Anti-IGFBP7 constructs and uses thereof

Abstract

The present application provides an anti-IGFBP7 construct (e.g., an anti-IGFBP7 antibody) that binds to IGFBP7, a nucleic acid molecule that encodes the amino acid sequence of anti-IGFBP7, a vector that includes the nucleic acid molecule, a host cell that includes the vector, a method for preparing the anti-IGFBP7 construct, a pharmaceutical composition that includes the anti-IGFBP7 construct, and a method for using the anti-IGFBP7 construct or composition. The anti-IGFBP7 construct may include a polypeptide that includes a single domain antibody (sdAb) portion that specifically recognizes IGFBP7.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 63 / 166,146, filed March 25, 2021, the contents of which are incorporated by reference in their entirety for all purposes.

[0002] Technical Field The present disclosure relates to anti-IGFBP7 constructs (such as anti-IGFBP7 antibodies) and uses thereof.

[0003] Submission of sequence listing as an ASCII text file The contents of the following submission in an ASCII text file are incorporated herein by reference in their entirety: Computer Readable Format (CRF) of the Sequence Listing (Filename: 193852000540SEQLIST.TXT, Recorded Date: March 24, 2022, Size: 56,698 bytes). [Background technology]

[0004] Background of the Application IGFBP7 (insulin-like growth factor binding protein 7) is a protein that in humans is encoded by the IGFBP7 gene. Insulin-like growth factors (IGFs) are proteins involved in promoting cell growth and division and preventing premature apoptosis. The main function of IGFBP7 is to regulate the availability of IGFs in body fluids and tissues, as well as regulating the binding of IGFs to their receptors. IGFBP7 is active in the inner lining of blood vessels; its interaction with IGFs and IGF receptors is important for controlling BRAF signaling, which is involved in directing cell growth. From these processes, IGFBP7 is thought to be involved in cell adhesion and cancer.

[0005] The disclosures of all publications, patents, patent applications and published patent applications mentioned herein are hereby incorporated by reference in their entirety. Summary of the Invention [Means for solving the problem]

[0006] BRIEF SUMMARY OF THE APPLICATION The following summary is illustrative only and is not intended to be limiting in any way. That is, the following summary is provided to introduce highlights, benefits, and advantages of the novel molecules and their uses. As such, the following summary is not intended to identify essential features of the claimed subject matter, nor is it intended for use in determining the scope of the claimed subject matter.

[0007] The present application provides in one aspect an anti-IGFBP7 construct comprising a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, the sdAb portion comprising: 1) CDR1 comprising the amino acid sequence of SEQ ID NO: 1, CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 2) CDR1 comprising the amino acid sequence of SEQ ID NO: 2 or 112, CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 3) CDR1 comprising the amino acid sequence of SEQ ID NO: 5 or 114, CDR2 comprising the amino acid sequence of SEQ ID NO: 6, and CDR3 comprising the amino acid sequence of SEQ ID NO: 7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 4) CDR1 comprising the amino acid sequence of SEQ ID NO:8, CDR2 comprising the amino acid sequence of SEQ ID NO:12, and CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 5) CDR1 comprising the amino acid sequence of SEQ ID NO: 9, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 6) CDR1 comprising the amino acid sequence of SEQ ID NO: 10, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 7) CDR1 comprising the amino acid sequence of SEQ ID NO: 11, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 8) CDR1 comprising the amino acid sequence of SEQ ID NO: 14, CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 9) CDR1 comprising the amino acid sequence of SEQ ID NO: 17, CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 10) CDR1 comprising the amino acid sequence of SEQ ID NO: 20, CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and CDR3 comprising the amino acid sequence of SEQ ID NO: 22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 11) CDR1 comprising the amino acid sequence of SEQ ID NO: 23, CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and CDR3 comprising the amino acid sequence of SEQ ID NO: 25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; 12) CDR1 comprising the amino acid sequence of SEQ ID NO: 26, CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and CDR3 comprising the amino acid sequence of SEQ ID NO: 28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; or 13) CDR1 comprising the amino acid sequence of SEQ ID NO:29, CDR2 comprising the amino acid sequence of SEQ ID NO:30, and CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDRs.

[0008] In another aspect, the present application provides an anti-IGFBP7 construct comprising a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in any of SEQ ID NOs: 32 to 51.

[0009] In some embodiments of any of the above anti-IGFBP7 constructs, the sdAb portion comprises the amino acid sequence of any one of SEQ ID NOs: 32-51, or a variant thereof having at least about 80% sequence identity to any one of SEQ ID NOs: 32-51.

[0010] In some embodiments according to any of the above anti-IGFBP7 constructs, the sdAb portion is camelid, chimeric, human, partially humanized, or fully humanized.

[0011] In some embodiments according to any of the above anti-IGFBP7 constructs, the sdAb portion is H It is an H antibody.

[0012] In some embodiments with any of the above anti-IGFBP7 constructs, the anti-IGFBP7 construct blocks binding of CD93 to IGFBP7. In some embodiments, the CD93 is human CD93.

[0013] In some embodiments according to any of the above anti-IGFBP7 constructs, the IGFBP7 is human IGFBP7.

[0014] In some embodiments according to any of the above anti-IGFBP7 constructs, the anti-IGFBP7 construct further comprises a second moiety. In some embodiments, the second moiety comprises an antibody moiety that specifically recognizes an antigen. In some embodiments, the antigen is PD-L1. In some embodiments, the antibody moiety is a full length antibody, Fab, Fab', (Fab')2, Fv, single chain Fv (scFv), scFv-scFv, minibody, diabody, or sdAb.

[0015] In some embodiments according to any of the above anti-IGFBP7 constructs, the anti-IGFBP7 construct further comprises a second moiety that comprises a half-life extending moiety.

[0016] In some embodiments according to any of the above anti-IGFBP7 constructs, the anti-IGFBP7 construct is an antibody-drug conjugate.

[0017] In another aspect, the present application provides an anti-IGFBP7 construct that specifically binds to IGFBP7 competitively with any one of the above anti-IGFBP7 constructs.

[0018] In another aspect, the application provides a pharmaceutical composition comprising any of the anti-IGFBP7 constructs described above and a pharma- ceutically acceptable carrier.

[0019] In another aspect, the present application provides a polynucleotide encoding a polypeptide or a portion thereof of any of the above anti-IGFBP7 constructs.

[0020] In another aspect, the application provides a nucleic acid construct comprising any of the above polynucleotides, optionally further comprising a promoter operably associated with the polynucleotide.

[0021] In another aspect, the application provides a vector comprising any of the above nucleic acid constructs.

[0022] In another aspect, the application provides an isolated host cell comprising any of the above-described polynucleotides, nucleic acid constructs or vectors.

[0023] In another aspect, the present application provides a culture medium comprising any of the above-mentioned anti-IGFBP7 constructs, polynucleotides, nucleic acid constructs, vectors or host cells.

[0024] In another aspect, the present application provides a method for producing an anti-IGFBP7 construct, the method comprising: a) culturing any of the above-described isolated host cells under conditions effective to express the polypeptide, and b) obtaining the polypeptide from the host cell.

[0025] In another aspect, the application provides a method of treating a disease or condition (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, comprising administering to an individual an effective amount of any of the above-described anti-IGFBP7 constructs or pharmaceutical compositions. In some embodiments, the disease of the condition is associated with abnormal vasculature. In some embodiments, the disease or condition is cancer. In some embodiments, the cancer is a solid tumor. In some embodiments, the cancer comprises CD93+ endothelial cells. In some embodiments, the cancer comprises IGFBP7+ blood vessels. In some embodiments, the cancer is characterized by tumor hypoxia. In some embodiments, the cancer is a locally advanced or metastatic cancer. In some embodiments, the cancer is selected from the group consisting of lymphoma, colon cancer, breast cancer, ovarian cancer, endometrial cancer, esophageal cancer, prostate cancer, cervical cancer, renal cancer, bladder cancer, gastric cancer, non-small cell lung cancer, melanoma, and pancreatic cancer. In some embodiments, the anti-IGFBP7 construct is administered parenterally to the individual. In some embodiments, the method further comprises administering a second treatment. In some embodiments, the second treatment is selected from the group consisting of surgery, radiation, gene therapy, immunotherapy, bone marrow transplant, stem cell transplant, hormone therapy, targeted therapy, cryotherapy, ultrasound therapy, photodynamic therapy, and chemotherapy. In some embodiments, the second treatment is immunotherapy. In some embodiments, the immunotherapy comprises administering an immunomodulatory agent. In some embodiments, the immunomodulatory agent is an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor comprises an anti-PD-L1 antibody or an anti-PD-1 antibody. In some embodiments, the individual is a human. [Brief description of the drawings]

[0026] [Figure 1] FIG. 1 shows anti-human IGFBP7 antibody titers in llama sera after immunization of llamas with human IGFBP7.

[0027] [Figure 2A] Figures 2A-B show the binding of various anti-IGFBP7 nanobodies to human IGFBP7-expressing (hIGFBP7) HEK293T cells. [Figure 2B] Figures 2A-B show the binding of various anti-IGFBP7 nanobodies to human IGFBP7-expressing (hIGFBP7) HEK293T cells.

[0028] [Figure 3A] Figures 3A-B show binding of various anti-IGFBP7 nanobodies to mouse IGFBP7-expressing (mIGFBP7) HEK293T cells. [Figure 3B] Figures 3A-B show binding of various anti-IGFBP7 nanobodies to mouse IGFBP7-expressing (mIGFBP7) HEK293T cells.

[0029] [Figure 4] FIG. 4 shows binding of various anti-IGFBP7 nanobody Fc fusion constructs to human IGFBP7-expressing (hIGFBP7) HEK293T cells.

[0030] [Diagram 5] FIG. 5 shows binding of various anti-IGFBP7 nanobody Fc fusion constructs to mouse IGFBP7-expressing (mIGFBP7) CHO-K1 cells.

[0031] [Figure 6] FIG. 6 shows the binding affinity of various anti-IGFBP7 Nanobody Fc fusion constructs to human IGFBP7.

[0032] [Figure 7] FIG. 7 shows the binding affinity of various anti-IGFBP7 nanobody Fc fusion constructs to mouse IGFBP7.

[0033] [Figure 8] FIG. 8 shows a schematic diagram of two anti-IGFBP7 bispecific molecules.

[0034] [Figure 9]FIG. 9 shows the binding affinity of various monospecific and bispecific anti-IGFBP7 Nanobody Fc fusion constructs to human IGFBP7.

[0035] [Figure 10] FIG. 10 shows the binding affinity of various monospecific and bispecific anti-IGFBP7 nanobody Fc fusion constructs to mouse IGFBP7.

[0036] [Figure 11] FIG. 11 shows the binding affinity of various monospecific and bispecific anti-IGFBP7 Nanobody Fc fusion constructs to cynomolgus IGFBP7.

[0037] [Figure 12A] 12A-B show binding of various monospecific and bispecific anti-IGFBP7 Nanobody Fc fusion constructs to cell surface displayed human IGFBP7. [Figure 12B] 12A-B show binding of various monospecific and bispecific anti-IGFBP7 Nanobody Fc fusion constructs to cell surface displayed human IGFBP7.

[0038] [Figure 13] FIG. 13 shows binding of various bispecific anti-IGFBP7 nanobody Fc fusion constructs to human and mouse IGFBP7 expressing HEK293T cells.

[0039] [Figure 14] FIG. 14 shows the blocking effect of the interaction between CD93 and IGFBP7 by various anti-IGFBP7 nanobodies in human CD93-expressing CHO cells.

[0040] [Figure 15A] Figures 15A-C show the blocking effect of the interaction between CD93 and IGFBP7 by various anti-IGFBP7 Nanobody Fc fusion constructs in human CD93-expressing CHO cells. [Figure 15B]Figures 15A-C show the blocking effect of the interaction between CD93 and IGFBP7 by various anti-IGFBP7 Nanobody Fc fusion constructs in human CD93-expressing CHO cells. [Figure 15C] Figures 15A-C show the blocking effect of the interaction between CD93 and IGFBP7 by various anti-IGFBP7 Nanobody Fc fusion constructs in human CD93-expressing CHO cells.

[0041] [Figure 16] FIG. 16 shows the blocking effect of the interaction between CD93 and IGFBP7 by various monospecific and bispecific anti-IGFBP7 Nanobody Fc fusion constructs in human CD93-expressing CHO cells.

[0042] [Figure 17A] 17A-B show inhibition of HUVEC tube formation by various anti-IGFBP7 nanobody Fc fusion constructs compared to controls. [Figure 17B] 17A-B show inhibition of HUVEC tube formation by various anti-IGFBP7 nanobody Fc fusion constructs compared to controls.

[0043] [Figure 18] FIG. 18 shows epitope binning of anti-IGFBP7 antibodies by Octet competition. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0044] Detailed Description of the Application The present application provides novel anti-IGFBP7 constructs that specifically bind to IGFBP7 (e.g., anti-IGFBP7 monoclonal or multispecific antibodies), methods for preparing the anti-IGFBP7 constructs, and methods for using the constructs (e.g., methods for treating a disease or condition).

[0045] I. Definition The term "antibody" is used in its broadest sense and encompasses a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), full-length antibodies and antigen-binding fragments thereof, so long as they exhibit the desired antigen-binding activity. The term "antibody portion" refers to a full-length antibody or an antigen-binding fragment thereof.

[0046] A full-length antibody contains two heavy chains and two light chains. The variable regions of the light and heavy chains are responsible for antigen binding. The variable domains of the heavy and light chains are each called "V H " and "V L ". The variable regions of both chains generally contain three highly variable loops called complementarity determining regions (CDRs) (light chain (LC) CDRs including LC-CDR1, LC-CDR2 and LC-CDR3, and heavy chain (HC) CDRs including HC-CDR1, HC-CDR2 and HC-CDR3). The CDR boundaries of the antibodies and antigen-binding fragments disclosed herein may be defined or identified by the conventions of Kabat, Chothia, or Al-Lazikani (Al-Lazikani 1997; Chothia 1985; Chothia 1987; Chothia 1989; Kabat 1987; Kabat 1991). The three CDRs of the heavy or light chain are interposed between adjacent stretches known as framework regions (FRs), which are more highly conserved than the CDRs and form a scaffold to support the hypervariable loops. The constant regions of the heavy and light chains are not involved in antigen binding, but exhibit various effector functions. Antibodies are assigned to classes based on the amino acid sequence of the constant region of their heavy chains. The five major classes or isotypes of antibodies are IgA, IgD, IgE, IgG, and IgM, characterized by the presence of α, δ, ε, γ, and μ heavy chains, respectively. Some of the major antibody classes are divided into subclasses, such as IgG1 (γ1 heavy chain), IgG2 (γ2 heavy chain), IgG3 (γ3 heavy chain), IgG4 (γ4 heavy chain), lgA1 (α1 heavy chain), or lgA2 (α2 heavy chain).

[0047] The term "antigen-binding fragment" as used herein refers to antibody fragments including, for example, diabodies, Fab, Fab', F(ab')2, Fv fragments, disulfide-stabilized Fv fragments (dsFv), (dsFv)2, bispecific dsFv (dsFv-dsFv'), disulfide-stabilized diabodies (ds diabodies), single-chain Fvs (scFv), scFv dimers (bivalent diabodies), multispecific antibodies formed from a portion of an antibody containing one or more CDRs, camelid single domain antibodies, nanobodies, domain antibodies, bivalent domain antibodies, or any other antibody fragment that binds to an antigen but does not contain a complete antibody structure. An antigen-binding fragment can bind to the same antigen that the parent antibody or parent antibody fragment (e.g., parent scFv) binds. In some embodiments, an antigen-binding fragment can contain one or more CDRs from a particular human antibody grafted onto framework regions from a different human antibody from the fragment.

[0048] The term "single domain antibody" or "sdAb" refers to a single antigen-binding polypeptide having three complementarity determining regions (CDRs). An sdAb can bind to an antigen alone, without pairing with a corresponding CDR-containing polypeptide. In some cases, sdAbs are engineered from Camelidae HCAbs, and their heavy chain variable domains are referred to herein as "V H The camelid sdAb is one of the smallest known antigen-binding antibody fragments (see, e.g., Hamers-Casterman et al., Nature 363:446-8 (1993); Greenberg et al., Nature 374:168-73 (1995); Hassanzadeh-Ghassabeh et al., Nanomedicine (Lond), 8:1013-26 (2013)).

[0049] "Fv" is the smallest antibody fragment that contains a complete antigen recognition and binding site. This fragment consists of a tightly non-covalently associated dimer of one heavy chain variable region domain and one light chain variable region domain. Folding of these two domains results in six hypervariable loops (three loops each from the heavy and light chains) that contribute amino acid residues for antigen binding and confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv containing only three CDRs specific for an antigen) has the ability to recognize and bind antigen, but with a lower affinity than the entire binding site.

[0050] A "single-chain Fv" (also abbreviated as "sFv" or "scFv") is a VFv that is linked to a single polypeptide chain. H and V L In some embodiments, the scFv polypeptide is an antibody fragment that contains an antibody domain, which enables the scFv to form the desired structure for antigen binding. H Domain and V L It further comprises a polypeptide linker between the domains. For a review of scFvs, see Plueckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds. Springer-Verlag, New York, pp. 269-315 (1994).

[0051] As used herein, the term "CDR" or "complementarity determining region" is intended to mean the discontinuous antigen-binding sites found within the variable regions of both heavy and light chain polypeptides. These specific regions were identified by Kabat et al., J.Biol.Chem.252:6609-6616(1977);Kabat et al.,USDept.of Health and Human Services, "Sequences of proteins of immunological interest"(1991);Chothia et al.,J.Mol.Biol.196:901-917(1987);Al-Lazikani B.et al.,J.Mol.Biol.,273:927-948(1997);MacCallum et al.,J.Mol.Biol.262:732-745(1996);Abhinandan and Martin,Mol.Immunol.,45:3832-3839(2008);Lefranc MPet al., Dev. Comp. Immunol., 27:55-77 (2003); and Honegger and Pluckthun, J. Mol. Biol., 309:657-670 (2001), where the definitions include overlapping or subsets of amino acid residues when compared with each other. Nevertheless, application of either definition to refer to the CDR of an antibody or grafted antibody or variant thereof is intended to be within the scope of the term as defined and used herein. The amino acid residues encompassing the CDRs defined by each of the above cited references are shown in Table 1 below for comparison. CDR prediction algorithms and interfaces are known in the art, e.g., Abhinandan and Martin, Mol. Immunol., 45:3832-3839 (2008); Ehrenmann F. et al., Nucleic Acids Res., 38:D301-D307 (2010); and Adolf-Bryfogle J. et al., Nucleic Acids Res., 43:D432-D438 (2015).The contents of the references cited in this paragraph are incorporated herein by reference in their entirety for use in this application and for possible inclusion in one or more claims hereof. [Table 1]

[0052] The phrases "Kabat variable domain residue numbering" or "Kabat amino acid position numbering" and variations thereof refer to the numbering system used for the heavy or light chain variable domains of the antibody compilation of Kabat et al., supra. Using this numbering system, the actual linear amino acid sequence may contain fewer or additional amino acids corresponding to a shortening or insertion of the FR or hypervariable region (HVR) of the variable domain. For example, a heavy chain variable domain may contain a single amino acid insertion after residue 52 of H2 (residue 52a according to Kabat) and an inserted residue after heavy chain FR residue 82 (e.g., residues 82a, 82b and 82c, etc., according to Kabat). The Kabat numbering of residues may be determined for a given antibody by alignment of the antibody's sequence with the "standard" Kabat numbering sequence at the regions of homology.

[0053] Unless otherwise indicated herein, the numbering of residues in an immunoglobulin heavy chain is that of the EU index in Kabat et al., supra. "EU index in Kabat" refers to the residue numbering of the human IgG1 EU antibody.

[0054] "Framework" or "FR" residues are those variable domain residues other than the CDR residues as herein defined.

