Anti-idiotypic antibody against anti-KLK2 antibody

Abstract

In certain aspects, the present disclosure relates to anti-idiotype antibodies and antigen-binding portions thereof, e.g., antibodies or antigen-binding portions thereof, that specifically bind to KL2B413-containing proteins. In some aspects, the anti-idiotype antibodies and antigen-binding portions thereof of the present disclosure can be used in methods for detecting and quantifying cells that express a chimeric antigen receptor comprising KL2B413.

Description

[Technical Field]

[0001] (CROSS-REFERENCE TO RELATED APPLICATIONS) This application claims priority to U.S. Provisional Patent Application No. 63 / 053,275, filed July 17, 2020. The entire contents of the foregoing application are incorporated herein by reference in their entirety.

[0002] (Sequence Listing) This application contains a Sequence Listing that has been submitted electronically in ASCII format, which is incorporated herein by reference in its entirety. The ASCII copy was created on July 7, 2021, is named JBI6351WOPCT1_SL.txt, and is 36,187 bytes in size.

[0003] FIELD OF THE INVENTION The present invention relates to anti-idiotypic antibodies and antigen-binding portions thereof, e.g., antibodies or antigen-binding portions thereof, that specifically bind to KL2B413-containing proteins. Methods for detecting and quantifying cells expressing a chimeric antigen receptor containing KL2B413 are also provided. [Background technology]

[0004] Recent advances in understanding the delivery and integration of genomic material into targeted genomes hold great potential for transforming standard of care for a variety of diseases. T cell therapy utilizes isolated T cells that have been genetically modified to enhance their specificity for specific tumor-associated antigens. Genetic modification can involve the expression of chimeric antigen receptors (CARs) or exogenous T cell receptors, providing new antigen specificity on T cells. T cells expressing chimeric antigen receptors (CAR-T cells) can induce tumor immune reactivity.

[0005] One particular CAR target of interest is kallikrein-related peptidase 2 (hK2, HK2), a trypsin-like enzyme specifically expressed in prostate tissue and prostate cancer, driven by the androgen receptor (AR). hK2 is activated by the transmembrane protease, serine 2 (TMPRSS2), and secreted into the prostate ducts, where it cleaves semenogelin, an extracellular matrix component in ejaculate, initiating a cascade that enhances sperm motility. While hK2 expression is restricted to prostate and prostate cancer tissue, it has recently been demonstrated that hK2 is detectable in breast cancer lines and primary patient samples after appropriate activation of the AR pathway by steroid hormones (U.S. Patent Application Publication No. 2018 / 0326102). When the highly structured tissue of the prostate is compromised during hypertrophy or malignant transformation, retrograde release of catalytically inactive hK2 into the blood occurs. Summary of the Invention [Problem to be solved by the invention]

[0006] Therefore, there is a need for CAR-T cell therapies to treat prostate cancer. There is also a need for anti-idiotypic antibodies directed against such CARs to detect, purify, or select proteins and cells that express CARs. [Means for solving the problem]

[0007] The present disclosure provides anti-idiotype antibodies and antigen-binding portions thereof, e.g., antibodies or antigen-binding portions thereof, that specifically bind to KL2B413-containing proteins. The present disclosure also provides nucleic acids encoding the anti-idiotype antibodies and antigen-binding portions thereof, methods for producing the anti-idiotype antibodies and antigen-binding portions thereof, methods for detecting KL2B413 (or cells expressing KL2B413) using the anti-idiotype antibodies and antigen-binding portions thereof, and kits containing the anti-idiotype antibodies and antigen-binding portions thereof.

[0008] In one aspect, the disclosure provides an anti-idiotypic antibody, or antigen-binding portion thereof, that specifically binds to an anti-hK2 targeting antibody, such as a targeting antibody comprising KL2B413. In some embodiments, the targeting antibody or antigen-binding portion comprises a VH domain having an amino acid sequence comprising SEQ ID NO:41 and a VL domain having an amino acid sequence comprising SEQ ID NO:42.

[0009] In other embodiments, the anti-idiotype antibody or antigen-binding portion is for use in detecting KL2B413 in a biological sample, wherein the detection comprises (a) providing a biological sample, (b) contacting the biological sample with the anti-idiotype antibody or antigen-binding portion, and (c) detecting the anti-idiotype antibody or antigen-binding portion.

[0010] In another aspect, the present disclosure provides an anti-idiotype antibody or antigen-binding portion that specifically binds to KL2B413, comprising a heavy chain variable (VH) domain including a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 to 7, a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11 to 14, and a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 19 to 20, and further comprising a light chain variable (VL) domain including a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 25 to 26, a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 29 to 30, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 33 to 34.

