CDR set forth in SEQ ID NO: 4. In some embodiments, the anti-integrin beta-2 binding domain includes HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 4, wherein two of the CDRs include substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 4. In some embodiments, the anti-integrin beta-2 binding domain includes HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 4, wherein all three of the CDRs include substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 4.

[0072] In some embodiments, the anti-integrin beta-2 binding domain includes LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 5, wherein one of the CDRs includes a substitution relative to the corresponding CDR set forth in SEQ ID NO: 5. In some embodiments, the anti-integrin beta-2 binding domain includes LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 5, wherein two of the CDRs include substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 5. In some embodiments, the anti-integrin beta-2 binding domain includes LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 5, wherein all three of the CDRs include substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 5.

[0073] In some embodiments, the anti-integrin β-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 4 or SEQ ID NO: 5. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 4 or SEQ ID NO: 5. In some embodiments, the anti-integrin β-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 4 and has a heavy chain variable region having at least 95% identity to SEQ ID NO: 4; and comprises the LCD1, LCDR2, LCDR3 sequences and has a light chain variable region having at least 95% identity to SEQ ID NO: 5.

[0074] In some embodiments, the anti-integrin β-2 binding domain of the present disclosure has at least 1, at least 2 or 3 CDRs of the variable domain sequence of SEQ ID NO: 6 or SEQ ID NO: 7. In some embodiments, the anti-integrin β-2 binding domain of the present disclosure comprises HCDR3 of SEQ ID NO: 6 and LCDR3 of SEQ ID NO: 7. In some embodiments, the anti-integrin β-2 binding domain comprises HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 6 and LCDR1, LCDR2 and LCDR3 of SEQ ID NO: 7.

[0075] In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 6, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 6. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 6, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 6. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 6, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 6.

[0076] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 7, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 7. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 7, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 7. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 7, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 7.

[0077] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 6 or SEQ ID NO: 7. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 6 or SEQ ID NO: 7. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 6 and has a heavy chain variable region having at least 95% identity to SEQ ID NO: 6; and comprises the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 7 and has a light chain variable region having at least 95% identity to SEQ ID NO: 7.

[0078] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure has at least 1, at least 2 or 3 CDRs of the variable domain sequence of SEQ ID NO: 8 or SEQ ID NO: 9. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises HCDR3 of SEQ ID NO: 8 and LCDR3 of SEQ ID NO: 9. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 8 and LCDR1, LCDR2 and LCDR3 of SEQ ID NO: 9.

[0079] In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 8, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 8. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 8, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 8. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 8, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 8.

[0080] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 9, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 9. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 9, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 9. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 9, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 9.

[0081] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 8 or SEQ ID NO: 9. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 8 or SEQ ID NO: 9. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 8 and has a heavy chain variable region having at least 95% identity to SEQ ID NO: 8; and comprises the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 9 and has a light chain variable region having at least 95% identity to SEQ ID NO: 9.

[0082] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure has at least one, at least two or three CDRs of the variable domain sequence of SEQ ID NO: 10 or SEQ ID NO: 11. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises HCDR3 of SEQ ID NO: 10 and LCDR3 of SEQ ID NO: 11. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 10 and LCDR1, LCDR2 and LCDR3 of SEQ ID NO: 11.

[0083] In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 10, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 10. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 10, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 10. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 10, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 10.

[0084] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 11, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 11. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 11, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 11. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 11, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 11.

[0085] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 10 or SEQ ID NO: 11. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 10 or SEQ ID NO: 11. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 10 and has a heavy chain variable region having at least 95% identity to SEQ ID NO: 10; and comprises the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 11 and has a light chain variable region having at least 95% identity to SEQ ID NO: 11.

[0086] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure has at least one, at least two or three CDRs of the variable domain sequence of SEQ ID NO: 12 or SEQ ID NO: 13. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises HCDR3 of SEQ ID NO: 12 and LCDR3 of SEQ ID NO: 13. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 12 and LCDR1, LCDR2 and LCDR3 of SEQ ID NO: 13.

[0087] In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 12, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 12. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 12, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 12. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 12, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 12.

[0088] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 13, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 13. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 13, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 13. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 13, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 13.