[0055] "Humanized" forms of non-human (e.g., rodent) antibodies are chimeric antibodies that contain minimal sequence derived from the non-human antibody. In most cases, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a hypervariable region (HVR) of the recipient are replaced by residues from a hypervariable region of a non-human species (donor antibody) such as mouse, rat, rabbit or non-human primate having the desired antibody specificity, affinity, and capacity. In some instances, framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, humanized antibodies can contain residues that are not found in the recipient antibody or the donor antibody. These modifications are made to further refine antibody performance. In general, humanized antibodies contain substantially all of at least one, and typically two, variable domains, with all or substantially all of the hypervariable loops corresponding to those of a non-human immunoglobulin and all or substantially all of the FRs being those of human immunoglobulin sequences. Humanized antibodies also optionally contain at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. For further details, see Jones et al., Nature 321:522-525 (1986); Riechmann et al., Nature 332:323-329 (1988); and Presta, Curr. Op. Struct. Biol. 2:593-596 (1992).

[0056] A "human antibody" is an antibody having an amino acid sequence that corresponds to that of an antibody produced by a human and / or has been produced using any of the techniques for producing human antibodies disclosed herein. This definition of a human antibody specifically excludes humanized antibodies that contain non-human antigen-binding residues. Human antibodies can be produced using a variety of techniques known in the art, including phage display libraries. Hoogenboom and Winter, J. Mol. Biol., 227:381 (1991); Marks et al., J. Mol. Biol., 222:581 (1991). Also available for the preparation of human monoclonal antibodies are the methods described in Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, p.77 (1985); Boerner et al., J. Immunol., 147(1):86-95 (1991). See also van Dijk and van de Winkel, Curr. Opin. Pharmacol., 5:368-74 (2001). Human antibodies can be prepared by administering antigen to transgenic animals, e.g., immunized xenogeneic animals, that have been engineered to produce such antibodies in response to antigen challenge but whose endogenous loci have been disabled (see, e.g., U.S. Patent Nos. 6,075,181 and 6,150,584 for XENOMOUSE™). See also Li et al., Proc. Natl. Acad. Sci. USA, 103:3557-3562 (2006) for human antibodies generated via human B-cell hybridoma technology.

[0057] "Percent (%) amino acid sequence identity" or "homology" with respect to the polypeptide and antibody sequences identified herein is defined as the percentage of amino acid residues in a candidate sequence that are identical to the amino acid residues in the polypeptide being compared, after the sequences are aligned, taking into account any conservative substitutions as part of the sequence identity. Alignment to determine percent amino acid sequence identity can be accomplished in a variety of ways that are within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, Megalign (DNASTAR), or MUSCLE software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms required to achieve maximum alignment over the entire length of the sequences being compared. However, for purposes of this specification, % amino acid sequence identity values ​​are generated using the sequence comparison computer program MUSCLE (Edgar, RC, Nucleic Acids Research 32(5):1792-1797, 2004; Edgar, RC, BMC Bioinformatics 5(1):113, 2004.

[0058] "Homologous" refers to sequence similarity or sequence identity between two polypeptides or two nucleic acid molecules. If both positions of two compared sequences are occupied by the same base or amino acid monomer subunit, for example, if each position of two protein molecules is occupied by lysine, or each position of two DNA molecules is occupied by adenine, then the molecules are homologous at that position. The percentage of homology between two sequences is a function of the number of matching or homologous positions shared by the two sequences multiplied by 100 divided by the number of positions compared. For example, if 6 out of 10 positions in two sequences are matching or homologous, then the two sequences are 60% homologous. As an example, the protein sequences SGTSTD and TGTSDA share 50% homology. Generally, comparisons are made when the two sequences are aligned to obtain maximum homology.

[0059] The term "constant domain" refers to the portion of an immunoglobulin molecule that has a more conserved amino acid sequence compared to the other portion of the immunoglobulin, the variable domain, which contains the antigen binding site. The constant domain is the C H 1. C H 2 and C H 3 domains (collectively C H ) and light chain CHL (or C L ) domain.

[0060] The "light chains" of antibodies (immunoglobulins) from any mammalian species can be assigned to one of two clearly distinct types, called kappa ("κ") and lambda ("λ"), based on the amino acid sequences of their constant domains.

[0061] The "CH1 domain" (also called "C1" for "H1" domain) typically spans from about amino acid 118 to about amino acid 215 (EU numbering system).

[0062] The "hinge region" is generally defined as the region in IgG corresponding to Glu216-Pro230 of human IgG1 (Burton, Molec. Immunol. 22:161-206 (1985)). Hinge regions of other IgG isotypes can be aligned with the IgG1 sequence by placing the first and last cysteine ​​residues that form the inter-heavy chain S-S bonds in the same positions.

[0063] The "CH2 domain" (also called the "C2" domain) of the human IgG Fc region typically extends from about amino acid 231 to about amino acid 340. The CH2 domain is unique in that it is not closely paired with another domain. Rather, two N-linked branched carbohydrate chains are sandwiched between the two CH2 domains in intact native IgG molecules. It has been speculated that the carbohydrates may provide a surrogate for domain-domain pairing and help stabilize the CH2 domain. Burton, Molec Immunol. 22:161-206 (1985).

[0064] The "CH3 domain" (also called the "C2" domain) comprises the stretch of residues C-terminal to the CH2 domain in the Fc region (i.e., from about amino acid residue 341 to the C-terminus of the antibody sequence, typically amino acid residue 446 or 447 for IgG).

[0065] The term "Fc region" or "fragment crystallizable region" herein is used to define the C-terminal region of an immunoglobulin heavy chain, including native sequence Fc regions and variant Fc regions. Although the boundaries of an immunoglobulin heavy chain Fc region may vary, the human IgG heavy chain Fc region is usually defined to extend from the amino acid residue at position Cys226 or the amino acid residue at Pro230 to its carboxyl terminus. The C-terminal lysine (residue 447 according to the EU numbering system) of the Fc region may be removed, for example, during production or purification of the antibody, or by recombinantly engineering the nucleic acid encoding the heavy chain of the antibody. Thus, a composition of intact antibodies may include an antibody population in which all K447 residues have been removed, an antibody population in which the K447 residue has not been removed, and an antibody population having a mixture of antibodies with and without the K447 residue. Suitable native sequence Fc regions for use in the antibodies described herein include human IgG1, IgG2 (IgG2A, IgG2B), IgG3, and IgG4.

[0066] "Fc receptor" or "FcR" describes a receptor that binds to the Fc region of an antibody. A preferred FcR is a native sequence human FcR. Additionally, a preferred FcR is one that binds an IgG antibody (gamma receptor) and includes receptors of the FcγRI, FcγRII, FcRN, and FcγRIII subclasses, including allelic variants and alternatively spliced ​​forms of these receptors, with FcγRII receptors including FcγRIIA ("activating receptor") and FcγRIIB ("inhibiting receptor"), which have similar amino acid sequences that differ primarily in their cytoplasmic domains. Activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. Inhibiting receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic domain. (See M. Daeron, Annu. Rev. Immunol. 15:203-234 (1997). FcRNs are important for the recycling of antibodies to the blood allowing for increased serum half-life of the antibody. FcRs are reviewed in Ravetch and Kinet, Annu. Rev. Immunol. 9:457-92 (1991); Capel et al., Immunomethods 4:25-34 (1994); and de Haas et al., J. Lab. Clin. Med. 126:330-41 (1995). Other FcRs, including those identified in the future, are encompassed by the term "FcR" herein.

[0067] The term "epitope" as used herein refers to the particular group of atoms or amino acids on an antigen to which an antibody or antibody portion binds. Two antibodies or antibody portions may bind to the same epitope in an antigen if they exhibit competitive binding to the antigen.

[0068] As used herein, a first antibody or fragment thereof "competes" with a second antibody or fragment thereof for binding to a target antigen if the first antibody or fragment thereof inhibits target antigen binding of the second antibody or fragment thereof by at least about 50% (e.g., at least about any one of 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99%) in the presence of an equimolar concentration of the first antibody or fragment thereof, or vice versa. A high throughput process for "binning" antibodies based on cross-competition is described in PCT Publication No. WO 03 / 48731.

[0069] As used herein, the terms "specifically bind," "specifically recognize," and "specific for" refer to a measurable and reproducible interaction, such as binding, between a target and an antibody or antibody portion that determines the presence of the target in the presence of a heterogeneous population of molecules, including biological molecules. For example, an antibody or antibody portion that specifically recognizes a target (which may be an epitope) is an antibody or antibody portion that binds to this target with higher affinity, avidity, more readily, and / or with longer duration than its binding to other targets. In some embodiments, the extent of binding of an antibody to an unrelated target is less than about 10% of the binding of the antibody to the target, as measured, for example, by radioimmunoassay (RIA). In some embodiments, an antibody that specifically binds to a target has a binding affinity of ≦10%. -5 M, ≦10 -6 M, ≦10 -7 M, ≦10 -8 M, ≦10 -9 M, ≦10 -10 M, ≦10 -11 M, or ≤ 10 -12 Dissociation constant of M (K D). In some embodiments, the antibody specifically binds to an epitope on a protein that is conserved among proteins from different species. In some embodiments, specific binding can include, but does not require, exclusive binding. The binding specificity of an antibody or antigen-binding domain can be determined experimentally by methods known in the art. Such methods include, but are not limited to, Western blot, ELISA-, RIA-, ECL-, IRMA-, EIA-, BLI, SPR, BIACORE™-test and peptide scan.

[0070] A "bispecific" or "multispecific" antibody can refer to an antibody that binds to two or more different antigens or an antibody that binds to two or more different epitopes of the same antigen.

[0071] An "isolated" antibody (or construct) is one that has been identified, separated and / or recovered from a component of its production environment (e.g., natural or recombinant). Preferably, an isolated polypeptide is free of association with all other components from its production environment.

[0072] An "isolated" nucleic acid molecule encoding a construct, antibody, or antigen-binding fragment thereof described herein is a nucleic acid molecule that is identified and separated from at least one contaminating nucleic acid molecule with which it is normally associated in the environment in which it is produced. Preferably, an isolated nucleic acid is not associated with all components associated with the production environment. An isolated nucleic acid molecule encoding a polypeptide and antibody described herein is in a form other than the form or situation in which it is found in nature. Thus, an isolated nucleic acid molecule is distinct from a nucleic acid encoding a polypeptide and antibody described herein that naturally occurs in a cell. An isolated nucleic acid includes a nucleic acid molecule contained in a cell that normally contains the nucleic acid molecule, but the nucleic acid molecule is present extrachromosomally or at a chromosomal location different from its natural chromosomal location.

[0073] The term "control sequence" refers to a DNA sequence required for the expression of an operably linked coding sequence in a particular host organism. Control sequences suitable for prokaryotes include, for example, a promoter, optionally an operator sequence, and a ribosome binding site. Eukaryotic cells are known to utilize promoters, polyadenylation signals, and enhancers.

[0074] A nucleic acid is "operably linked" when it is placed into a functional relationship with another nucleic acid sequence. For example, DNA for a presequence or secretory leader is operably linked to DNA for a polypeptide if it is expressed as a preprotein that participates in the secretion of the polypeptide; a promoter or enhancer is operably linked to a coding sequence if it affects the transcription of the sequence; or a ribosome binding site is operably linked to a coding sequence if it is positioned so as to promote translation. Generally, "operably linked" means that the DNA sequences being linked are contiguous, and in the case of a secretory leader, contiguous and in reading frame. Enhancers, however, need not be contiguous. Linking is accomplished by ligation at convenient restriction sites. If such sites do not exist, synthetic oligonucleotide adaptors or linkers are used in accordance with conventional practice.

[0075] The term "vector" as used herein refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is linked. The term includes vectors as self-replicating nucleic acid structures as well as vectors integrated into the genome of a host cell into which it is introduced. Certain vectors are capable of directing the expression of nucleic acids to which they are operatively linked. Such vectors are referred to herein as "expression vectors."

[0076] The terms "transfected" or "transformed" or "transduced" as used herein refer to the process by which exogenous nucleic acid is transferred or introduced into a host cell. A "transfected" or "transformed" or "transduced" cell is one that has been transfected, transformed or transduced with exogenous nucleic acid. The cell includes the primary subject cell and its progeny.

[0077] The terms "host cell," "host cell line," and "host cell culture" are used interchangeably and refer to cells into which exogenous nucleic acid has been introduced, including the progeny of such cells. Host cells include "transformants" and "transformed cells," which include the primary transformed cell and its progeny, regardless of the number of passages. The progeny may not be completely identical in nucleic acid content to the parent cell and may contain mutations. Mutant progeny that have the same function or biological activity as screened or selected for in the originally transformed cell are included herein.

[0078] The term "immunoconjugate" includes reference to the covalent attachment of a therapeutic agent or detectable label to an antibody, such as an antibody portion described herein. The linkage can be direct or indirect, such as through a linker (such as a peptide linker).

[0079] As used herein, "treatment" or "treating" is an approach to obtain beneficial or desired results, including clinical results. For purposes of this application, beneficial or desired clinical results include, but are not limited to, one or more of the following: alleviating one or more symptoms attributable to the disease, reducing the extent of the disease, stabilizing the disease (e.g., preventing or slowing the progression of the disease (e.g., metastasis), preventing or slowing the spread of the disease (e.g., metastasis), preventing or slowing the recurrence of the disease, slowing or slowing the progression of the disease, improving the disease state, providing remission (partial or total) of the disease, reducing the dose of one or more other medications required to treat the disease, slowing the progression of the disease, improving or improving quality of life, increasing weight gain, and / or prolonging survival. Reduction in the pathological consequences of cancer (e.g., tumor volume, etc.) is also encompassed by "treatment". The methods of this application contemplate any one or more of these aspects of treatment.

[0080] In the context of cancer, the term "treating" includes any or all of inhibiting the growth of cancer cells, inhibiting the replication of cancer cells, reducing the overall tumor burden, and ameliorating one or more symptoms associated with the disease.

[0081] The term "inhibition" or "inhibiting" refers to the reduction or cessation of any phenotypic characteristic, or the reduction or cessation of the incidence, degree, or likelihood of that characteristic. "Reducing" or "inhibiting" is the reduction, reduction, or cessation of an activity, function, and / or amount compared to a reference. In certain embodiments, "reducing" or "inhibiting" refers to the ability to cause an overall reduction of 20% or more. In another embodiment, "reducing" or "inhibiting" refers to the ability to cause an overall reduction of 50% or more. In yet another embodiment, "reducing" or "inhibiting" refers to the ability to cause an overall reduction of 75%, 85%, 90%, 95%, or more.

[0082] As used herein, "reference" refers to any sample, standard, or level used for comparison purposes. Reference can be obtained from healthy and / or non-disease samples. In some examples, reference can be obtained from untreated samples. In some examples, reference is obtained from non-disease or non-treated samples of individuals. In some examples, reference is obtained from one or more healthy individuals who are not individuals or patients.

[0083] As used herein, "delaying the onset of disease" means to postpone, hinder, slow, prevent, stabilize, inhibit and / or postpone the onset of disease (such as cancer). This delay can be of various lengths of time, depending on the disease being treated and / or the individual's medical history. As will be apparent to those skilled in the art, a sufficient or significant delay can, in effect, encompass prevention, in that the individual does not develop the disease. For example, late-stage cancer, such as the onset of metastasis, can be delayed.

[0084] "Preventing" as used herein includes providing prophylaxis against the occurrence or recurrence of a disease in an individual who may have a predisposition to the disease, but has not yet been diagnosed with the disease.

[0085] As used herein, "inhibiting" a function or activity is to decrease a function or activity when compared to conditions that are otherwise the same except for the condition or parameter of interest, or when compared to another condition, For example, an antibody that inhibits tumor growth reduces the rate of tumor growth compared to the rate of tumor growth in the absence of the antibody.

[0086] The terms "subject," "individual," and "patient" are used interchangeably herein to refer to a mammal, including, but not limited to, a human, bovine, equine, feline, canine, rodent, or primate. In some embodiments, an individual is a human.

[0087] An "effective amount" of an agent refers to an amount effective at the dosage and duration required to achieve the desired therapeutic or prophylactic result. The specific dosage may vary depending on one or more of the specific agent selected, the dosing regimen to be followed, whether it is administered in combination with other compounds, the timing of administration, the tissue to be imaged, and the physical delivery system in which it is carried.

[0088] The terms "pharmaceutical formulation" and "pharmaceutical composition" refer to a preparation that is in a form that allows the biological activity of the active ingredient(s) to be effective and does not contain additional components that are unacceptably toxic to the individual to whom the formulation is administered. Such formulations may be sterile.

[0089] "Pharmaceutically acceptable carrier" refers to a non-toxic solid, semi-solid or liquid filler, diluent, encapsulating material, formulation aid or carrier conventional in the art for use with therapeutic agents that together comprise a "pharmaceutical composition" for administration to an individual. A pharmaceutically acceptable carrier is non-toxic to a recipient at the dosage and concentration used and is compatible with other ingredients of the formulation. A pharmaceutically acceptable carrier is suitable for the formulation used.

[0090] A "sterile" preparation is sterile or essentially free of living microorganisms and their spores.

[0091] Administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive or sequential administration in any order.

[0092] The term "concurrently" is used herein to refer to the administration of two or more therapeutic agents where at least a portion of the administration overlaps in time or where the administration of one therapeutic agent falls within a short period of time compared to the administration of the other therapeutic agent. For example, the two or more therapeutic agents are administered within a time interval of about 60 minutes or less, e.g., about any of 30, 15, 10, 5, or 1 minutes or less.

[0093] The term "sequentially" is used herein to refer to administration of two or more therapeutic agents, where administration of one or more agents is continued after discontinuing administration of the other agent or agents. For example, administration of the two or more therapeutic agents is administered at intervals of more than about 15 minutes, e.g., about 20, 30, 40, 50, or 60 minutes, one day, two days, three days, one week, two weeks, or one month or more.

[0094] As used herein, "in combination with" refers to the administration of one treatment modality in addition to another treatment modality. Thus, "in combination with" refers to the administration of one treatment modality before, during or after the administration of another treatment modality to an individual.

[0095] The term "package insert" is used to refer to instructions typically included in commercial packaging of a therapeutic product that contain information regarding the indications, usage, dosage, administration, concomitant therapy, contraindications and / or warnings for the use of such therapeutic product.

[0096] An "article of manufacture" is any article of manufacture (e.g., package or container) or kit that contains at least one reagent, such as a pharmaceutical agent for treating a disease or disorder (e.g., cancer), or a probe for specifically detecting a biomarker described herein. In certain embodiments, the article of manufacture or kit is promoted, distributed, or sold as a unit for performing the methods described herein.

[0097] It is understood that embodiments of the present application described herein include "consisting of" and / or "consisting essentially of."

[0098] Reference herein to "about" a value or parameter includes (and accounts for) the variation that is directed to the value or parameter itself. For example, a statement of "about X" includes the statement "X."

[0099] As used herein, reference to a value or parameter "is not" generally means and describes a value or parameter "other than." For example, the method is not used to treat cancer of type X means that the method is used to treat cancers other than type X.

[0100] Here, "about X to Y" is synonymous with "about X to about Y."

[0101] As used in this specification and the appended claims, the singular forms "a," "or," and "the" include plural referents unless the context clearly dictates otherwise.

[0102] II. Anti-IGFBP7 Constructs The present application provides anti-IGFBP7 constructs comprising an anti-IGFBP7 antibody portion that specifically binds to IGFBP7 as described herein.

[0103] IGFBP7 Insulin-like growth factor (IGF) binding protein (IGFBP) 7, also known as Mac25, IGFBP-rp1, tumor-derived adhesion factor (TAF), prostacyclin-stimulating factor (PSF), and angiomodulin (AGM), is a secreted extracellular matrix (ECM) protein that belongs to the IGFBP family. See Hwa et al. Endocr Rev. 1999;20(6):761-87; Bach et al. Endocrinology. 2018;159(2):570-8. Members of the IGFBP family contain an IGF-binding (IB) domain at the N-terminus that binds to IGF1 and helps regulate the bioavailability of IGF1 in the blood. IGFBP7 lacks the C-terminal domain that functions to stabilize IGF1 binding, and therefore its affinity for IGF-1 is significantly lower than that of IGFBPs 1-6. See Oh et al. J Biol Chem. 1996;271(48):30322-5. IGFBP7 was found to be expressed in many normal tissues and cancer cells; however, the exact role of IGFBP7 in cancer has been controversial. On the other hand, IGFBP7 was shown to be released from cancer cells and act as a tumor suppressor that induces tumor apoptosis and inhibits angiogenesis (Wajapeyee et al. Cell. 2008;132(3):363-74). IGF1R was proposed as the receptor, and upon IGFBP7 binding, the interaction between IGF-1 and IGF1R was blocked, inhibiting the proliferation and aggressiveness of cancer stem-like cells. See Cao et al. Cancer Cell. 2017;31(1):110-26; Evdokimova et al. Sci Signal. 2012;5(255):ra92. Administration of IGFBP7 inhibited tumor growth in vivo, and IGFBP7- / - mice were susceptible to diethylnitrosamine-induced hepatocarcinogenesis. See Akiel et al., Cancer Res. 2017;77(15):4014-25; Darr et al. Oncogene. 2014;33(23):3024-32.On the other hand, IGFBP7 has been shown to be upregulated in blood vessels of cancer tissues and can promote angiogenesis (48,64). See Komiya et al. Cancer Med. 2014; 3(3): 537-49; Pen et al. Oncogene. 2008; 27(54): 6834-44. IGFBP7 can be strongly induced by VEGF in vascular ECs, and a synergistic effect between IGFBP7 and VEGF in angiogenesis has been reported. See Komiya et al. Cancer Med. 2014; 3(3): 537-49; Hooper et al. Circ Res. 2009; 105(2): 201-8. Each of the above-listed references is incorporated by reference in its entirety for all purposes.