[0011] In some embodiments, the anti-idiotype antibody, or antigen-binding portion thereof, comprises a VH domain having an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45, and a VL domain having an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 46. In some embodiments, the anti-idiotype antibody, or antigen-binding portion thereof, comprises a heavy chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37, and further comprises a light chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 39. In some embodiments, the anti-idiotype antibody, or antigen-binding portion thereof, comprises a VH domain having an amino acid sequence having at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO: 45. In some embodiments, the anti-idiotype antibody or antigen-binding portion thereof comprises a VL domain having an amino acid sequence having at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO:46.

[0012] In some embodiments, the anti-idiotype antibody, or antigen-binding portion thereof, comprises a VH domain having the amino acid sequence of SEQ ID NO: 45 and a VL domain having the amino acid sequence of SEQ ID NO: 46. In some other embodiments, the anti-idiotype antibody, or antigen-binding portion thereof, comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 37, and further comprises a light chain comprising the amino acid sequence of SEQ ID NO: 39.

[0013] In some embodiments, the antigen-binding portion is selected from a Fab, F(ab')2, or scFv. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the chimeric antibody comprises a mouse IgG2a framework. In some other embodiments, the antibody is fully human. In some embodiments, the anti-idiotype antibody or antigen-binding portion thereof is specific for KL2B413, where KL2B413 is within the antigen-binding domain of the extracellular portion of a chimeric antigen receptor (CAR). In some embodiments, KL2B413 is an scFv, and the anti-idiotype antibody or antigen-binding portion specifically binds to an epitope in the scFv of the CAR. In some embodiments, KL2B413 specifically binds to KLK2. In some embodiments, the antibody or antigen-binding portion does not cross-react with other KLK2 antibodies or other KLK2-binding CARs. In some embodiments, the CAR has an amino acid sequence selected from the group consisting of SEQ ID NOs: 43-44.

[0014] In some embodiments, the present disclosure provides nucleic acids encoding the heavy chain, the light chain, or both, of an anti-idiotype antibody or antigen-binding portion.

[0015] In another aspect, the present disclosure provides a nucleic acid encoding the heavy chain, the light chain, or both of an anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to KL2B413, the nucleic acid comprising the nucleotide sequence of SEQ ID NO: 38, the nucleotide sequence of SEQ ID NO: 40, or both. In another aspect, the present disclosure provides a vector comprising the nucleic acid sequence. In some embodiments, the vector is an expression vector. In another aspect, the present disclosure provides a host cell comprising the vector. In some embodiments, the host cell is a mammalian cell.

[0016] In another aspect, the disclosure provides a method for producing an anti-idiotype antibody or antigen-binding portion thereof that specifically binds to KL2B413, the method comprising culturing a host cell comprising nucleotide sequences encoding the heavy and light chains of the antibody or antigen-binding portion under conditions allowing expression of the antibody or antigen-binding portion, and isolating the antibody or antigen-binding portion from the culture. In some embodiments, the host cell encodes a vector comprising a nucleic acid encoding the anti-idiotype antibody or antigen-binding portion thereof.

[0017] In another aspect, the present disclosure provides a method for detecting KL2B413 in a biological sample, the method comprising: (a) preparing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion; and (c) detecting the anti-idiotype antibody or antigen-binding portion.

[0018] In another aspect, the present disclosure provides a method for detecting expression of a chimeric antigen receptor (CAR) comprising KL2B413 in a biological sample, the method comprising: (a) providing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion; and (c) detecting the anti-idiotype antibody or antigen-binding portion, thereby detecting expression of the CAR.

[0019] In some embodiments, the antibody comprises a detectable label. In some embodiments, the method further comprises contacting the anti-idiotype antibody or antigen-binding portion with a detectable label prior to detecting the anti-idiotype antibody or antigen-binding portion. In some embodiments, the biological sample is blood, serum, or urine.

[0020] In some aspects, the present disclosure provides a kit for detecting KL2B413 in a biological sample, the kit comprising: (a) an anti-idiotype antibody or antigen-binding portion; and (b) instructions for detecting the anti-idiotype antibody or antigen-binding portion.

[0021] In another aspect, the present disclosure provides a method for purifying KL2B413 from a sample, the method comprising: (a) preparing a biological sample containing KL2B413; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion of the present disclosure; and (c) capturing the anti-idiotype antibody or antigen-binding portion, thereby purifying KL2B413.

[0022] In another aspect, the disclosure provides a method of selecting CAR-T cells from a cell population, the method comprising: (a) providing a biological sample containing CAR-T cells; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion thereof; and (c) capturing the anti-idiotype antibody or antigen-binding portion thereof, thereby selecting the CAR-T cells. In some embodiments, the anti-idiotype antibody or antigen-binding portion thereof is specific for KL2B413.