[0089] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 12 or SEQ ID NO: 13. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 12 or SEQ ID NO: 13. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 12, a heavy chain variable region having at least 95% identity to SEQ ID NO: 12; and the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 13, a light chain variable region having at least 95% identity to SEQ ID NO: 13.

[0090] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure has at least 1, at least 2 or 3 CDRs of the variable domain sequence of SEQ ID NO: 14 or SEQ ID NO: 15. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises HCDR3 of SEQ ID NO: 14 and LCDR3 of SEQ ID NO: 15. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2 and HCDR3 of SEQ ID NO: 14 and LCDR1, LCDR2 and LCDR3 of SEQ ID NO: 15.

[0091] In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 14, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 14. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 14, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 14. In some embodiments, the anti-integrin beta-2 binding domain comprises HCDR1, HCDR2, and HCDR3 of SEQ ID NO: 14, wherein all three of the CDRs comprise substitutions relative to the corresponding CDR sequences set forth in SEQ ID NO: 14.

[0092] In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 15, wherein one of the CDRs comprises a substitution relative to the corresponding CDR set forth in SEQ ID NO: 15. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 15, wherein two of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 15. In some embodiments, the anti-integrin beta-2 binding domain comprises LCDR1, LCDR2, and LCDR3 of SEQ ID NO: 15, wherein all three of the CDRs comprise substitutions relative to the corresponding CDRs set forth in SEQ ID NO: 15.

[0093] In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises a variable region having at least 70%, 75%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of the variable region sequence of SEQ ID NO: 14 or SEQ ID NO: 15. In some embodiments, the variable domain comprises substitutions, insertions or deletions in the framework of the variable region shown in SEQ ID NO: 14 or SEQ ID NO: 15. In some embodiments, the anti-integrin beta-2 binding domain of the present disclosure comprises the HCD1, HCDR2 and HCDR3 sequences of SEQ ID NO: 14 and has a heavy chain variable region having at least 95% identity to SEQ ID NO: 14; and the LCD1, LCDR2, LCDR3 sequences of SEQ ID NO: 15 and has a light chain variable region having at least 95% identity to SEQ ID NO: 15.

[0094] Antibody format The anti-integrin beta-2 antibodies of the present disclosure can be incorporated into bivalent or multivalent antibodies that bind the same antigen or different antigens. In some embodiments, the anti-integrin beta-2 antibodies of the present disclosure can be incorporated into bispecific or multispecific antibodies that bind the antigen at different epitopes or bind different antigens. In some embodiments, such antibodies can comprise an Fc region. In some embodiments, the anti-integrin beta-2 antibodies of the present disclosure can be present as the antigen-binding domain of a larger molecule, for example, as the antigen-binding domain of a chimeric antigen receptor or a synthetic Notch receptor.

[0095] Nucleic acids and vectors encoding CAR Any method may be used to genetically modify effector cells, such as T cells or NK cells, to express a CAR comprising an anti-integrin beta-2 antibody of the present disclosure. Non-limiting examples of methods for genetically engineering immune cells include, but are not limited to, transduction mediated by retroviruses or lentiviruses. Other viral delivery systems include adenoviruses, adeno-associated viruses, herpes simplex virus vectors, poxvirus vectors, alphavirus vectors, poliovirus vectors, and other positive and negative strand RNA viruses, viroids, and virusoids, or portions thereof. Methods of transduction include, for example, direct co-culture of cells with producer cells by the method of Bregni, et al. (Blood 80:1418-1422 (1992)), or culture with viral virus supernatant alone or concentrated vector stock, with or without appropriate growth factors and polycations, by the methods of, for example, Xu, et al. Exp. Hemat. 22:223-230 (1994); and Hughes, et al. J. Clin. Invest. 89:1817 (1992).

[0096] In some embodiments, the genetic modification is performed using a transposase-based system for gene integration, CRISPR / Cas-mediated gene integration, TALEN, or zinc finger nuclease integration techniques. For example, CRISPR / Cas-mediated gene integration may be used to introduce a CAR or synthetic Notch receptor into immune effector cells, which may then be selected and expanded for administration to a patient.