[0104] The human IGFBP7 gene is located at 4q12 and encodes a polypeptide. One isoform of the polypeptide has 264 amino acid residues (SEQ ID NO:111, including a signal peptide domain (residues 1-26 of SEQ ID NO:111), an insulin-binding domain (IB domain, residues 28-106 of SEQ ID NO:111), a Kazal-like domain (residues 105-158 of SEQ ID NO:111), and an Ig-like C2-type domain (residues 160-264 of SEQ ID NO:111)).

[0105] Anti-IGFBP7 antibody part In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, wherein the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, wherein the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0106] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprises a single domain antibody (sdAb) comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 1, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0107] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:1, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113.

[0108] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:33.

[0109] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:33, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:33.

[0110] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, wherein the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, wherein the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:2 or 112, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0111] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprises a single domain antibody (sdAb) comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:2 or 112, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0112] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:2 or 112, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113.

[0113] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:32.

[0114] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:32, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:32.

[0115] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:46.

[0116] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:46, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:46.

[0117] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:47.

[0118] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:47, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:47.

[0119] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:48.

[0120] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:48, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:48.

[0121] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, wherein the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, wherein the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0122] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, and the antibody portion comprises a single domain antibody (sdAb) comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0123] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116.

[0124] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:34.

[0125] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:34, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:34.

[0126] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:49.

[0127] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:49, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:49.

[0128] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:50.

[0129] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:50, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:50.

[0130] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:51.

[0131] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:51, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:51.

[0132] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 8, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0133] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO:8, CDR2 comprising the amino acid sequence of SEQ ID NO:12, and CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0134] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:8, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0135] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:35.

[0136] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:35, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:35.

[0137] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 9, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0138] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO:9, CDR2 comprising the amino acid sequence of SEQ ID NO:12, and CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0139] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0140] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:36.

[0141] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:36, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:36.

[0142] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:37.

[0143] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:37, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:37.

[0144] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0145] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 10, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0146] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 10, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13.

[0147] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:38.

[0148] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:38, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:38.

[0149] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0150] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 11, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0151] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:11, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0152] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:39.

[0153] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:39, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:39.

[0154] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0155] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 14, CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0156] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16.

[0157] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:40.

[0158] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:40, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:40.

[0159] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0160] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 17, CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0161] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19.

[0162] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:41.

[0163] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:41, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:41.

[0164] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 20, a CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 22. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0165] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO:20, CDR2 comprising the amino acid sequence of SEQ ID NO:21, and CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0166] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22.

[0167] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:42.

[0168] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:42, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:42.

[0169] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 23, a CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 25. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0170] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO:23, CDR2 comprising the amino acid sequence of SEQ ID NO:24, and CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0171] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25.

[0172] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:43.

[0173] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:43, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:43.

[0174] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, where the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, where the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0175] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO:26, CDR2 comprising the amino acid sequence of SEQ ID NO:27, and CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0176] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28.

[0177] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:44.

[0178] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:44, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:44.

[0179] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, and the antibody portion competes for binding to IGFBP7 with an antibody or antibody fragment that comprises a single domain antibody (sdAb) portion, and the sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31. In some embodiments, the antibody portion is a single domain antibody (sdAb) portion.

[0180] In some embodiments, the anti-IGFBP7 construct comprises an antibody portion that specifically recognizes IGFBP7, the antibody portion comprising a single domain antibody (sdAb) comprising CDR1 comprising the amino acid sequence of SEQ ID NO:29, CDR2 comprising the amino acid sequence of SEQ ID NO:30, and CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0181] In some embodiments, the anti-IGFBP7 construct comprises a humanized antibody portion derived from a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0182] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, wherein the sdAb portion comprises the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in SEQ ID NO:45.

[0183] In some embodiments, the sdAb portion comprises the amino acid sequence of SEQ ID NO:45, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity to SEQ ID NO:45.

[0184] In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, the sdAb portion comprising a) a CDR1 comprising the amino acid sequence INTYL (SEQ ID NO:52), b) a CDR2 comprising the amino acid sequence AITSGGSINYADSVKG (SEQ ID NO:12), and c) a CDR3 comprising the amino acid sequence KAHPNPWGFDNDY (SEQ ID NO:13). In some embodiments, the anti-IGFBP7 construct comprises a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, the sdAb portion comprising a) a CDR1 comprising the amino acid sequence AX 1 CDR1 (wherein X 1 =N or I), b) a CDR2 comprising the amino acid sequence AITSGGSINYADSVKG (SEQ ID NO: 12), and c) a CDR3 comprising the amino acid sequence KAHPNPWGFDNDY (SEQ ID NO: 13).

[0185] In some embodiments, the anti-IGFBP7 sdAb portion is camelid, chimeric, human, partially humanized, or fully humanized.

[0186] In some embodiments, the anti-IGFBP7 sdAb portion is H It is an H antibody.

[0187] In some embodiments, the anti-IGFBP7 construct comprises or is an anti-IGFBP7 fusion protein.

[0188] In some embodiments, the anti-IGFBP7 construct comprises or is a multispecific anti-IGFBP7 construct (such as a bispecific antibody).

[0189] In some embodiments, the anti-IGFBP7 construct comprises or is an anti-IGFBP7 immunoconjugate.

[0190] In some embodiments, the anti-IGFBP7 construct blocks the binding of IGFBP7 and CD93. In some embodiments, the CD93 is human CD93. In some embodiments, the IGFBP7 is human IGFBP7. In some embodiments, the binding of IGFBP7 to CD93 is blocked by at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more after pre-incubation of the anti-IGFBP7 antibody with IGFBP7 or IGFBP7-expressing cells. In some embodiments, the dose of the anti-IGFBP7 antibody and IGFBP7 is in a ratio of about 1:10, 1:6, 1:3, 1:1.5, 1:1, 4:3, 2:1 or 5:1. In some embodiments, binding of IGFBP7 to CD93 is blocked by at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more following pre-incubation with an anti-IGFBP7 antibody at a concentration of about 50 μg / ml, 25 μg / ml, 10 μg / ml, 5 μg / ml, 2 μg / ml, 1 μg / ml, 0.8 μg / ml, 0.6 μg / ml or 0.4 μg / ml.

[0191] In some embodiments, the anti-IGFBP7 construct blocks the binding of IGFBP7 and MMRN2 to CD93. In some embodiments, the MMRN2 is human MMRN2. In some embodiments, the IGFBP7 is human IGFBP7. In some embodiments, the binding of IGFBP7 to MMRN2 is blocked by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more after pre-incubation of the anti-IGFBP7 antibody with IGFBP7 or IGFBP7-expressing cells. In some embodiments, the anti-IGFBP7 construct does not block the binding of IGFBP7 and MMRN2.

[0192] In some embodiments, the IGFBP7 is human IGFBP7. In some embodiments, the IGFBP7 is mouse IGFBP7. In some embodiments, the IGFBP7 is cynomolgus IGFBP7.

[0193] In some embodiments, the anti-IGFBP7 antibody portion binds to both human IGFBP7 and cynomolgus IGFBP7. In some embodiments, the anti-IGFBP7 antibody portion binds to both human IGFBP7 and mouse IGFBP7. In some embodiments, the anti-IGFBP7 antibody portion does not bind to cynomolgus IGFBP7 and / or mouse IGFBP7.

[0194] Anti-IGFBP7 constructs comprising at least two antibody moieties that specifically recognize IGFBP7 The present application also provides an anti-IGFBP7 construct comprising two or more antibody moieties, at least two of the two or more antibody moieties specifically recognizing IGFBP7.

[0195] In some embodiments, the at least two antibody moieties are different. In some embodiments, the at least two antibody moieties bind to two different epitopes of IGFBP7. In some embodiments, the at least two antibody moieties bind to the same IGFBP7 epitope.

[0196] In some embodiments, the anti-IGFBP7 construct comprises a tetrameric fusion protein comprising at least two different anti-IGFBP7 antibody moieties (e.g., two different VHH domains, e.g., A1 and D4). In some embodiments, the two different anti-IGFBP7 antibody moieties bind to two different epitopes of IGFBP7. In some embodiments, the fusion protein comprises an Fc fragment.

[0197] In some embodiments, at least two of the antibody moieties are the same.

[0198] In some embodiments, the at least two antibody moieties are both single domain antibody moieties (such as any of the sdAb moieties described herein).

[0199] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116.

[0200] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 9, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0201] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0202] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:14, a CDR2 comprising the amino acid sequence of SEQ ID NO:15, and a CDR3 comprising the amino acid sequence of SEQ ID NO:16.

[0203] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:17, a CDR2 comprising the amino acid sequence of SEQ ID NO:18, and a CDR3 comprising the amino acid sequence of SEQ ID NO:19.

[0204] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22.

[0205] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25.

[0206] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28.

[0207] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0208] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:1, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4.

[0209] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0210] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0211] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:14, a CDR2 comprising the amino acid sequence of SEQ ID NO:15, and a CDR3 comprising the amino acid sequence of SEQ ID NO:16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:14, a CDR2 comprising the amino acid sequence of SEQ ID NO:15, and a CDR3 comprising the amino acid sequence of SEQ ID NO:16.

[0212] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:17, a CDR2 comprising the amino acid sequence of SEQ ID NO:18, and a CDR3 comprising the amino acid sequence of SEQ ID NO:19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:17, a CDR2 comprising the amino acid sequence of SEQ ID NO:18, and a CDR3 comprising the amino acid sequence of SEQ ID NO:19.

[0213] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22.

[0214] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25.

[0215] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28.

[0216] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0217] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 9, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13.

[0218] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:14, a CDR2 comprising the amino acid sequence of SEQ ID NO:15, and a CDR3 comprising the amino acid sequence of SEQ ID NO:16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:14, a CDR2 comprising the amino acid sequence of SEQ ID NO:15, and a CDR3 comprising the amino acid sequence of SEQ ID NO:16.

[0219] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:17, a CDR2 comprising the amino acid sequence of SEQ ID NO:18, and a CDR3 comprising the amino acid sequence of SEQ ID NO:19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:17, a CDR2 comprising the amino acid sequence of SEQ ID NO:18, and a CDR3 comprising the amino acid sequence of SEQ ID NO:19.

[0220] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22.

[0221] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25.

[0222] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28.

[0223] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0224] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16.

[0225] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19.

[0226] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 20, a CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 20, a CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 22.

[0227] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 23, a CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 23, a CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 25.

[0228] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28.

[0229] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31.

[0230] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19.

[0231] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 20, a CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 20, a CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 22.

[0232] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 23, a CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 23, a CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 25.

[0233] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28.

[0234] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 14, a CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 16; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31.

[0235] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 20, a CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO: 17, CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and CDR3 comprising the amino acid sequence of SEQ ID NO: 19; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO: 20, CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and CDR3 comprising the amino acid sequence of SEQ ID NO: 22.

[0236] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 23, a CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO: 17, CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and CDR3 comprising the amino acid sequence of SEQ ID NO: 19; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO: 23, CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and CDR3 comprising the amino acid sequence of SEQ ID NO: 25.

[0237] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 26, a CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 28.

[0238] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 17, a CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 19; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 29, a CDR2 comprising the amino acid sequence of SEQ ID NO: 30, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 31.

[0239] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25.

[0240] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:20, CDR2 comprising the amino acid sequence of SEQ ID NO:21, and CDR3 comprising the amino acid sequence of SEQ ID NO:22; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:26, CDR2 comprising the amino acid sequence of SEQ ID NO:27, and CDR3 comprising the amino acid sequence of SEQ ID NO:28.

[0241] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:20, a CDR2 comprising the amino acid sequence of SEQ ID NO:21, and a CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:20, CDR2 comprising the amino acid sequence of SEQ ID NO:21, and CDR3 comprising the amino acid sequence of SEQ ID NO:22; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:29, CDR2 comprising the amino acid sequence of SEQ ID NO:30, and CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0242] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:23, CDR2 comprising the amino acid sequence of SEQ ID NO:24, and CDR3 comprising the amino acid sequence of SEQ ID NO:25; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:26, CDR2 comprising the amino acid sequence of SEQ ID NO:27, and CDR3 comprising the amino acid sequence of SEQ ID NO:28.

[0243] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:23, a CDR2 comprising the amino acid sequence of SEQ ID NO:24, and a CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:23, CDR2 comprising the amino acid sequence of SEQ ID NO:24, and CDR3 comprising the amino acid sequence of SEQ ID NO:25; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:29, CDR2 comprising the amino acid sequence of SEQ ID NO:30, and CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0244] In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:26, a CDR2 comprising the amino acid sequence of SEQ ID NO:27, and a CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs; and the second sdAb portion comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, an anti-IGFBP7 antibody construct is provided comprising: a) a first anti-IGFBP7 antibody portion comprising a first single domain antibody (sdAb) portion; and b) a second anti-IGFBP7 antibody portion comprising a second sdAb portion, wherein the first sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:26, CDR2 comprising the amino acid sequence of SEQ ID NO:27, and CDR3 comprising the amino acid sequence of SEQ ID NO:28; and the second sdAb portion comprises CDR1 comprising the amino acid sequence of SEQ ID NO:29, CDR2 comprising the amino acid sequence of SEQ ID NO:30, and CDR3 comprising the amino acid sequence of SEQ ID NO:31.

[0245] Two antibody moieties (e.g., two sdAbs) described herein can be fused to each other in any manner that allows for the appropriate properties of both moieties (e.g., binding to IGFBP7). In some embodiments, a first anti-IGFBP7 antibody moiety is fused (e.g., directly fused) to the N-terminus of a second anti-IGFBP7 antibody moiety. In some embodiments, a first anti-IGFBP7 antibody moiety is fused (e.g., directly fused) to the C-terminus of a second anti-IGFBP7 antibody moiety. In some embodiments, a construct comprises an Fc fragment comprising a first Fc domain and a second Fc domain, each Fc domain comprising a CH2 domain and a CH3 domain. In some embodiments, both antibody moieties are fused to the N-terminus of the two Fc domains. See, e.g., FIG. 8. In some embodiments, both antibody moieties are fused to the C-terminus of the two Fc domains. In some embodiments, a first anti-IGFBP7 antibody moiety is fused to the N-terminus of both Fc domains and a second anti-IGFBP7 antibody moiety is fused to the C-terminus of both Fc domains. In some embodiments, the first anti-IGFBP7 antibody moiety is fused to the C-terminus of both Fc domains, and the second anti-IGFBP7 antibody moiety is fused to the N-terminus of both Fc domains. In some embodiments, the first or second antibody moiety is fused to the C-terminus of one or both Fc domains via a linker (e.g., a GS linker). In some embodiments, the first or second antibody moiety is fused to the N-terminus of one or both Fc domains via a linker (e.g., a GS linker). In some embodiments, the Fc domain comprises a hinge region. In some embodiments, the Fc domain has a mutation that results in reduced effector function.

[0246] In some embodiments the construct comprises a first polypeptide and a second polypeptide, both polypeptides each comprising a) a first single domain antibody (sdAb) portion, b) a second sdAb portion, and c) an Fc domain, the two Fc domains forming an Fc fragment, the first sdAb comprising i) a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, the second sdAb portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO: 6, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 7, 115 or 116, the first sdAb is fused via its C-terminus directly to the N-terminus of the second sdAb and the second sdAb is fused via its C-terminus, optionally via a linker (e.g. a GS linker), to the N-terminus of both Fc domains.

[0247] In some embodiments the construct comprises a first polypeptide and a second polypeptide, both polypeptides each comprising a) a first single domain antibody (sdAb) portion, b) a second sdAb portion, and c) an Fc domain, the two Fc domains forming an Fc fragment, the first sdAb comprising i) a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, the second sdAb portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO: 6, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 7, 115 or 116, the first sdAb being fused via its C-terminus to the N-terminus of both Fc domains and the second sdAb being fused via its N-terminus, optionally via a linker (e.g. a GS linker), to the C-terminus of both Fc domains.

[0248] In some embodiments, the first or second anti-IGFBP7 antibody portion is a humanized antibody portion derived from any of the antibody portions described in this section.

[0249] In some embodiments, the first or second anti-IGFBP7 antibody portion is an antibody portion that competes with any of the antibody portions described in this section for a binding epitope of IGFBP7.

[0250] Multispecific anti-IGFBP7 constructs that bind to a second agent In some embodiments, the anti-IGFBP7 construct comprises a multispecific (e.g., bispecific) anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any one of the anti-IGFBP7 antibody portions described herein), wherein a second binding portion (such as a second antibody portion) specifically recognizes a second antigen distinct from IGFBP7. In some embodiments, the second antigen is an immune checkpoint molecule. In some embodiments, the second antigen is PD-1 or PD-L1. In some embodiments, the second antigen is a tumor antigen. In some embodiments, the second antigen is an angiogenic agent. In some embodiments, the angiogenic agent is VEGF (e.g., human VEGF). In some embodiments, the angiogenic agent is a VEGF receptor. In some embodiments, the angiogenic agent is VEGFR1 (e.g., human VEGFR1) or a fragment thereof. In some embodiments, the angiogenic agent is VEGFR2 (e.g., human VEGFR2) or a fragment thereof.

[0251] In some embodiments, the second binding moiety is fused to the anti-IGFBP7 antibody moiety via a linker (eg, any of the linkers described herein).

[0252] In some embodiments, the second antibody portion is a full length antibody, a Fab, a Fab', a (Fab') 2 , Fv, single chain Fv (scFv), scFv-scFv, minibody, diabody, nanobody, or sdAb.

[0253] In some embodiments, the anti-IGFBP7 construct is a multispecific (e.g., bispecific) anti-IGFBP7 construct comprising: a) an anti-IGFBP7 antibody portion according to any one of the anti-IGFBP7 antibody portions described herein; and b) a second antibody portion that specifically recognizes PD-L1 (an anti-PD-L1 antibody portion, such as any of those described herein) or a second antibody portion that specifically recognizes PD-1 (an anti-PD-1 antibody portion, such as any of those described herein).

[0254] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 or anti-PD-1 antibody portion comprising a full-length antibody comprising two heavy chains and two light chains, each of the two heavy chains comprising a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L a) an anti-PD-L1 or anti-PD-1 antibody portion comprising: a) an anti-IGFBP7 antibody portion comprising: a) an anti-IGFBP7 antibody portion comprising: a) an anti-IGFBP7 antibody portion comprising: a) an anti-IGFBP7 antibody portion comprising: a) an anti-IGFBP7 antibody portion comprising:

[0255] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 or anti-PD-1 antibody portion comprising a full-length antibody comprising two heavy chains and two light chains, each of the two heavy chains comprising a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L a) an anti-PD-L1 or anti-PD-1 antibody portion comprising: a) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to at least one or both of the light chains of an anti-PD-L1 or anti-PD-1 full antibody. In some embodiments, the anti-IGFBP7 antibody portion is fused to the N-terminus of both light chains. In some embodiments, the anti-IGFBP7 antibody portion is fused to the C-terminus of both light chains.

[0256] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0257] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, CDR2 comprising the amino acid sequence of SEQ ID NO:6, and CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 single amino acid substitution in the CDRs.