[0023] The present disclosure contemplates all combinations of any of the foregoing aspects and embodiments, as well as combinations with any of the embodiments described in the detailed description and examples. [Brief explanation of the drawings]

[0024] For the purpose of illustrating the invention, there are shown in the drawings certain embodiments of the present disclosure. However, the present disclosure is not limited to the precise arrangements and instrumentalities of the embodiments shown in the drawings. [Figure 1] 1 is a graphical representation of KL2B513 or KL2B610 specific binding enrichment after four rounds of panning as detected by polyclonal ELISA. [Figure 2] 1 is a graphical representation of monoclonal Fab binding screening results from KL2B513 or KL2B610 target binding assays compared to GCDB734 counterscreening reagent binding assays. [Figure 3] 1 is a graphical representation of a cell-based binding assay in which monoclonal antibodies were screened for binding to scFv-transfected SupT1 cells. [Figure 4] 1 is a graphical representation of the dose-dependent binding of A002B39 to KL2B413-LH SupT1 cells. [Figure 5] 1 is a graphic representation of the dose-dependent blocking of PE-A002B39 binding to KL2B413-LH SupT1 cells by KL2B413-LH scFv-Fc fusion protein. [Figure 6] 1 is a graphical representation of PE-A002B39 detection of KL2B413-LH-SupT1 cells spiked into whole blood. [Figure 7] 1 is a graphical representation of two different lots of PE-A002B39 that have similar binding profiles to KL2B413-LH-SupT1. DETAILED DESCRIPTION OF THE INVENTION

[0025] Overview The present disclosure provides anti-idiotypic antibodies and antigen-binding portions thereof, e.g., antibodies or antigen-binding portions thereof, that specifically bind to KL2B413-containing proteins. The anti-idiotypic antibodies and antigen-binding portions of the present disclosure can be used in methods for detecting and quantifying cells expressing a CAR containing KL2B413. Such methods may allow researchers to determine whether a given batch of in vitro-generated CAR-T cells expresses a desired CAR and, therefore, whether the cells are therapeutically useful for targeting a desired protein. In the present disclosure, the anti-idiotypic antibodies and antigen-binding portions target KL2B413, which itself targets KLK2, a protein associated with prostate tissue.

[0026] definition As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the content clearly dictates otherwise. Thus, for example, reference to "a cell" includes a combination of two or more cells, and the like.

[0027] The transitional phrases "comprising," "consisting essentially of," and "consisting of" are intended to connote their generally accepted meanings in patent language, i.e., (i) "comprising" is synonymous with "comprising," "containing," or "characterized by" and is inclusive or open-ended and does not exclude other unrecited elements or method steps; (ii) "consisting of" excludes any element, step, or ingredient not specified in the claim; and (iii) "consisting essentially of" limits the claim to the specified materials or steps and those that do not materially affect the basic and novel characteristics" of the claimed invention. Embodiments described with the phrase "comprising" (or its equivalents) are also provided as embodiments described independently with "consisting of" and "consisting essentially of."

[0028] "Activation" or "stimulation" or "activated" or "stimulated" refers to the induction of a change in the biological state of a cell that results in the expression of activation markers, cytokine production, or mediation of target cell proliferation or cytotoxicity. Cells can be activated by a primary stimulatory signal. A costimulatory signal can amplify the magnitude of the primary signal and suppress cell death after initial stimulation, resulting in a more durable activation state and, therefore, greater cytotoxic potential. A "costimulatory signal" refers to a signal that, in combination with a primary signal, such as TCR / CD3 ligation, causes T cell and / or NK cell proliferation and / or up- or down-regulation of key molecules.

[0029] An "anti-idiotype antibody" or "anti-idiotypic antibody" refers to an antibody that specifically binds to the variable region of another antibody. In the case of KLK2, an anti-idiotype antibody specifically binds to an anti-KLK2 antibody.

[0030] An "antigen-binding portion," "antigen-binding fragment," or "antigen-binding domain" refers to a portion of a protein that binds to an antigen. An antigen-binding portion may be a synthetic, enzymatically obtainable, or genetically engineered polypeptide, and includes antigen-binding portions of immunoglobulins, such as VH, VL, VH and VL, Fab, Fab', F(ab')2, Fd, and Fv fragments, domain antibodies (dAbs) consisting of one VH domain or one VL domain, shark variable IgNAR domains, camelized VH domains, VHH domains, minimal recognition units consisting of amino acid residues mimicking the CDRs of an antibody, such as the FR3-CDR3-FR4 portion, HCDR1, HCDR2, and / or HCDR3, and LCDR1, LCDR2, and / or LCDR3, alternative scaffolds that bind to antigen, and multispecific proteins comprising an antigen-binding portion. Antigen-binding moieties (such as VH and VL) can be linked to each other via synthetic linkers to form a variety of single-chain antibody designs in which, when the VH and VL domains are expressed as separate single chains, the VH / VL domains can pair intramolecularly or intermolecularly to form monovalent antigen-binding domains, e.g., single-chain Fvs (scFvs) or diabodies. Antigen-binding moieties may also be conjugated to other antibodies, proteins, antigen-binding moieties, or alternative scaffolds, which may be monospecific or multispecific, to engineer bispecific and multispecific proteins.