[0097] Antibody conjugate In a further aspect, the anti-integrin beta-2 antibodies of the present disclosure can be directly or indirectly conjugated or linked to a therapeutic moiety and / or an imaging moiety / detectable moiety. For example, in some embodiments, the antibodies of the present disclosure, or antigen-binding regions comprising the antibodies of the invention, can be conjugated to agents including, but not limited to, detectable markers, cytotoxic agents, imaging agents, therapeutic agents, or oligonucleotides. Methods for conjugating or linking an antibody or an antigen-binding region comprising an antibody to a desired molecular moiety are well known in the art. The moiety can be linked to the antibody by covalent or non-covalent bonds.

[0098] In some embodiments, the anti-integrin beta-2 antibodies of the present disclosure, or antigen-binding domains comprising the anti-integrin beta-2 antibodies of the present disclosure, are conjugated to a cytotoxic moiety or other moiety that inhibits cell proliferation.

[0099] In some embodiments, the anti-integrin beta-2 antibodies of the present disclosure are conjugated to cytotoxic agents including, but not limited to, for example, lysin A chain, doxorubicin, daunorubicin, maytansinoid, taxol, ethidium bromide, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, dihydroxyanthracinedione, methotrexact, actinomycin, diphtheria toxin, Pseudomonas-derived exotoxin A, Pseudomonas exotoxin 40, abrin, abrin A chain, modeccin A chain, alpha-sarcin, gelonin, mitogelin, restrictocin, cobra venom factor, ribonuclease, engineered Shiga toxin, phenomycin, enomycin, curicin, crotonin, calicheamicin, Saponaria officinalis inhibitor, glucocorticoid, auristatin, auroomycin, yttrium, bismuth, combrestatin, duocarmycin, dostarlimab, cc1065 or cisplatin. In some embodiments, the antibody may be linked to agents such as enzyme inhibitors, growth inhibitors, lysing agents, DNA or RNA synthesis inhibitors, membrane permeability modifiers, DNA metabolites, dichloroethyl sulfide derivatives, protein production inhibitors, ribosome inhibitors, or inducers of apoptosis.

[0100] In some embodiments, the anti-integrin beta-2 antibodies of the present disclosure, or antigen-binding domains comprising the anti-integrin beta-2 antibodies of the present disclosure, may be conjugated to radionuclides, iron-related compounds, dyes, fluorescent agents, or contrast agents. In some embodiments, the antibody may be linked to agents such as, but not limited to, metals; metal chelating agents; lanthanides; lanthanide chelating agents; radioactive metals; radioactive metal chelating agents; positron-emitting nuclei; microbubbles (for ultrasound); liposomes; molecules microencapsulated in liposomes or nanospheres; single-crystalline iron oxide nanocompounds; magnetic resonance imaging contrast agents; light absorbers, reflectors and / or scatterers; colloidal particles; fluorophores such as near-infrared fluorophores.