[0258] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO:9, CDR2 comprising the amino acid sequence of SEQ ID NO:12, and CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0259] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0260] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 14, CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0261] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 17, CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0262] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO:20, CDR2 comprising the amino acid sequence of SEQ ID NO:21, and CDR3 comprising the amino acid sequence of SEQ ID NO:22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0263] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO:23, CDR2 comprising the amino acid sequence of SEQ ID NO:24, and CDR3 comprising the amino acid sequence of SEQ ID NO:25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0264] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO:26, CDR2 comprising the amino acid sequence of SEQ ID NO:27, and CDR3 comprising the amino acid sequence of SEQ ID NO:28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0265] In some embodiments, the anti-IGFBP7 construct comprises: a) an anti-PD-L1 antibody portion or an anti-PD-1 antibody portion that comprises a full-length antibody comprising two heavy chains and two light chains, wherein each of the two heavy chains comprises a heavy chain variable region (V H ), and each of the two light chains contains a light chain variable region (V L ), and b) an anti-IGFBP7 antibody portion (such as any of the antibody portions described herein) fused to the anti-PD-L1 antibody portion or full anti-PD-1 antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising CDR1 comprising the amino acid sequence of SEQ ID NO:29, CDR2 comprising the amino acid sequence of SEQ ID NO:30, and CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs.

[0266] In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0267] In some embodiments, an anti-IGFBP7 construct is provided that comprises a) an anti-IGFBP7 antibody portion and b) a binding portion that specifically recognizes VEGF (e.g., human VEGF) or VEGFR (e.g., human VEGFR1, e.g., human VEGFR2). In some embodiments, the binding portion that specifically recognizes VEGF or VEGFR comprises the amino acid sequence of SEQ ID NO: 98. In some embodiments, the anti-IGFBP7 sdAb comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the anti-IGFBP7 sdAb is fused to the binding portion that specifically recognizes VEGF or VEGFR via a linker or a constant region of an immunoglobulin (such as human IgG).

[0268] In some embodiments, an anti-IGFBP7 construct is provided that comprises a) an anti-IGFBP7 antibody portion and b) a binding portion that specifically recognizes VEGF (e.g., human VEGF) or VEGFR (e.g., human VEGFR1, e.g., human VEGFR2). In some embodiments, the binding portion that specifically recognizes VEGF or VEGFR comprises the amino acid sequence of SEQ ID NO: 98. In some embodiments, the anti-IGFBP7 sdAb comprises a CDR1 that comprises the amino acid sequence of SEQ ID NO: 5 or 114, a CDR2 that comprises the amino acid sequence of SEQ ID NO: 6, and a CDR3 that comprises the amino acid sequence of SEQ ID NO: 7, 115 or 116, or a variant thereof that comprises up to 5, 4, 3, 2, or 1 amino acid substitution in the CDR. In some embodiments, the anti-IGFBP7 sdAb is fused to the binding portion that specifically recognizes VEGF or VEGFR via a linker or a constant region of an immunoglobulin (such as human IgG).

[0269] In some embodiments, an anti-IGFBP7 construct is provided that comprises: a) an anti-VEGF (e.g., anti-human VEGF) or anti-VEGFR (e.g., anti-human VEGFR1, e.g., anti-human VEGFR2) full-length antibody comprising two heavy chains and two light chains; and b) an anti-IGFBP7 single domain antibody (sdAb), wherein the anti-IGFBP7 sdAb is fused to one or both of the heavy and / or light chains of the full-length antibody. In some embodiments, the anti-IGFBP7 sdAb is fused to the C-terminus of both heavy chains of the full-length antibody. In some embodiments, the anti-IGFBP7 sdAb is fused to the N-terminus of both heavy chains of the full-length antibody. In some embodiments, the anti-IGFBP7 sdAb is fused to the C-terminus of both light chains of the full-length antibody. In some embodiments, the anti-IGFBP7 sdAb is fused to the N-terminus of both light chains of the full-length antibody. In some embodiments, the anti-IGFBP7 sdAb comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the anti-IGFBP7 sdAb comprises a CDR1 comprising the amino acid sequence of SEQ ID NO: 5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO: 6, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 7, 115, or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the full length antibody comprises a heavy chain variable region (V H ) and the light chain variable region (V L ) and specifically recognizes VEGF, H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 99, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 100, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 101, L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 102, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO: 103, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO: 104. In some embodiments, the full length antibody comprises a heavy chain variable region (V H ) and the light chain variable region (V L) and specifically recognizes VEGFR2, H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 108, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO: 109, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO: 110. In some embodiments, the anti-IGFBP7 sdAb is fused to the full-length antibody via a linker (such as any of the linkers described above).

[0270] In some embodiments, the second binding (e.g., second antibody moiety) and the anti-IGFBP7 antibody moiety are fused to each other via a linker, such as any linker described herein, in any operable form that allows for proper function of the binding moieties. In some embodiments, the linker is a GS linker. In some embodiments, the linker is selected from the group consisting of SEQ ID NOs: 54-61.

[0271] Exemplary Anti-PD-L1 Antibody Moieties Exemplary anti-PD-L1 antibody portions include, but are not limited to, those described in WO2019228514A1, WO2019227490A1, and WO2020019232A1.

[0272] In some embodiments, the anti-PD-L1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), the antibody portion comprising a second heavy chain variable region (V H-2 ) and a second light chain variable region (V L-2 ) and competes with an antibody or antibody fragment containing V H-2 comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 62, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 63, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 64, L-2comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO:65, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO:66, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO:67.

[0273] In some embodiments, the anti-PD-L1 moiety has a sequence as set forth in SEQ ID NO: 68, 69, or 70. H HC-CDR1, HC-CDR2 and HC-CDR3, each of which contains the amino acid sequences of CDR1, CDR2 and CDR3 within the chain region, and V having the sequence shown in SEQ ID NO: 71, 72 or 73. L The chain regions include LC-CDR1, LC-CDR2 and LC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively.

[0274] In some embodiments, the anti-PD-L1 antibody portion (such as an scFv) used in a multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), including a) V H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 62, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 63, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 64, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the HC-CDRs; b) V L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 65, an LC-CDR2 comprising the amino acid sequence of any one of SEQ ID NO: 66, and an LC-CDR3 comprising the amino acid sequence of any one of SEQ ID NO: 67, or a variant thereof comprising a total of up to about 5, 4, 3, 2, or 1 amino acid substitutions in the LC-CDRs.

[0275] In some embodiments, V H includes variants comprising an amino acid sequence of SEQ ID NO: 68, 69, or 70, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity; LIn some embodiments, V includes variants comprising an amino acid sequence of SEQ ID NO: 71, 72, or 73, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity. H includes a variant comprising an amino acid sequence of SEQ ID NO:68, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity; L In some embodiments, V includes a variant comprising an amino acid sequence of SEQ ID NO: 71, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity. H includes a variant comprising an amino acid sequence of SEQ ID NO:69, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity; L In some embodiments, V includes a variant comprising an amino acid sequence of SEQ ID NO: 72, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity. H includes a variant comprising an amino acid sequence of SEQ ID NO: 70, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity; L includes variants comprising an amino acid sequence of SEQ ID NO: 73, or an amino acid sequence having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0276] Exemplary Anti-PD-1 Antibody Moieties Exemplary anti-PD-1 antibody portions include, but are not limited to, those described in WO2018133842 and WO2018133837.

[0277] In some embodiments, the anti-PD-1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), the antibody portion comprising a second heavy chain variable region (V H-2 ) and a second light chain variable region (V L-2 ) and competes with an antibody or antibody fragment containing V H-2 comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 74, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 75, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 76, L-2 comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO:77, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO:78, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO:79.

[0278] In some embodiments, the anti-PD-1 moiety has the sequence set forth in SEQ ID NO:92. H HC-CDR1, HC-CDR2 and HC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively, within the chain region, and V having the sequence shown in SEQ ID NO:93 L The chain regions include LC-CDR1, LC-CDR2 and LC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively.

[0279] In some embodiments, the anti-PD-1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), including a) V Hcomprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 74, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 75, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 76, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the HC-CDRs; b) V L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 77, an LC-CDR2 comprising the amino acid sequence of any one of SEQ ID NO: 78, and an LC-CDR3 comprising the amino acid sequence of any one of SEQ ID NO: 79, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the LC-CDRs. In some embodiments, the amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0280] In some embodiments, the second antibody portion comprises a heavy chain variable region (V H ) and a light chain variable region (V L ) and

[0281] In some embodiments, the anti-PD-1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), the antibody portion comprising a second heavy chain variable region (V H-2 ) and a second light chain variable region (V L-2 ) and competes with an antibody or antibody fragment containing V H-2 comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 80, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 81, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 82, L-2 comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO:83, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO:84, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO:85.

[0282] In some embodiments, the anti-PD-1 moiety has the sequence set forth in SEQ ID NO:94. H HC-CDR1, HC-CDR2 and HC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively, within the chain region, and V having the sequence shown in SEQ ID NO:95 L The chain regions include LC-CDR1, LC-CDR2 and LC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively.

[0283] In some embodiments, the anti-PD-1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), including a) V H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 80, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 81, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 82, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the HC-CDRs; b) V L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 83, an LC-CDR2 comprising the amino acid sequence of any one of SEQ ID NO: 84, and an LC-CDR3 comprising the amino acid sequence of any one of SEQ ID NO: 85, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the LC-CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0284] In some embodiments, the anti-PD-1 antibody portion comprises a heavy chain variable region (V) comprising the amino acid sequence set forth in SEQ ID NO:94. H ) and a light chain variable region (V L ) and

[0285] In some embodiments, the anti-PD-1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), the antibody portion comprising a second heavy chain variable region (V H-2 ) and a second light chain variable region (V L-2 ) and competes with an antibody or antibody fragment containing V H-2 comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 86, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 87, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 88; L-2 comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO:89, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO:90, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO:91.

[0286] In some embodiments, the anti-PD-1 moiety has the sequence set forth in SEQ ID NO:96. H HC-CDR1, HC-CDR2 and HC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively, within the chain region, and V having the sequence shown in SEQ ID NO:97 L The chain regions include LC-CDR1, LC-CDR2 and LC-CDR3, which contain the amino acid sequences of CDR1, CDR2 and CDR3, respectively.

[0287] In some embodiments, the anti-PD-1 antibody portion (such as an scFv) used in the multispecific anti-IGFBP7 construct comprises a heavy chain variable region (V H ) and the light chain variable region (V L ), including a) V H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 86, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 87, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 88, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the HC-CDRs; b) V Lcomprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 89, an LC-CDR2 comprising the amino acid sequence of any one of SEQ ID NO: 90, and an LC-CDR3 comprising the amino acid sequence of any one of SEQ ID NO: 91, or a variant thereof comprising up to about 5, 4, 3, 2, or 1 total amino acid substitutions in the LC-CDRs. In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0288] In some embodiments, the second antibody portion comprises a heavy chain variable region (V H ) and a light chain variable region (V L ) and

[0289] Exemplary Binding Moieties that Specifically Recognize VEGF Exemplary binding moieties that specifically recognize VEGF include, but are not limited to, Avastin, Ramucirumab or VEGF trap (Aflibercept), or variants or functional portions thereof.

[0290] In some embodiments, the binding moiety that specifically recognizes VEGF used in the multispecific anti-IGFBP7 construct is a heavy chain variable region (V H ) and the light chain variable region (V L ), and V H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 99, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 100, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 101, L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO:102, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO:103, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO:104.

[0291] In some embodiments, the binding moiety that specifically recognizes VEGF used in the multispecific anti-IGFBP7 construct is a heavy chain variable region (V H ) and the light chain variable region (V L ), and V H comprises an HC-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, an HC-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and an HC-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; L comprises an LC-CDR1 comprising the amino acid sequence of SEQ ID NO: 108, an LC-CDR2 comprising the amino acid sequence of SEQ ID NO: 109, and an LC-CDR3 comprising the amino acid sequence of SEQ ID NO: 110.

[0292] In some embodiments, the binding moiety that specifically recognizes VEGF used in the multispecific anti-IGFBP7 construct comprises the amino acid sequence of SEQ ID NO:99.

[0293] Anti-IGFBP7 fusion protein In some embodiments, the anti-IGFBP7 construct comprises an anti-IGFBP7 antibody portion (eg, an anti-IGFBP7 sdAb portion) and a second portion.

[0294] In some embodiments, the second moiety is a ligand (e.g., a ligand that interacts with another molecule). In some embodiments, the second moiety is a peptide. In some embodiments, the second moiety is a cytokine.

[0295] In some embodiments, the second moiety comprises a half-life extending moiety.

[0296] In some embodiments, the half-life extending moiety is an albumin binding moiety (e.g., an albumin binding antibody moiety). In some embodiments, the anti-IGFBP7 antibody moiety and the half-life extending moiety are linked via a linker (such as any of the linkers described in the "Linkers" section).

[0297] In some embodiments, the half-life extending moiety is an Fc fragment. In some embodiments, the Fc fragment is selected from the group consisting of Fc fragments from IgG, IgA, IgD, IgE, IgM, and combinations and hybrids thereof. In some embodiments, the Fc fragment is selected from the group consisting of Fc fragments from IgG1, IgG2, IgG3, IgG4, and combinations and hybrids thereof. In some embodiments, the Fc has one or more amino acid modifications that result in an extension of the half-life of the antibody moiety in serum. In some embodiments, the Fc fragment has reduced effector function compared to the corresponding wild-type Fc fragment. In some embodiments, the Fc fragment has enhanced effector function compared to the corresponding wild-type Fc fragment.

[0298] In some embodiments, the Fc fragment is derived from IgG2a (such as murine IgG2a).

[0299] In some embodiments, the second moiety is a lipid. In some embodiments, the lipid is conjugated to an anti-IGFBP7 antibody moiety (such as an anti-IGFBP7 antibody) and is capable of binding to albumin, thereby extending the half-life of the anti-IGFBP7 antibody moiety.

[0300] In some embodiments, the second moiety is albumin or a portion of albumin (eg, human albumin, eg, human serum albumin).

[0301] Anti-IGFBP7 immunoconjugates The present application also provides an anti-IGFBP7 immunoconjugate comprising an anti-IGFBP7 antibody moiety (such as any IGFBP7 antibody moiety described herein) and a second agent. In some embodiments, the second agent is a therapeutic agent. In some embodiments, the second agent is a label.

[0302] In some embodiments, the second agent is a cytotoxic agent. In some embodiments, the cytotoxic agent is a chemotherapeutic agent. In some embodiments, the cytotoxic agent is a growth inhibitory agent. In some embodiments, the cytotoxic agent is a toxin (e.g., a protein toxin, an enzymatically active toxin of bacterial, fungal, plant or animal origin, or a fragment thereof). In some embodiments, the cytotoxic agent is a radioisotype (i.e., a radioconjugate).

[0303] Immunoconjugates allow targeted delivery of, and in some embodiments intracellular accumulation of, drug moieties to tumors, where systemic administration of unconjugated drugs can result in unacceptable levels of toxicity to normal cells (Polakis P. (2005) Current Opinion in Pharmacology 5:382-387).

[0304] Production of the immunoconjugates described herein can be found, for example, in US Pat. No. 9,562,099 and US Pat. No. 7,541,034, which are incorporated herein by reference in their entireties.

[0305] Linker In some embodiments, the anti-IGFBP7 constructs described herein include one or more linkers between two moieties (e.g., the anti-IGFBP7 antibody moiety and the half-life extending moiety, the anti-IGFBP7 antibody moiety and the second binding moiety in the multispecific constructs described above). The length, degree of flexibility and / or other properties of the linker(s) used in the anti-IGFBP7 constructs may have some effect on the properties including, but not limited to, affinity, specificity or avidity for one or more specific antigens or epitopes. For example, a longer linker may be selected to ensure that two adjacent domains do not sterically interfere with each other. In some embodiments, the linker (such as a peptide linker) includes flexible residues (such as glycine and serine) so that the adjacent domains can move freely relative to each other. For example, a glycine-serine duplex may be a suitable peptide linker. In some embodiments, the linker is a non-peptide linker. In some embodiments, the linker is a peptide linker. In some embodiments, the linker is a non-cleavable linker. In some embodiments, the linker is a cleavable linker.

[0306] Other linker considerations include their effect on the physical or pharmacokinetic properties of the resulting compound, such as solubility, lipophilicity, hydrophilicity, hydrophobicity, stability (more or less stable and programmed degradation), rigidity, flexibility, immunogenicity, modulation of antibody binding, ability to be incorporated into micelles or liposomes, etc.

[0307] Peptide Linker The peptide linker may have a naturally occurring or non-naturally occurring sequence. For example, a sequence derived from the hinge region of a heavy chain-only antibody may be used as a linker. See, for example, WO1996 / 34103.

[0308] The peptide linker can be any suitable length. In some embodiments, the peptide linker is at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 50, 75, 100 or more amino acids in length. In some embodiments, the peptide linker is no more than about 100, 75, 50, 40, 35, 30, 25, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5 or less amino acids in length. In some embodiments, the length of the peptide linker is from about 1 amino acid to about 10 amino acids, from about 1 amino acid to about 20 amino acids, from about 1 amino acid to about 30 amino acids, from about 5 amino acids to about 15 amino acids, from about 10 amino acids to about 25 amino acids, from about 5 amino acids to about 30 amino acids, from about 10 amino acids to about 30 amino acids in length, from about 30 amino acids to about 50 amino acids, from about 50 amino acids to about 100 amino acids, or from about 1 amino acid to about 100 amino acids.

[0309] The essential technical feature of such peptide linkers is that said peptide linkers do not contain polymerization activity. The features of peptide linkers, including the absence of promotion of secondary structures, are known in the art and are described, for example, in Dall'Acqua et al. (Biochem. (1998) 37, 9266-9273), Cheadle et al. (Mol Immunol (1992) 29, 21-30) and Raag and Whitlow (FASEB (1995) 9(1), 73-80). A particularly preferred amino acid in the context of "peptide linkers" is Gly. Furthermore, peptide linkers that do not promote secondary structures are preferred. The linkage of domains to each other can be provided, for example, by genetic engineering. Methods for preparing fused, operably linked bispecific single-chain constructs and expressing them in mammalian cells or bacteria are well known in the art (e.g., WO 99 / 54440; Ausubel, Current Protocols in Molecular Biology, Green Publishing Associates and Wiley Interscience, NY 1989 and 1994 or Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 2001).

[0310] The peptide linker may be a stable linker that is not cleaved by proteases, particularly matrix metalloproteinases (MMPs).

[0311] The linker may be a flexible linker. Exemplary flexible linkers include glycine polymers (G) n (SEQ ID NO:54), glycine-serine polymers (e.g., (GS) n (SEQ ID NO:55), (GSGGS) n (SEQ ID NO:56), (GGGGS) n (SEQ ID NO:57), and (GGGS) n(SEQ ID NO:58), where n is an integer of at least 1), glycine-alanine polymers, alanine-serine polymers, and other flexible linkers known in the art. Glycine and glycine-serine polymers are relatively unstructured and can therefore function as neutral tethers between components. Glycine has access to much more phi-psi space than alanine and is much less restricted than residues with longer side chains (see Scheraga, Rev. Computational Chem. 11 173-142 (1992)). Those skilled in the art will recognize that the design of antibody fusion proteins can include linkers that are wholly or partially flexible, such that the linker can include a flexible linker portion as well as one or more portions that provide a less flexible structure to provide the desired antibody or fusion protein structure.

[0312] Additionally, exemplary linkers also include (GGGGS) n (SEQ ID NO: 57), where n is an integer from 1 to 8, for example, (GGGGS) 3 (SEQ ID NO:59; hereinafter referred to as "(G4S)3" or "GS3"), or (GGGGS) 6 (SEQ ID NO: 60; hereinafter referred to as "(G4S)6" or "GS6"). In some embodiments, the peptide linker is (GSTSGSGKPGSGEGS) n (SEQ ID NO: 61), where n is an integer from 1 to 3.

[0313] Non-peptide linkers The coupling of the two moieties can be achieved by any chemical reaction that binds the two molecules, so long as both components retain their respective activities, for example, binding to IGFBP7 in an anti-IGFBP7 multispecific antibody and to a second agent, respectively. This binding can include many chemical mechanisms, such as covalent binding, affinity binding, intercalation, coordinate binding, and complexation. In some embodiments, the binding is a covalent bond. Covalent binding can be achieved either by direct condensation of existing side chains or by incorporation of an external crosslinking molecule. Many bivalent or polyvalent linking agents can be useful in coupling protein molecules in this context. For example, representative coupling agents can include organic compounds such as thioesters, carbodiimides, succinimide esters, diisocyanates, glutaraldehyde, diazobenzene, and hexamethylenediamine. This list is not intended to be exhaustive of the various classes of coupling agents known in the art, but rather, is illustrative of the more common coupling agents (see Killen and Lindstrom, Jour. Immun. 133:1335-2549 (1984); Jansen et al., Immunological Reviews 62:185-216 (1982); and Vitetta et al., Science 238:1098 (1987)).

[0314] Linkers that may be applied in the present application have been described in the literature (see, for example, Ramakrishnan, S. et al., Cancer Res. 44:201-208 (1984) which describes the use of MBS (M-maleimidobenzoyl-N-hydroxysuccinimide ester)). In some embodiments, non-peptide linkers used herein include: (i) EDC (1-ethyl-3-(3-dimethylamino-propyl) carbodiimide hydrochloride; (ii) SMPT (4-succinimidyloxycarbonyl-alpha-methyl-alpha-(2-pyridyl-dithio)-toluene (Pierce Chem. Co., Cat. (21558G); (iii) SPDP (succinimidyl-6[3-(2-pyridyldithio)propionamido]hexanoate (Pierce Chem. Co., Cat. #21651G); (iv) sulfo-LC-SPDP (sulfosuccinimidyl-6[3-(2-pyridyldithio)-propianamido]hexanoate (Pierce Chem. Co., Cat. #2165-G); and (v) sulfo-NHS (N-hydroxysulfo-succinimide: Pierce Chem. Co., Cat. #2165-G) conjugated to EDC. Chem. Co., Cat. #24510).

[0315] The above linkers contain components with different attributes and therefore may result in bispecific antibodies with different physiochemical properties. For example, sulfo-NHS esters of alkyl carboxylates are more stable than sulfo-NHS esters of aromatic carboxylates. NHS ester-containing linkers are less soluble than sulfo-NHS esters. The linker SMPT also contains a sterically hindered disulfide bond, which can form a highly stable antibody fusion protein. Disulfide bonds are generally less stable than other bonds because disulfide bonds are cleaved in vitro, resulting in less available antibody fusion protein. In particular, sulfo-NHS can increase the stability of carbodiimide coupling. Carbodiimide coupling (such as EDC) when used in combination with sulfo-NHS forms esters that are more resistant to hydrolysis than the carbodiimide coupling reaction alone.

[0316] Anti-IGFBP7 antibody moiety or construct variants a) Antibody affinity The binding specificity of an anti-IGFBP7 antibody moiety can be determined experimentally by methods known in the art, including, but not limited to, Western blot, ELISA-, RIA-, ECL-, IRMA-, EIA-, BLI, SPR, BIACORE™-test and peptide scan.

[0317] In some embodiments, the K D is about 10 -7 M~about 10 -12 M, about 10 -7 M~about 10 -8 M, about 10 -8 M~about 10 -9 M, about 10 -9 M~about 10 -10 M, about 10 -10 M~about 10 -11 M, about 10 -11 M~about 10 -12 M, about 10 -7 M~about 10 -12 M, about 10 -8 M~about 10 -12 M, about 10 -9 M~about 10 -12 M, about 10 -10 M~about 10 -12 M, about 10 -7 M~about 10 -11 M, about 10 -8 M~about 10 -11 M, about 10 -9 M~about 10 -11 M, about 10 -7 M~about 10 -10 M, about 10 -8 M~about 10 -10 M, or about 10 -7 M~about 10 -9 In some embodiments, the K of binding between the antibody moiety and IGFBP7 is M. D is about 10 -7 M, 10 -8 M, 10 -9 M, 10-10 M, 10 -11 M, or 10 -12 M. In some embodiments, the IGFBP7 is human IGFBP7.

[0318] In some embodiments, the K of binding between the antibody moiety and IGFBP7 is on is about 10 3 M -1 s -1 ~about 10 8 M -1 s -1 , about 10 3 M -1 s -1 ~about 10 4 M -1 s -1 , about 10 4 M -1 s -1 ~about 10 5 M -1 s -1 , about 10 5 M -1 s -1 ~about 10 6 M -1 s -1 , about 10 6 M -1 s -1 ~about 10 7 M -1 s -1 , or about 10 7 M -1 s -1 ~about 10 8 M -1 s -1 In some embodiments, the K of binding between the antibody moiety and IGFBP7 is on is about 10 3 M -1 s -1 ~about 10 5 M -1 s -1 , about 10 4 M -1 s -1 ~about 10 6 M -1 s -1 , about 10 5 M -1 s -1 ~about 107 M -1 s -1 , about 10 6 M -1 s -1 ~about 10 8 M -1 s -1 , about 10 4 M -1 s -1 ~about 10 7 M -1 s -1 , or about 10 5 M -1 s -1 ~about 10 8 M -1 s -1 In some embodiments, the K of binding between the antibody moiety and IGFBP7 is on is about 10 3 M -1 s -1 , 10 4 M -1 s -1 , 10 5 M -1 s -1 , 10 6 M -1 s -1 , 10 7 M -1 s -1 or 10 8 M -1 s -1 In some embodiments, the IGFBP7 is human IGFBP7.

[0319] In some embodiments, the K of binding between the antibody moiety and IGFBP7 is off takes about 1 s -1 ~about 10 -6 s -1 , about 1s -1 ~about 10 -2 s -1 , about 10 -2 s -1 ~about 10 -3 s -1 , about 10 -3 s -1 ~about 10 -4 s -1 , about 10 -4 s-1 ~about 10 -5 s -1 , about 10 -5 s -1 ~about 10 -6 s -1 , about 1s -1 ~about 10 -5 s -1 , about 10 -2 s -1 ~about 10 -6 s -1 , about 10 -3 s -1 ~about 10 -6 s -1 , about 10 -4 s -1 ~about 10 -6 s -1 , about 10 -2 s -1 ~about 10 -5 s -1 , or about 10 -3 s -1 ~about 10 -5 s -1 In some embodiments, the K of binding between the antibody moiety and IGFBP7 is off is at least about 1s -1 , 10 -2 s -1 , 10 -3 s -1 , 10 -4 s -1 , 10 -5 s -1 or 10 -6 s -1 In some embodiments, the IGFBP7 is human IGFBP7.

[0320] In some embodiments, the binding affinity of the anti-IGFBP7 antibody portion or anti-IGFBP7 construct is higher (e.g., K ) than an existing anti-IGFBP7 antibody (e.g., an anti-human IGFBP7 antibody, e.g., MM01). D value is smaller).

[0321] b) Chimeric or humanized antibodies In some embodiments, the anti-IGFBP7 antibody portion is a chimeric antibody. Certain chimeric antibodies are described, for example, in U.S. Patent No. 4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984). In some embodiments, the chimeric antibody comprises a non-human variable region (e.g., a variable region from a llama) and a human constant region. In some embodiments, the chimeric antibody is a "class-switched" antibody whose class or subclass has been changed from that of the parent antibody. The chimeric antibody includes antigen-binding fragments thereof.

[0322] In some embodiments, the anti-IGFBP7 antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce immunogenicity to humans while retaining the specificity and affinity of the parent non-human antibody. Generally, a humanized antibody comprises one or more variable domains in which the HVR, e.g., CDR (or a portion thereof) is derived from a non-human antibody and the FR (or a portion thereof) is derived from a human antibody sequence. The humanized antibody also optionally comprises at least a portion of a human constant region. In some embodiments, some FR residues in the humanized antibody are replaced with corresponding residues from a non-human antibody (e.g., the antibody from which the HVR residues are derived), e.g., to restore or improve the specificity or affinity of the antibody.

[0323] Humanized antibodies and methods for making them are reviewed, e.g., in Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008), and further described, e.g., in Riechmann et al., Nature 332:323-329 (1988); Queen et al., Proc. Nat'l Acad. Sci. USA 86:10029-10033 (1989); U.S. Patent Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409; Kashmiri et al., Methods 36:25-34 (2005) (describing SDR (a-CDR) grafting); Padlan, Mol. Immunol. 28:489-498 (1991) (describing "resurfacing"); Dall'Acqua et al., Methods 36:43-60 (2005) (describing "FR shuffling"); and Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer, 83:252-260 (2000) (describing a "guided selection" approach to FR shuffling).

[0324] Human framework regions that can be used for humanization include, but are not limited to, framework regions selected using the "best-fit" method (see, e.g., Sims et al. J. Immunol. 151:2296 (1993)); framework regions derived from consensus sequences of human antibodies of particular subgroups of light or heavy chain variable regions (see, e.g., Carter et al. Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al. J. Immunol., 151:2623 (1993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008)); and framework regions derived from screening of FR libraries (see, e.g., Baca et al. al., J. Biol. Chem. 272:10678-10684 (1997) and Rosok et al., J. Biol. Chem. 271:22611-22618 (1996).

[0325] It is understood that humanization of non-human derived antibodies is a common and routinely used technique.Therefore, it is understood that any and all humanized formats of anti-IGFBP7 antibodies disclosed in sequence listing can be used in preclinical or clinical situations.When any humanized format of the referenced anti-IGFBP7 antibodies or its antigen-binding region is used in such preclinical or clinical situations, the humanized format is expected to have the same or similar biological activity and profile as the original non-humanized format.

[0326] c) Human antibodies In some embodiments, the anti-IGFBP7 antibody portion is a human antibody (known as a human domain antibody or human DAb). Human antibodies can be produced using various techniques known in the art. Human antibodies are generally described in van Dijk and van de Winkel, Curr.Opin.Pharmacol.5:368-74(2001), Lonberg, Curr.Opin.Immunol.20:450-459(2008), and Chen, Mol.Immunol.47(4):912-21(2010). Transgenic mice or rats capable of producing fully human single domain antibodies (or DAbs) are known in the art. See, e.g., US20090307787A1, U.S. Patent No. 8,754,287, US20150289489A1, US20100122358A1, and WO2004049794.

[0327] Human antibodies (e.g., human DAbs) can be prepared by administering immunogens to transgenic animals that have been modified to produce intact human antibodies or intact antibodies with human variable regions in response to antigen challenge. Such animals typically contain all or part of a human immunoglobulin locus that replaces the endogenous immunoglobulin locus, is extrachromosomal, or is randomly integrated into the animal's chromosomes. In such transgenic mice, the endogenous immunoglobulin locus is generally inactivated. For a review of methods for obtaining human antibodies from transgenic animals, see Lonberg, Nat. Biotech. 23:1117-1125 (2005). See, e.g., U.S. Pat. Nos. 6,075,181 and 6,150,584 (describing XENOMOUSE™ technology); U.S. Pat. No. 5,770,429 (describing HuMab®); U.S. Pat. No. 7,041,870 (describing KM MOUSE®), and U.S. Patent Application No. 2007 / 0061900 (describing VelociMouse®). The human variable regions from intact antibodies produced by such animals can be further modified, for example, by combining with different human constant regions.

[0328] Human antibodies (e.g., human DAbs) can also be produced by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for producing human monoclonal antibodies have been described (see, for example, Kozbor J.Immunol., 133:3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp.51-63 (Marcel Dekker, Inc., New York, 1987); and Boerner et al., J.Immunol., 147:86 (1991)). Human antibodies generated via human B-cell hybridoma technology are also described in Li et al., Proc.Natl.Acad.Sci.USA, 103:3557-3562 (2006). Further methods include those described, for example, in U.S. Patent No. 7,189,826 (describing the production of monoclonal human IgM antibodies from hybridoma cell lines) and Ni, Xiandai Mianyixue, 26(4):265-268 (2006) (describing human-human hybridomas). Human hybridoma technology (trioma technology) is also described in Vollmers and Brandlein, Histology and Histopathology, 20(3):927-937 (2005) and Vollmers and Brandlein, Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-91 (2005).

[0329] Human antibodies (e.g., human DAbs) can also be made by isolating Fv clone variable domain sequences selected from a human-derived phage display library. Such variable domain sequences can then be combined with the desired human constant domains. Techniques for selecting human antibodies from an antibody library are described below.

[0330] d) Library-derived antibodies The anti-IGFBP7 antibody portions described herein can be isolated by screening combinatorial libraries for antibodies with the desired activity or activities. For example, various methods are known in the art for generating phage display libraries and screening such libraries for antibodies with the desired binding characteristics. Such methods are reviewed, for example, in Hoogenboom et al., Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, 2001), and further described, for example, in McCafferty et al., Nature 348:552-554; Clackson et al., Nature 352:624-628 (1991); Marks et al., J. Mol. Biol. 222:581-597 (1992); Marks and Bradbury, in Methods in Molecular Biology 248:161-175 (Lo, ed., Human Press, Totowa, NJ, 2003); Sidhu et al., J. Mol. Biol. 338(2):299-310 (2004); Lee et al. al., J. Mol. Biol. 340(5):1073-1093 (2004); Fellouse, Proc. Natl. Acad. Sci. USA 101(34):12467-12472 (2004); and Lee et al., J. Immunol. Methods 284(1-2):119-132 (2004). Methods for constructing single domain antibody libraries have been described, see, e.g., U.S. Patent No. 7,371,849.

[0331] In certain phage display methods, V H and V LGene repertoires can be cloned separately by polymerase chain reaction (PCR) and randomly recombined in phage libraries, which can then be screened for antigen-binding phages as described in Winter et al., Ann. Rev. Immunol., 12:433-455 (1994). Phages typically display antibody fragments as either scFv fragments or Fab fragments. Libraries from immunized sources provide high affinity antibodies to immunogens without the need to construct hybridomas. Alternatively, naive repertoires can be cloned (e.g., from humans) without any immunization to provide a single source of antibodies against a wide range of non-self and self antigens, as described in Griffiths et al., EMBO J, 12:725-734 (1993). Finally, naive libraries can also be generated synthetically by cloning unrearranged V gene segments from stem cells and using PCR primers containing random sequences to encode highly variable CDR3 regions and achieve rearrangement in vitro, as described by Hoogenboom and Winter, J. Mol. Biol., 227:381-388 (1992). Patent publications describing human antibody phage libraries include, for example, U.S. Pat. No. 5,750,373, and U.S. Patent Application Publication Nos. 2005 / 0079574, 2005 / 0119455, 2005 / 0266000, 2007 / 0117126, 2007 / 0160598, 2007 / 0237764, 2007 / 0292936, and 2009 / 0002360.

[0332] Antibodies or antibody fragments isolated from a human antibody library are considered herein to be human antibodies or human antibody fragments.

[0333] e) Substitutions, insertions, deletions and variants In some embodiments, anti-IGFBP7 antibody variants are provided with one or more amino acid substitutions. Sites of interest for substitution mutagenesis include HVRs (or CDRs) and FRs. Conservative substitutions are shown in Table 2 under the heading of "preferred substitutions". More substantial changes are provided in Table 2 under the heading of "exemplary substitutions" and are further described below with reference to amino acid side chain classes. Amino acid substitutions can be introduced into the antibody of interest and the product screened for the desired activity, for example, retained / improved antigen binding, reduced immunogenicity, or improved ADCC or CDC. [Table 2]

[0334] Amino acids can be grouped according to general side chain properties: (1) hydrophobic: norleucine, Met, Ala, Val, Leu, Ile; (2) neutral hydrophilic: Cys, Ser, Thr, Asn, Gln; (3) acidic: Asp, Glu; (4) basic: His, Lys, Arg; (5) residues that affect chain orientation: Gly, Pro; and (6) aromatic: Trp, Tyr, Phe.

[0335] Non-conservative substitutions involve exchanging a member of one of these classes for another class.

[0336] One type of substitution variant involves substituting one or more hypervariable region residues of a parent antibody (e.g., a humanized or human antibody). Generally, the resulting variant(s) selected for further study have alterations (e.g., improvements) in certain biological properties (e.g., increased affinity, reduced immunogenicity, improved solubility) compared to the parent antibody and / or substantially retain certain biological properties of the parent antibody. An exemplary substitution variant is an affinity matured antibody, which can be conveniently generated using, for example, phage display-based affinity maturation techniques such as those described herein. Briefly, one or more HVR residues are mutated and the variant antibodies are displayed on phage and screened for a particular biological activity (e.g., binding affinity).

[0337] For example, modifications (e.g., substitutions) may be made to HVRs to improve antibody affinity. Such modifications may be made at HVR "hot spots," i.e., codons that undergo high frequency mutation during the somatic maturation process (see, e.g., Chowdhury, Methods Mol. Biol. 207:179-196 (2008)) and / or residues encoded by the SDRs (a-CDRs), resulting in variant V H or V Lare tested for binding affinity. Affinity maturation by constructing and reselecting a secondary library is described, for example, by Hoogenboom et al. in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, (2001)). In some embodiments of affinity maturation, diversity is introduced into the variable genes selected for maturation by any of a variety of methods (e.g., error-prone PCR, chain shuffling, or oligonucleotide-directed mutagenesis). A secondary library is then created. The library is then screened to identify any antibody variants with the desired affinity. Another method of introducing diversity involves an HVR-directed approach, in which several HVR residues (e.g., 4-6 residues at a time) are randomized. HVR residues involved in antigen binding can be specifically identified, for example, using alanine scanning mutagenesis or modeling. In particular, CDR-H3 and CDR-L3 are often targeted.

[0338] In some embodiments, substitutions, insertions or deletions can occur within one or more HVRs, so long as such changes do not substantially reduce the ability of the antibody to bind to the antigen. For example, conservative changes (e.g., conservative substitutions provided herein) that do not substantially reduce binding affinity can be made in HVRs. Such changes can be outside of HVR "hot spots" or CDRs.

[0339] A useful method for identifying antibody residues or regions that can be targeted for mutagenesis is called "alanine scanning mutagenesis" described in Cunningham and Wells (1989) Science, 244:1081-1085. In this method, residues or groups of target residues (e.g., charged residues such as Arg, Asp, His, Lys and Glu) are identified and replaced with neutral or negatively charged amino acids (e.g., alanine or polyalanine) to determine whether the interaction between the antibody and antigen is affected. Further substitutions can be introduced at amino acid positions that show functional sensitivity to the initial substitution. Alternatively, or in addition, a crystal structure of an antigen-antibody complex to identify contact points between the antibody and antigen. Such contact and adjacent residues can be targeted or eliminated as candidates for substitution. The variants can be screened to determine whether they contain the desired properties.

[0340] Amino acid sequence insertions include amino- and / or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. An example of a terminal insertion is an antibody with an N-terminal methionyl residue. Other insertional variants of the antibody molecule include the fusion to the N- or C-terminus of the antibody to an enzyme (e.g., for ADEPT) or a polypeptide which increases the serum half-life of the antibody.

[0341] f) Glycosylation variants In some embodiments, the anti-IGFBP7 construct is altered to increase or decrease the extent to which the construct is glycosylated. Addition or deletion of glycosylation sites to an antibody can be conveniently accomplished by altering the amino acid sequence such that one or more glycosylation sites are created or removed.

[0342] If the anti-IGFBP7 construct contains an Fc region, the carbohydrate attached thereto may vary. Native antibodies produced by mammalian cells typically contain the C of the Fc region.H The oligosaccharides of the antibody moiety include branched, biantennary oligosaccharides that are generally linked by N-linkage to Asn297 of the 2 domain. See, for example, Wright et al. TIBTECH 15:26-32 (1997). The oligosaccharides may contain various carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose and sialic acid, as well as fucose linked to GlcNAc in the "stem" of the biantennary oligosaccharide structure. In some embodiments, modification of the oligosaccharides of the antibody moiety may be performed to generate antibody variants with specific improved properties.