[0101] The foregoing description provides examples of one or more antibodies and nucleic acids, but it will be recognized that other compositions may also fall within the scope of the claims, as will be interpreted by those skilled in the art. Polypeptide sequence: SEQ ID NO:1 Uniprot P05107-1 amino acid sequence MLGLRPPLLA LVGLLSLGCV LSQECTKFKV SSCRECIESG PGCTWCQKLN FTGPGDPDSI RCDTRPQLLM RGCAADDIMD PTSLAETQED HNGGQKQLSP QKVTLYLRPG QAAAFNVTFR RAKGYPIDLY YLMDLSYSML DDLRNVKKLG GDLLRALNEI TESGRIGFGS FVDKTVLPFV NTHPDKLRNP CPNKEKECQP PFAFRHVLKL TNNSNQFQTE VGKQLISGNL DAPEGGLDAM MQVAACPEEI GWRNVTRLLV FATDDGFHFA GDGKLGAILT PNDGRCHLED NLYKRSNEFD YPSVGQLAHK LAENNIQPIF AVTSRMVKTY EKLTEIIPKS AVGELSEDSS NVVQLIKNAY NKLSSRVFLD HNALPDTLKV TYDSFCSNGV THRNQPRGDC DGVQINVPIT FQVKVTATEC IQEQSFVIRA LGFTDIVTVQ VLPQCECRCR DQSRDRSLCH GKGFLECGIC RCDTGYIGKN CECQTQGRSS QELEGSCRKD NNSIICSGLG DCVCGQCLCH TSDVPGKLIY GQYCECDTIN CERYNGQVCG GPGRGLCFCG KCRCHPGFEG SACQCERTTE GCLNPRRVEC SGRGRCRCNV CECHSGYQLP LCQECPGCPS PCGKYISCAE CLKFEKGPFG KNCSAACPGL QLSNNPVKGR TCKERDSEGC WVAYTLEQQD GMDRYLIYVD ESRECVAGPN IAAIVGGTVA GIVLIGILLL VIWKALIHLS DLREYRRFEK EKLKSQWNND NPLFKSATTT VMNPKFAES Sequence number 2 antibody - 7065 heavy chain variable region; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT ISYYYM HWVRQAPGKGLEWVA SISSSSGYTY YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR GAM DYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: ISYYYM HCDR2: SISSSSGYTY HCDR3: GAM Sequence number 3 antibody - 7065 light chain variable region; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ FSSGSWAPI TFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: FSSGSWAPI Sequence number 4 antibody - 7060 heavy chain variable region; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT LSYSSM HWVRQAPGKGLEWVA YIYPSYGYTY YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR WSPGSGWAF DYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: LSYSSM HCDR2: YIYPSYGYTY HCDR3: WSPGSGWAF Sequence number 5, variable region of antibody-7060 light chain; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ YHGSLI TFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: YHGSLI Sequence number 6, variable region of antibody-7062 heavy chain; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT ISSYSI HWVRQAPGKGLEWVA SIYSYYGYTS YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR YWGYPYAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: ISSYSI HCDR2: SIYSYYGYTS HCDR3: YWGYPYAM Sequence number 7, variable region of antibody-7062 light chain; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ YYYAASLF TFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: YYYAASLF Sequence number 8, variable region of antibody-7063 heavy chain; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT LSYSYM HWVRQAPGKGLEWVASIYSYYSSTS YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR SYHYSYYAGL DYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: LSYSYM HCDR2: SIYSYYSSTS HCDR3: SYHYSYYAGL Sequence number 9, light chain variable region of antibody - 7063; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ WYFLI TFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: WYFLI Sequence number 10, heavy chain variable region of antibody - 7064; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT LSYSSM HWVRQAPGKGLEWVA YIYSSSGYTY YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR WGWYAHAGM DYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: LSYSSM HCDR2: YIYSSSGYTY HCDR3: WGWYAHAGM Sequence number 11 Antibody - 7064 light chain variable region; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ WVHGLI TFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: WVHGLI Sequence number 12, heavy chain variable region of antibody - 7056; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT LYYYSM HWVRQAPGKGLEWVA YIYPYYGYTS YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR TVRGSKKPYFSGWAM DYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: LYYYSM HCDR2: YIYPYYGYTS HCDR3: TVRGSKKPYFSGWAM Sequence number 13, light chain variable region of antibody - 7056; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ WGAWGPLI TFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: WGAWGPLI SEQ ID NO: 14 Antibody-7341 heavy chain variable region; CDRs are underlined EVQLVESGGGLVQPGGSLRLSCAASGFT LSYYYM HWVRQAPGKGLEWVA SISSYYGYTS YADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAR GAL DYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK HCDR1: LSYYYM HCDR2: SISSYYGYTS HCDR3: GAL SEQ ID NO: 15 Antibody-7341 light chain variable region; CDRs are underlined DIQMTQSPSSLSASVGDRVTITCRASQ SVSSA VAWYQQKPGKAPKLLIY SASSLYS GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQ FYGGYSLITFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC LCDR1: SVSSA LCDR2: SASSLYS LCDR3: FYGGYSLI

Claims

1. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing ISYYYYM, an HCDR2 sequence containing SISSSSGYT, and an HCDR3 sequence containing GAM; and A light chain variable region (V) comprising an LCDR1 sequence containing SVSSA, an LCDR2 sequence containing SASSLYS; and an LCDR3 sequence containing FSSGSWAPI Ｌ ) An antibody.

2. Said V Ｈ comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 2; and / or Said V Ｌ comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 3 The antibody according to claim 1.

3. Said V Ｈ comprises the amino acid sequence of SEQ ID NO: 2; and / or Said V Ｌ The antibody according to claim 1, wherein V comprises the amino acid sequence of SEQ ID NO:

3.

4. The antibody according to any one of claims 1 to 3, which is a single-chain variable fragment (scFv).

5. Said V Ｌ being on the N-terminal side with respect to said V Ｈ The antibody according to claim 4.

6. The aforementioned V Ｈ is the aforementioned V Ｌ The antibody according to claim 4, which is on the N-terminal side with respect to the aforementioned V.