[0343] In some embodiments, the anti-IGFBP7 construct has a carbohydrate structure that lacks fucose attached (directly or indirectly) to the Fc region. For example, the amount of fucose in such antibodies can be 1%-80%, 1%-65%, 5%-65% or 20%-40%. The amount of fucose is determined by calculating the average amount of fucose in the glycan at Asn297 relative to the sum of all glycan structures (e.g., complex, hybrid and high mannose structures) attached to Asn297 as measured by MALDI-TOF mass spectrometry, e.g., as described in WO 2008 / 077546. Asn297 refers to an asparagine residue located at about position 297 of the Fc region (EU numbering of Fc region residues); however, Asn297 may also be located about ±3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300, due to minor sequence variations in the antibody. Such fucosylated variants may have improved ADCC function. For example, U.S. Patent Application Publication Nos. US 2003 / 0157108 (Presta, L.); US 2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd.). Examples of publications related to "defucosylated" or "fucose-deficient" antibody variants include: US 2003 / 0157108; WO 2000 / 61739; WO 2001 / 29246; US 2003 / 0115614; US 2002 / 0164328; US 2004 / 0093621; US ​​2004 / 0132140; US 2004 / 0110704; US 2004 / 0110282; US 2004 / 0109865; WO 2003 / 085119; WO 2003 / 084570; WO 2005 / 035586; WO 2005 / 035778;WO2005 / 053742;WO2002 / 031140;Okazaki et al.J.Mol.Biol.336:1239-1249(2004);Yamane-Ohnuki et al.Biotech.Bioeng.87:614(2004).Examples of cell lines capable of producing defucosylated antibodies include Lec13 CHO cells, which are deficient in protein fucosylation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); U.S. Patent Application No. US 2003 / 0157108 A1, Presta, L; and WO 2004 / 056312 A1, Adams et al., especially Example 11), and knockout cell lines, such as alpha-1,6-fucosyltransferase gene, FUT8, knockout CHO cells (e.g., Yamane-Ohnuki et al. Biotech. Bioeng. 87:614 (2004); Kanda, Y. et al., Biotechnol. Bioeng., 94(4):680-688 (2006); and WO2003 / 085107).

[0344] In some embodiments, the anti-IGFBP7 construct has bisected oligosaccharides, for example, biantennary oligosaccharides attached to the Fc region of the antibody are bisected by GlcNAc. Such antibody variants may have reduced fucosylation and / or improved ADCC function. Examples of such antibody variants are described, for example, in WO 2003 / 011878 (Jean-Mairet et al.); U.S. Patent No. 6,602,684 (Umana et al.); and US 2005 / 0123546 (Umana et al.). Also provided are antibody variants having at least one galactose residue in the oligosaccharide attached to the Fc region. Such antibody variants may have improved CDC function. Such antibody variants are described, for example, in WO 1997 / 30087 (Patel et al.); WO 1998 / 58964 (Raju, S.); and WO 1999 / 22764 (Raju, S.).

[0345] g) Fc region variant In some embodiments, the anti-IGFBP7 construct comprises an Fc fragment.

[0346] The term "Fc region", "Fc domain", "Fc fragment" or "Fc" refers to the C-terminal non-antigen binding region of an immunoglobulin heavy chain that contains at least a portion of the constant region. This term includes native and variant Fc regions. In some embodiments, a human IgG heavy chain Fc region extends from Cys226 to the carboxyl terminus of the heavy chain. However, the C-terminal lysine (Lys447) of the Fc region may be present or absent without affecting the structure or stability of the Fc region. Unless otherwise specified herein, the numbering of amino acid residues in an IgG or Fc region follows the EU numbering system for antibodies, also called the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0347] In some embodiments, the Fc fragment is derived from an immunoglobulin selected from the group consisting of IgG, IgA, IgD, IgE, IgM, and combinations and hybrids thereof. In some embodiments, the Fc fragment is derived from an immunoglobulin selected from the group consisting of IgG1, IgG2, IgG3, IgG4, and combinations and hybrids thereof.

[0348] In some embodiments, the Fc fragment has reduced effector function compared to a corresponding wild-type Fc fragment (e.g., at least about 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90% or 95% reduced effector function as measured by the level of antibody-dependent cellular cytotoxicity (ADCC).

[0349] In some embodiments, the Fc fragment is an IgG1 Fc fragment. In some embodiments, the IgG1 Fc fragment comprises a L234A mutation and / or a L235A mutation. In some embodiments, the IgG1 Fc fragment comprises a L235A mutation and / or a G237A mutation. In some embodiments, the Fc fragment is an IgG2 or IgG4 Fc fragment. In some embodiments, the Fc fragment is an IgG4 Fc fragment comprising S228P, F234A and / or L235A mutations. In some embodiments, the Fc fragment comprises a N297A mutation. In some embodiments, the Fc fragment comprises a N297G mutation.

[0350] In some embodiments, one or more amino acid modifications can be introduced into the Fc region of an antibody moiety, thereby generating an Fc region variant. The Fc region variant can comprise a human Fc region sequence (e.g., a human IgG1, IgG2, IgG3, or IgG4 Fc region) that contains an amino acid modification (e.g., a substitution) at one or more amino acid positions.

[0351] In some embodiments, the Fc fragment has some, but not all, effector functions, making it a desirable candidate for applications where the half-life of the antibody portion in vivo is important, but where certain effector functions (such as complement and ADCC) are unnecessary or detrimental. In vitro and / or in vivo cytotoxicity assays can be performed to confirm the reduction / depletion of CDC and / or ADCC activity. For example, Fc receptor (FcR) binding assays can be performed to ensure that the antibody lacks FcγR binding (and thus likely lacks ADCC activity) but retains FcRn binding ability. NK cells, the primary cells for mediating ADCC, express only FcγRIII, while monocytes express FcγRI, FcγRII and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 2 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol. 9:457-492 (1991). Non-limiting examples of in vitro assays to assess ADCC activity of a molecule of interest are described in U.S. Pat. No. 5,500,362 (see, e.g., Hellstrom, I. et al. Proc. Nat'l Acad. Sci. USA 83:7059-7063 (1986)) and Hellstrom, I et al., Proc. Nat'l Acad. Sci. USA 82:1499-1502 (1985); 5,821,337 (see, Bruggemann, M. et al., J. Exp. Med. 166:1351-1361 (1987)). Alternatively, non-radioactive assay methods may be used (see, for example, the ACTI™ Non-Radioactive Cytotoxicity Assay for Flow Cytometry (CellTechnology, Inc. Mountain View, CA; and CytoTox 96® Non-Radioactive Cytotoxicity Assay (Promega, Madison, WI)). Useful effector cells for such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells.Alternatively, or in addition, the ADCC activity of the molecule of interest can be evaluated in vivo, for example, in an animal model as disclosed in Clynes et al. Proc. Nat'l Acad. Sci. USA 95:652-656 (1998). A C1q binding assay can also be performed to confirm that the antibody cannot bind to C1q and therefore lacks CDC activity. See, for example, the C1q and C3c binding ELISAs in WO 2006 / 029879 and WO 2005 / 100402. To assess complement activation, a CDC assay can be performed (see, e.g., Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996); Cragg, MS et al., Blood 101:1045-1052 (2003); and Cragg, MS and MJ Glennie, Blood 103:2738-2743 (2004)). FcRn binding and in vivo clearance / half-life determinations can also be performed using methods known in the art (see, e.g., Petkova, SB et al., Int'l. Immunol. 18(12):1759-1769 (2006)).

[0352] Antibodies with reduced effector function include those with substitutions at one or more of Fc region residues 238, 265, 269, 270, 297, 327, and 329 (U.S. Patent No. 6,737,056). Such Fc variants include Fc variants with substitutions at two or more of amino acid positions 265, 269, 270, 297, and 327, including the so-called "DANA" Fc variant in which residues 265 and 297 are substituted with alanine (U.S. Patent No. 7,332,581). In some embodiments, the Fc fragment comprises a N297A mutation. In some embodiments, the Fc fragment comprises a N297G mutation.

[0353] Certain antibody variants with improved or diminished binding to FcRs have been described. (See, e.g., U.S. Pat. No. 6,737,056; WO 2004 / 056312; and Shields et al., J. Biol. Chem. 9(2):6591-6604 (2001).)

[0354] In some embodiments, the Fc fragment is an IgG1 Fc fragment. In some embodiments, the IgG1 Fc fragment comprises a L234A mutation and / or a L235A mutation. In some embodiments, the IgG1 Fc fragment comprises a L235A mutation and / or a G237A mutation. In some embodiments, the Fc fragment is an IgG2 or IgG4 Fc fragment. In some embodiments, the Fc fragment is an IgG4 Fc fragment comprising S228P, F234A and / or L235A mutations.

[0355] In some embodiments, the antibody portion comprises an Fc region with one or more amino acid substitutions that improve ADCC, e.g., substitutions at positions 298, 333 and / or 334 of the Fc region (EU numbering of residues).

[0356] In some embodiments, modifications are made to the Fc region that result in altered (i.e., either improved or decreased) C1q binding and / or complement dependent cytotoxicity (CDC), e.g., as described in U.S. Pat. No. 6,194,551, WO 99 / 51642, and Idusogie et al. J. Immunol. 164:4178-4184 (2000).

[0357] In some embodiments, the Fc fragment has one or more mutations at Thr250, Met252, Ser254, The256, Thr307. Glu380, Met428, His433, and / or Asn434.

[0358] In some embodiments, the anti-IGFBP7 construct comprises a variant Fc region that comprises one or more amino acid substitutions that alter half-life and / or alter binding to the neonatal Fc receptor (FcRn). Antibodies with increased half-life and improved binding to the neonatal Fc receptor (FcRn), responsible for maternal IgG transfer to the fetus (Guyer et al., J. Immunol. 117:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)), are described in US2005 / 0014934A1 (Hinton et al.). These antibodies comprise an Fc region with one or more substitutions that alter binding of the Fc region to FcRn. Such Fc variants include those with substitutions at one or more Fc region residues (e.g., Fc region residue 434) (U.S. Patent No. 7,371,826).

[0359] See also Duncan & Winter, Nature 322:738-40 (1988); U.S. Patent No. 5,648,260; U.S. Patent No. 5,624,821; U.S. Patent No. 8,163,881; and WO 94 / 29351 for other examples of Fc region variants.

[0360] h) Cysteine ​​engineered antibody variants In some embodiments, it may be desirable to generate cysteine ​​engineered antibody moieties, e.g., "thioMAbs," in which one or more residues of an antibody are replaced with cysteine ​​residues. In certain embodiments, the replaced residues are located at accessible sites of the antibody. By replacing those residues with cysteine, reactive thiol groups are thereby placed at accessible sites of the antibody, which can be used to conjugate the antibody to other moieties, such as drug moieties or linker-drug moieties, to generate immunoconjugates, as further described herein. In some embodiments, any one or more of the following residues can be replaced with cysteine: A118 (EU numbering) of the heavy chain; and S400 (EU numbering) of the heavy chain Fc region. Cysteine ​​engineered antibody moieties can be generated, for example, as described in U.S. Pat. No. 7,521,541.

[0361] i) Antibody derivative In some embodiments, the anti-IGFBP7 antibody moieties or constructs described herein may be further modified to include additional non-proteinaceous moieties that are known and readily available in the art. Moieties suitable for derivatization of antibodies include, but are not limited to, water-soluble polymers. Non-limiting examples of water-soluble polymers include, but are not limited to, polyethylene glycol (PEG), ethylene glycol / propylene glycol copolymers, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymers, polyamino acids (either homopolymers or random copolymers), and dextran or poly(n-vinylpyrrolidone) polyethylene glycol, propylene glycol homopolymer, propylene oxide / ethylene oxide copolymer, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may have advantages in manufacturing due to its stability in water. The polymer may be of any molecular weight and may be branched or unbranched. The number of polymers attached to the antibody may vary, and when more than one polymer is attached, they may be the same or different molecules. In general, the number and / or type of polymers used for derivatization may be determined based on considerations including, but not limited to, the specific properties or functions of the antibody to be improved, whether the antibody derivative is to be used for diagnosis under defined conditions, etc.

[0362] In some embodiments, the anti-IGFBP7 antibody moiety or construct may be further modified to include one or more biologically active proteins, polypeptides, or fragments thereof. "Biological activity" or "biologically active", as used interchangeably herein, means to exhibit biological activity in the body to perform a specific function. For example, it may mean combination with a specific biomolecule, such as a protein, DNA, and then promoting or inhibiting the activity of such a biomolecule. In some embodiments, the biologically active protein or fragment thereof includes proteins and polypeptides administered to patients as active drug substances for the prevention or treatment of disease or symptoms, as well as proteins and polypeptides used for diagnostic purposes, such as enzymes used in diagnostic tests or in vitro assays, and proteins and polypeptides administered to patients to prevent disease (such as vaccines).

[0363] III.Preparation method In some embodiments, methods are provided for preparing anti-IGFBP7 constructs or antibody moieties that specifically bind to IGFBP7, and compositions such as polynucleotides, nucleic acid constructs, vectors, host cells, or culture media produced during the preparation of anti-IGFBP7 constructs or antibody moieties. The anti-IGFBP7 constructs or antibody moieties or compositions described herein can be prepared by several processes as generally described below, and more specifically in the Examples.

[0364] Antibody expression and production The anti-IGFBP7 constructs described herein, or portions thereof (e.g., anti-IGFBP7 antibody portions, e.g., anti-IGFBP7 bispecific antibodies), can be prepared using any method known in the art, including those described below and in the Examples.

[0365] Single Domain Antibodies (sdAbs) Methods for preparing sdAbs have been described, see, e.g., Els Pardon et al., Nature Protocols, 2014;9(3):674. H H) can be obtained using methods known in the art, such as immunizing a camelid species (such as a camel or llama) and obtaining hybridomas therefrom, or by cloning a library of single domain antibodies using molecular biology techniques known in the art, followed by selection by ELISA using individual clones of the unselected library, or using phage display.

[0366] For recombinant production of sdAbs, the nucleic acid encoding the single domain antibody is isolated and inserted into a replicable vector for further cloning (amplification of the DNA) or expression. DNA encoding the single domain antibody is readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes capable of specifically binding to genes encoding the heavy and light chains of the antibody). Many vectors are available. The choice of vector depends in part on the host cell to be used. In general, preferred host cells are of prokaryotic or eukaryotic (generally mammalian) origin, including those described below.

[0367] Monoclonal antibodies A monoclonal antibody or antibody portion can be obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies constituting the population are identical except for possible naturally occurring mutations and / or post-translational modifications (e.g., isomerization, amidation) that may be present in minor amounts. Thus, the modifier "monoclonal" indicates the character of the antibody as not being a mixture of individual antibodies. For example, monoclonal antibodies can be produced using the hybridoma method first described by Kohler et al., Nature, 256:495 (1975), or can be produced by recombinant DNA methods (U.S. Pat. No. 4,816,567). In the hybridoma method, a mouse or other suitable host animal, such as a hamster or llama, is immunized as described above to elicit lymphocytes that produce or are capable of producing antibodies that specifically bind to the protein used for immunization. Alternatively, lymphocytes can be immunized in vitro. The lymphocytes are then fused with myeloma cells using a suitable fusing agent, such as polyethylene glycol, to form hybridoma cells. Goding, Monoclonal Antibodies: Principles and Practice, pp. 59-103 (Academic Press, 1986). See also Example 1 for immunization in llamas.

[0368] The immunizing agent typically includes an antigen protein or a fusion variant thereof. Generally, peripheral blood lymphocytes ("PBL") are used if cells of human origin are desired, or spleen cells or lymph node cells are used if non-human mammalian sources are desired. The lymphocytes are then fused with an immortalized cell line using a suitable fusing agent, such as polyethylene glycol, to form a hybridoma cell. Goding, Monoclonal Antibodies: Principles and Practice, Academic Press (1986), pp. 59-103.

[0369] Immortalized cell lines are usually transformed mammalian cells, especially myeloma cells of rodent, bovine and human origin. Usually, rat or mouse myeloma cell lines are used. The hybridoma cells thus prepared are seeded and grown in a suitable culture medium, preferably containing one or more substances that inhibit the growth or survival of unfused parental myeloma cells. For example, if parental myeloma cells lack the enzyme hypoxanthine guanine phosphoribosyltransferase (HGPRT or HPRT), the culture medium of hybridoma typically contains hypoxanthine, aminopterin and thymidine, which are substances that inhibit the growth of HGPRT-deficient cells (HAT medium).

[0370] Preferred immortalized myeloma cells are those that fuse efficiently, support stable high-level production of antibody by selected antibody-producing cells, and are sensitive to a medium such as HAT medium. Among these, murine myeloma lines, such as MOPC-21 and MPC-11 mouse tumors (available from the Salk Institute Cell Distribution Center, San Diego, Calif. USA), and SP-2 cells (and their derivatives, such as X63-Ag8-653) (available from the American Type Culture Collection, Manassas, Va. USA), are preferred. Human myeloma and mouse-human heteromyeloma cell lines have also been described for the production of human monoclonal antibodies (Kozbor, J. Immunol., 133:3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp.51-63 (Marcel Dekker, Inc., New York, 1987)).

[0371] The culture medium in which the hybridoma cells are growing is assayed for production of monoclonal antibodies against the antigen. Preferably, the binding specificity of the monoclonal antibodies produced by the hybridoma cells is determined by immunoprecipitation or by an in vitro binding assay, such as radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA).

[0372] The culture medium in which hybridoma cells are cultured can be assayed for the presence of monoclonal antibodies against desired antigens.Preferably, the binding affinity and specificity of monoclonal antibodies can be determined by immunoprecipitation or by in vitro binding assays, such as radioimmunoassay (RIA) or enzyme-linked assay (ELISA).Such techniques and assays are known in the art.For example, binding affinity can be determined by Scatchard analysis of Munson et al., Anal.Biochem.,107:220(1980).

[0373] After hybridoma cells that produce antibodies of desired specificity, affinity and / or activity are identified, the clones can be subcloned by limiting dilution procedures and grown by standard methods (Goding, supra). Suitable culture media for this purpose include, for example, D-MEM or RPMI-1640 medium. In addition, hybridoma cells can be grown in vivo as tumors in mammals.

[0374] The monoclonal antibodies secreted by the subclones are suitably separated from the culture medium, ascites fluid, or serum by conventional immunoglobulin purification procedures such as, for example, protein G-Sepharose, protein A-Sepharose, hydroxylapatite chromatography, gel electrophoresis, dialysis, or affinity chromatography.

[0375] Monoclonal antibodies can also be produced as described above by recombinant DNA methods, such as those described in U.S. Pat. No. 4,816,567. DNA encoding monoclonal antibodies is easily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that can specifically bind to genes encoding heavy and light chains of mouse antibodies). Hybridoma cells serve as a preferred source of such DNA. Once isolated, the DNA may be placed into an expression vector, which is then transfected into host cells such as E. coli cells, monkey COS cells, Chinese hamster ovary (CHO) cells, HEK cells, or myeloma cells (e.g., NS0 cells) that do not produce immunoglobulin proteins, and the monoclonal antibodies are synthesized in such recombinant host cells. In general, preferred host cells are of prokaryotic or eukaryotic (generally mammalian) origin, including those described below. Review articles on recombinant expression in bacteria of DNA encoding the antibody include Skerra et al., Curr. Opinion in Immunol., 5:256-262 (1993) and Pluckthun, Immunol. Revs. 130:151-188 (1992).

[0376] In further embodiments, antibodies can be isolated from antibody phage libraries generated using the techniques described in McCafferty et al., Nature, 348:552-554 (1990). Clackson et al., Nature, 352:624-628 (1991) and Marks et al., J. Mol. Biol., 222:581-597 (1991) describe the isolation of mouse and human antibodies, respectively, using phage libraries. Subsequent publications have described the production of high affinity (nM range) human antibodies by chain shuffling (Marks et al., Bio / Technology, 10:779-783 (1992)), as well as combinatorial infection and in vivo recombination (Waterhouse et al., Nucl. Acids Res., 21:2265-2266 (1993)) as strategies for constructing very large phage libraries. Therefore, these techniques are viable alternatives to traditional monoclonal antibody hybridoma techniques for isolating monoclonal antibodies.

[0377] The DNA can also be modified, for example, by substituting the coding sequence for human heavy and light chain constant domains for the homologous murine sequences (U.S. Pat. No. 4,816,567; Morrison et al., Proc. Natl Acad. Sci. USA, 81:6851 (1984)), or by covalently linking all or part of the coding sequence of a non-immunoglobulin polypeptide to the immunoglobulin coding sequence. Typically, such a non-immunoglobulin polypeptide is used in place of the constant domains of an antibody or in place of the variable domains of one antigen-binding site of an antibody to create a chimeric bivalent antibody containing one antigen-binding site with specificity for an antigen and another antigen-binding site with specificity for a different antigen.

[0378] The monoclonal antibodies described herein may be monovalent, and their preparation is well known in the art. For example, one method involves recombinant expression of immunoglobulin light chains and modified heavy chains. The heavy chain is generally truncated at any point in the Fc region to prevent heavy chain cross-linking. Alternatively, the relevant cysteine ​​residues may be replaced with another amino acid residue or deleted to prevent cross-linking. In vitro methods are also suitable for preparing monovalent antibodies. Digestion of antibodies to produce their fragments, particularly Fab fragments, can be accomplished using routine techniques known in the art.

[0379] Chimeric or hybrid antibodies can also be prepared in vitro using known methods in synthetic protein chemistry, including those involving crosslinking agents.For example, immunotoxins can be constructed using disulfide exchange reactions or by forming thioether bonds.Examples of suitable reagents for this purpose include iminothiolate and methyl-4-mercaptobutyrimidate.

[0380] Recombinant production in prokaryotic cells a) Vector construction Polynucleic acid sequences encoding the antibodies of the present application can be obtained using standard recombinant techniques. The desired polynucleic acid sequences can be isolated and sequenced from antibody producing cells, such as hybridoma cells. Alternatively, polynucleotides can be synthesized using a nucleotide synthesizer or PCR techniques. Once obtained, the sequences encoding the polypeptides are inserted into a recombinant vector capable of replicating and expressing heterologous polynucleotides in a prokaryotic host. For the purposes of the present invention, many vectors available and known in the art can be used. The selection of an appropriate vector depends primarily on the size of the nucleic acid to be inserted into the vector and the specific host cell to be transformed with the vector. Each vector contains various components depending on its function (amplification or expression of heterologous polynucleotides, or both) and its compatibility with the specific host cell in which it resides. Vector components generally include, but are not limited to, an origin of replication, a selectable marker gene, a promoter, a ribosome binding site (RBS), a signal sequence, a heterologous nucleic acid insert, and a transcription termination sequence.

[0381] The present invention provides an expression system that allows the quantitative ratio of expressed polypeptide components to be adjusted to maximize the yield of secreted and properly assembled antibodies of the present application. Such adjustment is achieved at least in part by simultaneously adjusting the translation strength of the polypeptide components. One technique for adjusting translation strength is disclosed in Simmons et al., U.S. Pat. No. 5,840,523. It utilizes variants of the translation initiation region (TIR) ​​within a cistron. For a given TIR, a series of amino acid or nucleic acid sequence variants can be made with a range of translation strengths, thereby providing a convenient means of tuning this factor to the desired expression level of a particular chain. TIR variants can be generated by conventional mutagenesis techniques that result in codon changes that can change the amino acid sequence, although silent changes in the nucleic acid sequence are preferred. Changes in the TIR can include, for example, changes in the number or spacing of Shine-Dalgarno sequences, along with changes in the signal sequence. One method for generating mutant signal sequences is to generate a "codon bank" at the beginning of the coding sequence that does not change the amino acid sequence of the signal sequence (i.e., the changes are silent). This can be accomplished by altering the third nucleotide position of each codon. Additionally, some amino acids, such as leucine, serine, and arginine, have multiple first and second positions which can add complexity when generating banks.

[0382] Preferably, a set of vectors is generated that has a range of TIR strengths for each cistron therein. This limited set provides a comparison of the expression level of each chain as well as the yield of the desired protein product under various TIR strength combinations. TIR strength can be determined by quantifying the expression level of a reporter gene, as described in detail in Simmons et al., U.S. Patent No. 5,840,523. Based on the comparison of translation strengths, the desired individual TIRs are selected to be combined in the expression vector construct of the present application.

[0383] b) Prokaryotic host cells Suitable prokaryotic host cells for expressing the antibodies of the present application include Archaea and Eubacteria, such as Gram-negative or Gram-positive organisms. Examples of useful bacteria include Escherichia (e.g., E. coli), Bacilli (e.g., B. subtilis), Enterobacteria, Pseudomonas species (e.g., P. aeruginosa), Salmonella typhimurium, Serratia marcescans, Klebsiella, Proteus, Shigella, Rhizobia, Vitreoscilla, or Paracoccus. In some embodiments, Gram-negative cells are used. In some embodiments, E. coli cells are used as hosts for the present invention. Examples of E. coli strains include strain W3110 and its derivatives, with the genotypes W3110 AfhuA(AtonA)ptr3 lac Iq lacL8 AompT A(nmpc-fepE)degP41 kan R (U.S. Patent No. 5,639,635). Other strains such as E. coli 294 (ATCC 31,446), E. coli B, E. coli 1776 (ATCC 31,537) and E. coli RV308 (ATCC 31,608) and their derivatives are also suitable. These examples are illustrative and not limiting. In general, the appropriate bacterium should be selected taking into account the replicability of the replicon in the bacterial cell. For example, E. coli, Serratia or Salmonella species can be suitably used as hosts when the replicon is supplied using well-known plasmids such as pBR322, pBR325, pACYC177 or pKN410.

[0384] Typically, the host cell should secrete minimal amounts of proteolytic enzymes and additional protease inhibitors may desirably be incorporated into the cell culture.

[0385] c) Protein Production Host cells are transformed with the above expression vectors and cultured in conventional nutrient media modified as appropriate for inducing promoters, selecting transformants, or amplifying genes encoding the desired sequences. Transformation refers to the introduction of DNA into a prokaryotic host so that the DNA is replicable, either as an extrachromosomal element or by chromosomal integrant. Depending on the host cell used, transformation is performed using standard techniques appropriate for such cells. Calcium treatment using calcium chloride is commonly used for bacterial cells that contain substantial cell wall barriers. Another method for transformation uses polyethylene glycol / DMSO. Yet another technique used is electroporation.

[0386] Host cells are transformed with the above expression vectors and cultured in conventional nutrient media modified as appropriate for inducing promoters, selecting transformants, or amplifying genes encoding the desired sequences. Transformation refers to the introduction of DNA into a prokaryotic host so that the DNA is replicable, either as an extrachromosomal element or by chromosomal integrant. Depending on the host cell used, transformation is performed using standard techniques appropriate for such cells. Calcium treatment using calcium chloride is commonly used for bacterial cells that contain substantial cell wall barriers. Another method for transformation uses polyethylene glycol / DMSO. Yet another technique used is electroporation.

[0387] The prokaryotic cells used to produce the antibodies of the present application are grown in a medium known in the art and suitable for the culture of the selected host cells. An example of a suitable medium includes Luria Broth (LB) + necessary nutritional supplements. In some embodiments, the medium also contains a selection agent selected based on the construction of the expression vector to selectively allow the growth of prokaryotic cells containing the expression vector. For example, ampicillin is added to the medium for the growth of cells expressing an ampicillin resistance gene.

[0388] d) Protein purification The constructs produced herein or portions thereof can be further purified to obtain substantially homogeneous preparations for further assays and uses. Standard protein purification methods known in the art can be used. The following procedures are examples of suitable purification procedures: fractionation on immunoaffinity or ion exchange columns, ethanol precipitation, reverse phase HPLC, chromatography on silica or cation exchange resins, such as DEAE, chromatofocusing, SDS-PAGE, ammonium sulfate precipitation, and gel filtration, such as using Sephadex G-75.

[0389] In some embodiments, Protein A immobilized on a solid phase is used for immunoaffinity purification of antibodies comprising the Fc region of the present application. Protein A is a 411 (D cell wall protein from Staphylococcus aureas that binds with high affinity to the Fc region of antibodies (Lindmark et al (1983) J. Immunol. Meth. 62:1-13). The solid phase to which Protein A is immobilized is preferably a column comprising a glass or silica surface, more preferably a controlled pore glass column or a silicic acid column. In some applications, the column is coated with a reagent such as glycerol to prevent nonspecific attachment of contaminants. The solid phase is then washed to remove contaminants nonspecifically bound to the solid phase. Finally, the antibody of interest is recovered from the solid phase by elution.

[0390] Recombinant production in eukaryotic cells For eukaryotic expression, the vector components generally include, but are not limited to, a signal sequence, an origin of replication, one or more marker genes, and one or more of an enhancer element, a promoter, and a transcription termination sequence.

[0391] a) Signal Sequence Component Vectors for use with eukaryotic hosts may also have an insert encoding a signal sequence or other polypeptide having a specific cleavage site at the N-terminus of the mature protein or polypeptide. The heterologous signal sequence selected is preferably one that is recognized and processed (i.e., cleaved by a signal peptidase) by the host cell. In mammalian cell expression, mammalian signal sequences as well as viral secretory leaders, such as the herpes simplex gD signal, are available.

[0392] The DNA for such precursor region is ligated in reading frame to DNA encoding the antibody of the present application.

[0393] b) Origin of replication Generally, the origin of replication component is not needed for mammalian expression vectors (the SV40 origin may typically be used only because it contains the early promoter).

[0394] c) Selective Gene Components Expression and cloning vectors may contain a selection gene, also called a selectable marker. Typical selection genes encode proteins that (a) confer resistance to antibiotics or other toxins, such as ampicillin, neomycin, methotrexate, or tetracycline, (b) complement an auxotrophic deficiency, or (c) supply vital nutrients unavailable from complex media (e.g., the gene encoding D-alanine racemase in Bacillus).

[0395] d) Promoter Component Expression and cloning vectors usually contain a promoter that is recognized by the host organism and is operably linked to a nucleic acid encoding a desired polypeptide sequence. Virtually all eukaryotic genes have an AT-rich region located about 25-30 bases upstream from the site where transcription begins. Another sequence found 70-80 bases upstream from the start of transcription of many genes is a CNCAAT region (N can be any nucleotide). At the 3' end of most eukaryotic genes is an AATAAA sequence that may be a signal for the addition of a poly A tail to the 3' end of the coding sequence. All of these sequences can be inserted into eukaryotic expression vectors.

[0396] Other promoters suitable for use with prokaryotic hosts include the phoA promoter, -lactamase and lactose promoter systems, alkaline phosphatase promoter, tryptophan (trp) promoter system, and hybrid promoters such as the tac promoter. However, other known bacterial promoters are suitable. Promoters for use in bacterial systems also contain a Shine-Dalgarno (SD) sequence operably linked to the DNA encoding the antibody.

[0397] Transcription of the polypeptide from the vector in a mammalian host cell is controlled by, for example, a promoter derived from the genome of a virus such as polyoma virus, fowlpox virus, adenovirus (such as adenovirus 2), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, retrovirus, Hepatitis B virus, and most preferably Simian Virus 40 (SV40), a heterologous mammalian promoter, such as a promoter derived from the actin promoter or an immunoglobulin promoter, a heat shock promoter, provided that such a promoter is compatible with the host cell system.

[0398] e) Enhancer element components Transcription of DNA encoding the antibody of the present application by higher eukaryotes is often increased by inserting an enhancer sequence into the vector. Currently, many enhancer sequences are known from mammalian genes (globin, elastase, albumin, α-fetoprotein, insulin). Typically, however, enhancers from eukaryotic cell viruses are used. Examples include the SV40 enhancer on the late side of the replication origin (100-270 bp), the cytomegalovirus early promoter enhancer, the polyoma enhancer on the late side of the replication origin, and adenovirus enhancers. The enhancer can be spliced ​​into the vector at a position 5' or 3' to the polypeptide-encoding sequence, but is preferably located at a site 5' from the promoter.

[0399] f) Transcription termination component Expression vectors used in eukaryotic host cells (yeast, fungi, insects, plants, animals, humans, or nucleated cells from other multicellular organisms) also contain sequences necessary for the termination of transcription and stabilization of mRNA. Such sequences are commonly available from the 5' and sometimes 3' untranslated regions of eukaryotic or viral DNA or cDNA. These regions contain nucleotide segments that are transcribed as polyadenylated fragments in the untranslated portion of the polypeptide-encoding mRNA. One useful transcription termination component is the bovine growth hormone polyadenylation region. See WO 94 / 11026 and the expression vectors disclosed therein.

[0400] g) Selection and transformation of host cells Suitable host cells for cloning or expressing the DNA in the vectors herein include the higher eukaryotic cells described herein, including vertebrate host cells. Propagation of vertebrate cells in culture (tissue culture) has become a routine procedure. Examples of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7, ATCC CRL 1651); human embryonic kidney line (293 or 293 cells subcloned for growth in suspension culture, Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK, ATCC CCL 10); Chinese hamster ovary cells / -DHFR (CHO, Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)); mouse Sertoli cells (TM4, Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1 ATCC CCL 70); African green monkey kidney cells (VERO-76, ATCC CRL-1587); human cervical carcinoma cells (HELA, ATCC CCL 2); canine kidney cells (MDCK, ATCC CCL 34); buffalo rat hepatocytes (BRL 3A, ATCC CRL 1442); human lung cells (W138, ATCC CCL 75); human hepatocytes (Hep G2, HB 8065); mouse mammary tumor (MMT 060562, ATCC CCL51); TR1 cells (Mather et al., Annals NYAcad. Sci. 383:44-68 (1982)); MRC 5 cells; FS4 cells; and a human hepatoma line (Hep G2).

[0401] Host cells are transformed with the above-described expression or cloning vectors for antibody production and cultured in conventional nutrient media modified as appropriate for inducing promoters, selecting transformants, or amplifying the genes encoding the desired sequences.

[0402] h) Cultivation of host cells The host cells used to produce the antibodies of the present application may be cultured in a variety of media. Commercially available media such as Ham's F10 (Sigma), Minimum Essential Medium ((MEM), (Sigma), RPMI-1640 (Sigma), and Dulbecco's Modified Eagle's Medium ((DMEM), Sigma) are suitable for culturing the host cells. The media may be supplemented as necessary with hormones and / or other growth factors (such as insulin, transferrin, or epidermal growth factor), salts (such as sodium chloride, calcium, magnesium, phosphates, etc.), buffers (such as HEPES), nucleotides (such as adenosine and thymidine), antibiotics (such as GENTAMYCIN™ drugs), trace elements (usually defined as inorganic compounds present at final concentrations in the micromolar range), and glucose or an equivalent energy source. Any other necessary supplements may also be included at appropriate concentrations that would be known to one of skill in the art. Culture conditions such as temperature, pH, etc. will be those previously used with the host cell selected for expression and will be apparent to one of skill in the art.

[0403] In some embodiments, a glutamine synthetase (GS)-CHO expression system is used to culture host cells and / or express the anti-IGFBP7 constructs or portions thereof described herein. See, e.g., Fan et al., J Biotechnol. 2013 Dec;168(4):652-8.

[0404] i) Protein purification When using recombinant techniques, antibodies can be produced intracellularly, in the periplasmic space, or directly secreted into the medium. If the antibody is produced intracellularly, as a first step, particulate debris, either host cells or lysed fragments, is removed, for example, by centrifugation or ultrafiltration. Carter et al., Bio / Technology 10:163-167 (1992) describes a procedure for isolating antibodies secreted into the periplasmic space of E. coli. Briefly, cell paste is thawed in the presence of sodium acetate (pH 3.5), EDTA, and phenylmethylsulfonyl fluoride (PMSF) for about 30 minutes. Cell debris can be removed by centrifugation. If the antibody is secreted into the medium, the supernatant from such expression systems is generally first concentrated using a commercially available protein concentration filter, for example, an Amicon or Millipore Pellicon ultrafiltration unit. A protease inhibitor such as PMSF may be included in any of the foregoing steps to inhibit proteolysis and antibiotics may be included to prevent the growth of adventitious contaminants.

[0405] Protein compositions prepared from cells can be purified using, for example, hydroxylapatite chromatography, gel electrophoresis, dialysis, multimodal chromatography, ion exchange chromatography, and affinity chromatography, with affinity chromatography being the preferred purification technique. The suitability of Protein A as an affinity ligand depends on the species and isotype of any immunoglobulin Fc domain present in the antibody. Protein A can be used to purify antibodies based on human immunoglobulins containing 1, 2, or 4 heavy chains. Protein G is recommended for all mouse isotypes and human 3. The matrix to which the affinity ligand is attached is most often agarose, although other matrices are available. Mechanically stable matrices such as controlled pore glass or poly(styrene-divinyl)benzene allow for faster flow rates and shorter processing times than can be achieved with agarose. If the antibody is C HIf three domains are involved, Bakerbond ABX™ resin (JT Baker, Phillipsburg, NJ) is useful for purification. Other techniques for protein purification, such as ultrafiltration / diafiltration (UF / DF), fractionation on ion exchange columns, ethanol precipitation, reversed-phase HPLC, chromatography on silica, chromatography on heparin, SEPHAROSE™ chromatography on anion or cation exchange resins (e.g., polyaspartic acid columns), chromatography on hydrophobic resins, chromatography on mixed-mode resins, chromatofocusing, SDS-PAGE, and ammonium sulfate precipitation, are also available depending on the antibody to be recovered.

[0406] Following any preliminary purification step(s), the mixture containing the antibody of interest and contaminants can be subjected to low pH hydrophobic interaction chromatography, preferably performed at low salt concentrations (e.g., about 0-0.25 M salt), using an elution buffer at a pH of about 2.5-4.5. Ion exchange is often also used for the polishing step.

[0407] Humanized antibodies Humanized forms of non-human antibodies include chimeric immunoglobulins, immunoglobulin chains or fragments thereof (e.g., Fv, Fab, Fab', F(ab')) that contain minimal sequence derived from non-human immunoglobulin. 2or other antigen-binding subsequences of antibodies). Humanized antibodies include human immunoglobulins (recipient antibody) in which residues from a CDR of the recipient are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat, rabbit, camel or llama having the desired specificity, affinity, and capacity. In some instances, Fv framework residues of the human immunoglobulin are replaced by corresponding non-human residues. Humanized antibodies can also comprise residues that are found neither in the recipient antibody nor in the imported CDR or framework sequences. In general, humanized antibodies can comprise substantially all of at least one, and typically two, variable domains, with all or substantially all of the CDR regions corresponding to those of a non-human immunoglobulin and all or substantially all of the FR regions being those of a human immunoglobulin consensus sequence. In some embodiments, the humanized antibody comprises at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. See, e.g., Jones et al., Nature, 321:522-525 (1986); Riechmann et al., Nature, 332:323-329 (1988); Presta, Curr. Op. Struct. Biol., 2:593-596 (1992).

[0408] Methods for humanizing non-human antibodies are well known in the art. Generally, a humanized antibody has one or more amino acid residues introduced from a source that is non-human. These non-human amino acid residues are often called "import" residues, which are typically taken from an "import" variable domain. Humanization can essentially be performed according to the method of Winter and coworkers (Jones et al., Nature 321:522-525 (1986); Riechmann et al., Nature 332:323-327 (1988); Verhoeyen et al., Science 239:1534-1536 (1988)), or by substituting rodent CDRs or CDR sequences for the corresponding sequences of a human antibody. Thus, such "humanized" antibodies are chimeric antibodies (U.S. Patent No. 4,816,567) in which substantially less than an intact human variable domain is replaced by the corresponding sequence from a non-human species. In practice, humanized antibodies are typically human antibodies in which some CDR residues and possibly some FR residues are substituted by residues from analogous sites in rodent antibodies.

[0409] The choice of human variable domains (both light and heavy) used in making humanized antibodies is very important to reduce antigenicity. According to the so-called "best-fit" method, the sequence of the variable domain of a rodent antibody is screened against the entire library of known human variable domain sequences. The human sequence that is closest to the rodent sequence is then accepted as the human framework (FR) of the humanized antibody. Sims et al., J.Immunol., 151:2296 (1993); Chothia et al., J.Mol.Biol., 196:901 (1987). Another method uses a specific framework derived from the consensus sequence of all human antibodies of a specific subgroup of light or heavy chains. The same framework can be used for several different humanized antibodies.

[0410] It is further important that antibodies be humanized with retention of high affinity for the antigen and other favorable biological properties. To this end, according to a preferred method, humanized antibodies are prepared by a process of analyzing the parental sequences and various conceptual humanized products using three-dimensional models of the parental and humanized sequences. Three-dimensional immunoglobulin models are commonly available and are familiar to those skilled in the art. Computer programs are available which illustrate and display probable three-dimensional conformational structures of selected candidate immunoglobulin sequences. Inspection of these displays allows analysis of the likely role of the residues in the functioning of the candidate immunoglobulin sequence, i.e., analysis of residues that influence the ability of the candidate immunoglobulin to bind to its antigen. In this way, FR residues can be selected and combined from the recipient and import sequences such that the desired antibody characteristic, such as increased affinity for the target antigen(s), is achieved. In general, the CDR residues are directly and most substantially involved in influencing antigen binding.