7. said V Ｈ and said V Ｌ The antibody according to any one of claims 5 to 6, wherein are separated by a flexible linker.

8. The antibody according to claim 7, wherein the flexible linker contains Gly-Ser.

9. The antibody according to claim 8, wherein the flexible linker contains one or more Gly-Ser sequences.

10. A bispecific or multispecific antibody comprising the antibody according to any one of claims 1 to 9.

11. A chimeric antigen receptor (CAR) comprising an antigen-binding domain, wherein the antigen-binding domain comprises the antibody according to any one of claims 1 to 9.

12. An immune effector cell comprising the CAR according to claim 11.

13. The immune effector cell according to claim 12, wherein the cell is a T cell or an NK cell.

14. A polynucleotide encoding the antibody according to any one of claims 1 to 11.

15. A vector comprising the polynucleotide according to claim 14.

16. An immune effector cell comprising the vector according to claim 15.

17. Antibody V Ｈ and / or V Ｌ A nucleic acid encoding, wherein said V Ｈ comprises an HCDR1 sequence containing ISYYYYM, an HCDR2 sequence containing SISSSSGYT Y; and an HCDR3 sequence containing GAM; and Said V Ｌ is a nucleic acid comprising an LCDR1 sequence containing SVSSA, an LCDR2 sequence containing SASSLYS; and an LCDR3 sequence containing FSSGSWAPI.

18. Said V Ｈ comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 2; and / or Said V Ｌ The nucleic acid according to claim 17, wherein V comprises an amino acid sequence having at least 95% identity to SEQ ID NO:

3.

19. Said V Ｈ comprises the amino acid sequence of SEQ ID NO: 2; and / or Said V Ｌ The nucleic acid according to claim 17, wherein V comprises the amino acid sequence of SEQ ID NO:

3.

20. A vector comprising the nucleic acid according to any one of claims 17 to 19.

21. A host cell comprising the vector according to claim 20.

22. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing LSYS SM, an HCDR2 sequence containing YIYPSYGYTY; and an HCDR3 sequence containing WSPGSG WAF; and A light chain variable region (V Ｌ ) containing an LCDR1 sequence including SVSSA, an LCDR2 sequence including SASSLYS; and an LCDR3 sequence including YHGSLI An antibody.

23. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing LYYYSM, an HCDR2 sequence containing YIYPYYGYTS; and an HCDR3 sequence containing TVRGSKKPYFSGWAM; and A light chain variable region (V Ｌ ) containing an LCDR1 sequence including SVSSA, an LCDR2 sequence including SASSLYS; and an LCDR3 sequence including WGAWGPLL An antibody.

24. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing LSYYYM, an HCDR2 sequence containing SISSYYGYT, and an HCDR3 sequence containing GAL; and A light chain variable region (V Ｌ ) containing an LCDR1 sequence including SVSSA, an LCDR2 sequence including SASSLYS; and an LCDR3 sequence including FYGGYSLI An antibody.

25. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing ISSYSI, an HCDR2 sequence containing SIYYYGYTS; and an HCDR3 sequence containing YWGYPYAM; and A light chain variable region (V) comprising an LCDR1 sequence containing SVSSA, an LCDR2 sequence containing SASSLYS; and an LCDR3 sequence containing YYYAASLF Ｌ ) An antibody.

26. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing LSYSYM, an HCDR2 sequence containing SIYSYYSSST, and an HCDR3 sequence containing SYHYSYYAGL; and A light chain variable region (V Ｌ ) containing an LCDR1 sequence including SVSSA, an LCDR2 sequence including SASSLYS; and an LCDR3 sequence including WYFLI An antibody.

27. An antibody that specifically binds to integrin beta-2 and contains an integrin beta-2 binding domain containing A heavy chain variable region (V Ｈ ) comprising an HCDR1 sequence containing LSYSSM, an HCDR2 sequence containing YIYSSSGYT, and an HCDR3 sequence containing WGWYAHAGM; and A light chain variable region (V Ｌ ) containing an LCDR1 sequence including SVSSA, an LCDR2 sequence including SASSLYS; and an LCDR3 sequence including WVHGLI An antibody.

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