[0411] In some embodiments, the sdAb is modified, such as humanized, to reduce its immunogenicity with respect to the heterologous species without reducing the domain's natural affinity for the antigen. For example, the antibody variable domain (V H H) amino acid residues can be determined such that, for example, one or more of the camelid amino acids in the framework regions are replaced by their human counterparts found in the human consensus sequence without the polypeptide losing its typical characteristics, i.e., the humanization does not significantly affect the antigen-binding capacity of the resulting polypeptide. Humanization of camelid sdAbs requires the introduction and mutagenesis of a limited amount of amino acids in a single polypeptide chain. This is in contrast to humanization of scFv, Fab', (Fab')2 and IgG, which requires the introduction of amino acid changes in two chains, light and heavy, and the preservation of the assembly of both chains.

[0412] Human antibodies As an alternative to humanization, human antibodies can be generated. For example, it is now possible to produce transgenic animals (e.g., mice) that, upon immunization, are capable of producing a full repertoire of human antibodies in the absence of endogenous immunoglobulin production. For example, the expression of the antibody heavy chain joining region (J) in chimeric and germ-line mutant mice has been demonstrated. H It has been described that homozygous deletion of the .) gene results in complete inhibition of endogenous antibody production. Transfer of the human germ-line immunoglobulin gene array into such germ-line mutant mice results in the production of human antibodies upon antigen challenge.

[0413] Alternatively, phage display technology can be used to identify human antibodies and antibody fragments in vitro from immunoglobulin variable (V) domain gene repertoires from unimmunized donors. McCafferty et al., Nature 348:552-553 (1990); Hoogenboom and Winter, J. Mol. Biol. 227:381 (1991).

[0414] Human antibodies can also be made by in vitro activated B cells (see U.S. Pat. Nos. 5,567,610 and 5,229,275) or by using a variety of techniques known in the art, including phage display libraries (Hoogenboom and Winter, J. Mol. Biol., 227:381 (1991); Marks et al., J. Mol. Biol., 222:581 (1991).

[0415] Nucleic acid molecules encoding antibody portions In some embodiments, a polynucleotide is provided that encodes any one of the anti-IGFBP7 constructs or antibody moieties described herein. In some embodiments, a polynucleotide is provided that is prepared using any one of the methods as described herein. In some embodiments, a nucleic acid molecule comprises a polynucleotide that encodes any one of the anti-IGFBP7 single domain antibody (sdAb) moieties described herein. In some embodiments, a polynucleotide comprises a nucleotide sequence that encodes a leader sequence that is located at the N-terminus of the sdAb when translated.

[0416] In some embodiments, the polynucleotide is DNA. In some embodiments, the polynucleotide is RNA. In some embodiments, the RNA is mRNA.

[0417] The nucleic acid molecule can be constructed using recombinant DNA techniques conventional in the art. In some embodiments, the nucleic acid molecule is an expression vector suitable for expression in a selected host cell.

[0418] nucleic acid construct In some embodiments, a nucleic acid construct is provided that includes any one of the polynucleotides described herein. In some embodiments, a nucleic acid construct is provided that is prepared using any of the methods described herein.

[0419] In some embodiments, the nucleic acid construct further comprises a promoter operably linked to the polynucleotide, hi some embodiments, the polynucleotide corresponds to a gene and the promoter is the wild-type promoter of the gene.

[0420] vector In some embodiments, a vector is provided that includes any polynucleotide encoding any one of the antibody moieties (e.g., anti-IGFBP7 antibody moieties) or nucleic acid constructs described herein. In some embodiments, a vector is provided that is prepared using any method described herein. Also provided is a vector that includes a polynucleotide encoding any one of the anti-IGFBP7 constructs, such as anti-IGFBP7 sdAbs, fusion proteins, or other forms of constructs described herein. Such vectors include, but are not limited to, DNA vectors, phage vectors, viral vectors, retroviral vectors, and the like.

[0421] In some embodiments, a vector is selected that is optimized for expression of a polypeptide in CHO or CHO-derived cells or NSO cells. Exemplary such vectors are described, for example, in Running Deer et al., Biotechnol. Prog. 20:880-889 (2004).

[0422] host cell In some embodiments, a host cell is provided that comprises any of the polypeptides, nucleic acid constructs and / or vectors described herein. In some embodiments, a host cell is provided that is prepared using any of the methods described herein. In some embodiments, the host cell is capable of producing any of the antibody portions described herein under fermentation conditions.

[0423] In some embodiments, the antibody moieties (e.g., anti-IGFBP7 antibody moieties) described herein may be expressed in prokaryotic cells, e.g., bacterial cells; or eukaryotic cells, e.g., fungal cells (such as yeast), plant cells, insect cells, and mammalian cells. Such expression may be performed, for example, according to procedures known in the art. Exemplary eukaryotic cells that may be used to express the polypeptides include, but are not limited to, COS cells, including COS7 cells; 293 cells, including 293-6E cells; CHO-S, DG44; CHO cells, including Lec13 CHO cells, FUT8 CHO cells, and CHO GS cells (Sigma); PER.C6® cells (Crucell); and NSO cells. In some embodiments, the antibody moieties (e.g., anti-IGFBP7 antibody moieties) described herein may be expressed in yeast. See, e.g., U.S. Patent Application Publication No. 2006 / 0270045. In some embodiments, a particular eukaryotic host cell is selected based on its ability to effect desired post-translational modifications on the antibody moiety or construct. For example, in some embodiments, CHO cells produce polypeptides that have a higher level of sialylation than the same polypeptides produced in 293 cells.

[0424] Introduction of one or more nucleic acids into a desired host cell can be accomplished by any method, including, but not limited to, calcium phosphate transfection, DEAE-dextran mediated transfection, cationic lipid mediated transfection, electroporation, transduction, infection, etc. Non-limiting exemplary methods are described, for example, in Sambrook et al., Molecular Cloning, A Laboratory Manual, 3rd ed. Cold Spring Harbor Laboratory Press (2001). Nucleic acids can be transiently or stably transfected into a desired host cell according to any suitable method.

[0425] The present application also provides a host cell comprising any of the polynucleotides or vectors described herein. In some embodiments, the present invention provides a host cell comprising an anti-IGFBP7 antibody. Any host cell capable of overexpressing heterologous DNA can be used for the purpose of isolating the gene encoding the antibody, polypeptide or protein of interest. Non-limiting examples of mammalian host cells include, but are not limited to, COS, HeLa and CHO cells. See also PCT Publication No. WO87 / 04462. Suitable non-mammalian host cells include prokaryotes (e.g., E. coli or B. subtillis) and yeast (e.g., S. cerevisae, S. pombe; or K. lactis).

[0426] In some embodiments, the antibody portion is produced in a cell-free system. Non-limiting exemplary cell-free systems are described, for example, in Sitaraman et al., Methods Mol. Biol. 498:229-44 (2009); Spirin, Trends Biotechnol. 22:538-45 (2004); Endo et al., Biotechnol. Adv. 21:695-713 (2003).

[0427] Culture medium In some embodiments, a culture medium is provided that comprises any of the antibody portions, polynucleotides, nucleic acid constructs, vectors, and / or host cells described herein. In some embodiments, a culture medium is provided that is prepared using any of the methods described herein.

[0428] In some embodiments, the medium contains hypoxanthine, aminopterin and / or thymidine (e.g., HAT medium). In some embodiments, the medium does not contain serum. In some embodiments, the medium is chemically defined. In some embodiments, the medium contains serum. In some embodiments, the medium is D-MEM or RPMI-1640 medium.

[0429] Purification of antibody moieties Anti-IGFBP7 constructs (e.g., anti-IGFBP7 monoclonal or multispecific antibodies) can be purified by any suitable method. Such methods include, but are not limited to, the use of affinity matrices or hydrophobic interaction chromatography. Suitable affinity ligands include ligands that bind to ROR1 ECD and antibody constant regions. For example, protein A, protein G, protein A / G, antibody affinity columns, or ultrafiltration / diafiltration (UF / DF) can be used to bind to the constant region and purify anti-IGFBP7 constructs that contain Fc fragments. Hydrophobic interaction chromatography, such as butyl or phenyl columns, can also be suitable for the purification of some polypeptides, such as antibodies. Ion exchange chromatography (e.g., anion exchange chromatography and / or cation exchange chromatography) can also be suitable for the purification of some polypeptides, such as antibodies. Mixed-mode chromatography (e.g., reversed-phase / anion exchange, reversed-phase / cation exchange, hydrophilic interaction / anion exchange, hydrophilic interaction / cation exchange, etc.) may also be suitable for the purification of some polypeptides, such as antibodies. Many methods of purifying polypeptides are known in the art.

[0430] V. Treatment Method Also provided herein is a method of treating a disease or condition in an individual, the method comprising administering to the individual (e.g., a mammal, such as a human) an anti-IGFBP7 construct described herein.

[0431] In some embodiments, a method is provided for treating a disease or condition (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, comprising administering to an individual an effective amount of an anti-IGFBP7 construct (such as any of the anti-IGFBP7 constructs described herein).

[0432] In some embodiments, methods are provided for inhibiting abnormal blood vessel growth in a tissue in an individual, comprising administering to the individual an effective amount of an anti-IGFBP7 construct (such as any of the anti-IGFBP7 constructs described herein). In some embodiments, the individual has cancer (such as a solid tumor). In some embodiments, the tissue is a cancer / tumor tissue.

[0433] In some embodiments, there is provided a method of treating a disease or condition (e.g., cancer, e.g., a solid tumor) in an individual or inhibiting abnormal blood vessel growth in a tissue, the method comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises the amino acid sequence of any of SEQ ID NOs: 32-33 and 46-48, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0434] In some embodiments, there is provided a method of treating a disease or condition in an individual (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises an amino acid sequence of any of SEQ ID NOs: 34 and 49-51, or a variant thereof having at least about 80% sequence identity (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%).

[0435] In some embodiments, there is provided a method of treating a disease or condition (e.g., cancer, e.g., a solid tumor) in an individual or inhibiting abnormal blood vessel growth in a tissue, the method comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises the amino acid sequence of any of SEQ ID NOs:33 and 35-57, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0436] In some embodiments, there is provided a method of treating a disease or condition in an individual (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises the amino acid sequence of any of SEQ ID NOs: 38 and 39, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0437] In some embodiments, there is provided a method of treating a disease or condition in an individual (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 or anti-PD-1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:1 or 2, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises an amino acid sequence of any of SEQ ID NOs: 32-33 and 46-48, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0438] In some embodiments, there is provided a method of treating a disease or condition in an individual (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, the method comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 or anti-PD-1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises an amino acid sequence of any of SEQ ID NOs: 34 and 49-51, or a variant thereof having at least about 80% sequence identity (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%).

[0439] In some embodiments, methods are provided for treating a disease or condition (e.g., cancer, e.g., a solid tumor) in an individual or inhibiting abnormal blood vessel growth in a tissue, comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 or anti-PD-1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises an amino acid sequence of any of SEQ ID NOs:35-37, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0440] In some embodiments, methods are provided for treating a disease or condition in an individual (e.g., cancer, e.g., a solid tumor) or inhibiting abnormal blood vessel growth in a tissue, comprising administering to the individual an effective amount of an anti-IGFBP7 construct comprising an anti-IGFBP7 antibody portion (e.g., any of the antibody portions described herein) fused to an anti-PD-L1 or anti-PD-1 full length antibody, wherein the anti-IGFBP7 antibody portion comprises a single domain antibody (sdAb) portion comprising a CDR1 comprising the amino acid sequence of SEQ ID NO: 10 or 11, a CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in the CDRs. In some embodiments, the sdAb portion comprises the amino acid sequence of any of SEQ ID NOs: 38 and 39, or a variant thereof having at least about 80% (e.g., at least about any one of 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) sequence identity.

[0441] In some embodiments, the above amino acid substitutions are limited to the "exemplary substitutions" shown in Table 2 of the present application. In some embodiments, the amino acid substitutions are limited to the "preferred substitutions" shown in Table 2 of the present application.

[0442] Disease or condition The methods described herein are applicable to any disease or condition associated with abnormal vasculature. In some embodiments, the disease or condition is age-related macular degeneration (ARMD). In some embodiments, the disease or condition is skin psoriasis. In some embodiments, the disease or condition is a benign tumor. In some embodiments, the disease or condition is cancer.

[0443] cancer In some embodiments, the disease ...

Claims

1. 1. An anti-IGFBP7 construct comprising a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, said sdAb portion comprising: 1) a CDR1 comprising the amino acid sequence of SEQ ID NO:1, a CDR2 comprising the amino acid sequence of SEQ ID NO:3, and a CDR3 comprising the amino acid sequence of SEQ ID NO:4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 2) a CDR1 comprising the amino acid sequence of SEQ ID NO: 2 or 112, a CDR2 comprising the amino acid sequence of SEQ ID NO: 3, and a CDR3 comprising the amino acid sequence of SEQ ID NO: 4 or 113, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 3) a CDR1 comprising the amino acid sequence of SEQ ID NO:5 or 114, a CDR2 comprising the amino acid sequence of SEQ ID NO:6, and a CDR3 comprising the amino acid sequence of SEQ ID NO:7, 115 or 116, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitution in said CDRs; 4) a CDR1 comprising the amino acid sequence of SEQ ID NO:8, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 5) a CDR1 comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:12, and a CDR3 comprising the amino acid sequence of SEQ ID NO:13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 6) CDR1 comprising the amino acid sequence of SEQ ID NO: 10, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 7) CDR1 comprising the amino acid sequence of SEQ ID NO: 11, CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and CDR3 comprising the amino acid sequence of SEQ ID NO: 13, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 8) CDR1 comprising the amino acid sequence of SEQ ID NO: 14, CDR2 comprising the amino acid sequence of SEQ ID NO: 15, and CDR3 comprising the amino acid sequence of SEQ ID NO: 16, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 9) CDR1 comprising the amino acid sequence of SEQ ID NO: 17, CDR2 comprising the amino acid sequence of SEQ ID NO: 18, and CDR3 comprising the amino acid sequence of SEQ ID NO: 19, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 10) CDR1 comprising the amino acid sequence of SEQ ID NO: 20, CDR2 comprising the amino acid sequence of SEQ ID NO: 21, and CDR3 comprising the amino acid sequence of SEQ ID NO: 22, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 11) CDR1 comprising the amino acid sequence of SEQ ID NO: 23, CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and CDR3 comprising the amino acid sequence of SEQ ID NO: 25, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; 12) CDR1 comprising the amino acid sequence of SEQ ID NO: 26, CDR2 comprising the amino acid sequence of SEQ ID NO: 27, and CDR3 comprising the amino acid sequence of SEQ ID NO: 28, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs; or 13) An anti-IGFBP7 construct comprising a CDR1 comprising the amino acid sequence of SEQ ID NO:29, a CDR2 comprising the amino acid sequence of SEQ ID NO:30, and a CDR3 comprising the amino acid sequence of SEQ ID NO:31, or a variant thereof comprising up to 5, 4, 3, 2, or 1 amino acid substitutions in said CDRs.

2. An anti-IGFBP7 construct comprising a polypeptide comprising a single domain antibody (sdAb) portion that specifically recognizes IGFBP7, said sdAb portion comprising the amino acid sequences of CDR1, CDR2, and CDR3 within the amino acid sequence set forth in any of SEQ ID NOs: 32-51.

3. 3. The anti-IGFBP7 construct of claim 1 or 2, wherein the sdAb portion comprises the amino acid sequence of any one of SEQ ID NOs: 32-51, or a variant thereof having at least about 80% sequence identity to any one of SEQ ID NOs: 32-51.

4. The anti-IGFBP7 construct of any one of claims 1 to 3, wherein the sdAb portion is camelid, chimeric, human, partially humanized, or fully humanized.

5. The sdAb portion is V Ｈ The anti-IGFBP7 construct according to any one of claims 1 to 4, which is an H antibody.

6. An anti-IGFBP7 construct according to any one of claims 1 to 5, which blocks the binding of CD93 to IGFBP7.

7. The anti-IGFBP7 construct according to any one of claims 1 to 6, wherein said IGFBP7 is human IGFBP7.

8. The anti-IGFBP7 construct of claim 6 or claim 7, wherein the CD93 is human CD93.

9. The anti-IGFBP7 construct of any one of claims 1 to 8, further comprising a second moiety.

10. The anti-IGFBP7 construct of claim 9 , wherein the second portion comprises an antibody portion that specifically recognizes an antigen.

11. The anti-IGFBP7 construct of claim 10, wherein the antigen is PD-L1 or PD-1.

12. The second antibody portion may be a full length antibody, a Fab, a Fab', a (Fab') ２ , Fv, single chain Fv (scFv), scFv-scFv, minibody, diabody, or sdAb.

13. The anti-IGFBP7 construct of claim 9 , wherein the second portion comprises a half-life extending moiety.

14. The anti-IGFBP7 construct of claim 9, wherein the construct is an antibody-drug conjugate.

15. An anti-IGFBP7 construct that specifically binds to IGFBP7 in a competitive manner with the anti-IGFBP7 construct of any one of claims 1 to 14.

16. A pharmaceutical composition comprising an anti-IGFBP7 construct according to any one of claims 1 to 15 and a pharma- ceutically acceptable carrier.

17. A polynucleotide encoding a polypeptide or a part thereof of the anti-IGFBP7 construct according to any one of claims 1 to 15.

18. 20. A nucleic acid construct comprising the polynucleotide of claim 17, optionally further comprising a promoter operably associated with said polynucleotide.

19. A vector comprising the nucleic acid construct of claim 18.

20. 20. An isolated host cell comprising the polynucleotide of claim 17, the nucleic acid construct of claim 18 or the vector of claim 19.

21. A culture medium comprising a polypeptide of an anti-IGFBP7 construct according to any one of claims 1 to 15, a polynucleotide according to claim 17, a nucleic acid construct according to claim 18, a vector according to claim 19, or a host cell according to claim 20.

22. 1. A method for producing an anti-IGFBP7 construct, comprising: a) culturing the isolated host cell of claim 20 under conditions effective to express said polypeptide; and b) obtaining said polypeptide from said host cell.

23. A composition comprising an anti-IGFBP7 construct according to any one of claims 1 to 15, or a pharmaceutical composition according to claim 16, for treating a disease or condition in an individual or inhibiting abnormal blood vessel growth in a tissue.

24. The composition of claim 23 , wherein the disease or condition is associated with abnormal vasculature.

25. 25. The composition of claim 23 or claim 24, wherein the disease or condition is cancer.

26. The composition of claim 25 , wherein the cancer is a solid tumor.

27. The composition of claim 25 or claim 26, wherein the cancer comprises CD93+ endothelial cells.

28. The composition of any one of claims 25 to 27, wherein the cancer comprises IGFBP7+ blood vessels.

29. The composition of any one of claims 25 to 28, wherein the cancer is characterized by tumor hypoxia.

30. The composition of any one of claims 25 to 29, wherein the cancer is locally advanced or metastatic cancer.

31. 31. The composition of any one of claims 25 to 30, wherein the cancer is selected from the group consisting of lymphoma, colon cancer, breast cancer, ovarian cancer, endometrial cancer, esophageal cancer, prostate cancer, cervical cancer, renal cancer, bladder cancer, gastric cancer, non-small cell lung cancer, melanoma, and pancreatic cancer.

32. A composition described in any one of claims 23 to 31, characterized in that it is administered parenterally to the individual.

33. The composition according to any one of claims 23 to 32, characterized in that it is administered in combination with a second treatment.

34. 34. The composition of claim 33, wherein the second treatment is selected from the group consisting of surgery, radiation, gene therapy, immunotherapy, bone marrow transplant, stem cell transplant, hormone therapy, targeted therapy, cryotherapy, ultrasound therapy, photodynamic therapy, and chemotherapy.

35. The composition of claim 34, wherein the second treatment is an immunotherapy.

36. 36. The composition of claim 35, wherein the immunotherapy comprises administering an immunomodulatory agent.

37. 37. The composition of claim 36, wherein the immunomodulatory agent is an immune checkpoint inhibitor.

38. 38. The composition of claim 37, wherein the immune checkpoint inhibitor comprises an anti-PD-L1 antibody or an anti-PD-1 antibody.

39. The composition of any one of claims 23 to 38, wherein the individual is a human.

Images