[0031] "Cancer" refers to a broad group of diverse diseases characterized by the uncontrolled growth of abnormal cells in the body. Uncontrolled cell division and growth leads to the formation of malignant tumors that invade adjacent tissues and may metastasize to distant parts of the body via the lymphatic system or bloodstream. "Cancer" or "cancerous tissue" can include tumors.

[0032] A "full-length antibody" is composed of two heavy chains (HC) and two light chains (LC) interconnected by disulfide bonds, and multimers thereof (e.g., IgM). Each heavy chain is composed of a heavy chain variable domain (VH) and a heavy chain constant domain, which in turn is composed of the subdomains CH1, hinge, CH2, and CH3. Each light chain is composed of a light chain variable domain (VL) and a light chain constant domain (CL). VH and VL can be further subdivided into regions of hypervariability called complementarity determining regions (CDRs), which are interspersed with framework regions (FRs). Each VH and VL is composed of three CDR and four FR segments, arranged from the amino terminus to the carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

[0033] "Complementarity-determining region (CDR)" is the antigen-binding site in an antibody. CDRs can be defined using various terms: (i) The three complementarity-determining regions (CDRs) in VH (HCDR1, HCDR2, HCDR3) and the three in VL (LCDR1, LCDR2, LCDR3) are based on sequence diversity (Wu and Kabat, J Exp Med 132:211-50, 1970; Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991). (ii) "Hypervariable regions," "HVRs," or "HVs," three in VH (H1, H2, H3) and three in VL (L1, L2, L3), refer to regions of antibody variable domains that are hypervariable in structure as defined by Chothia and Lesk (Chothia and Lesk Mol Biol 196:901-17, 1987). The International ImMunoGeneTics (IMGT) database (http: / / www_imgt_org) provides standard numbering and definitions for antigen-binding sites. The correspondence between CDRs, HVs, and IMGT profiles is described in Lefranc et al., Dev Comparat Immunol 27:55-77, 2003. As used herein, the terms "CDR," "HCDR1," "HCDR2," "HCDR3," "LCDR1," "LCDR2," and "LCDR3," unless otherwise specified herein, include CDRs defined by any of the methods described by Kabat, Chothia, or IMGT, supra.

[0034] A "human antibody" refers to an antibody optimized to evoke a minimal immune response when administered to a human subject. The variable regions of a human antibody are derived from human immunoglobulin sequences. If a human antibody contains a constant region or a portion of a constant region, the constant region is also derived from human immunoglobulin sequences. A human antibody contains heavy and light chain variable regions "derived" from sequences of human origin when the variable regions of the human antibody are obtained from a system using human germline immunoglobulins or rearranged immunoglobulin genes. Exemplary such systems include phage-displayed human immunoglobulin gene libraries and transgenic non-human animals, such as mice or rats, carrying human immunoglobulin loci. A "human antibody" typically contains amino acid differences compared to immunoglobulins expressed in humans due to differences in the systems used to obtain human antibodies and human immunoglobulin loci, the introduction of somatic mutations or intentional substitutions into frameworks or CDRs, or both. Typically, a "human antibody" is at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical in amino acid sequence to the amino acid sequence encoded by human germline immunoglobulin or rearranged immunoglobulin genes. Optionally, a "human antibody" may contain consensus framework sequences obtained from human framework sequence analysis, e.g., as described in Knappik et al., (2000) J Mol Biol 296:57-86, or synthetic HCDR3s incorporated into a phage-displayed human immunoglobulin gene library, e.g., as described in Shi et al., (2010) J Mol Biol 397:385-96 and WO 2009 / 085462. Antibodies in which at least one CDR is derived from a non-human species are not included in the definition of "human antibody."

[0035] "Humanized antibody" refers to an antibody in which at least one CDR is derived from a non-human species and at least one framework is derived from a human immunoglobulin sequence. Humanized antibodies can contain substitutions in the framework, so that the framework may not be an exact copy of an expressed human immunoglobulin or human immunoglobulin germline gene sequence.

[0036] "Isolated" refers to a homogenous population of molecules (e.g., synthetic polynucleotides or polypeptides) that have been substantially separated and / or purified from other components of the system in which the molecule is produced, such as in a recombinant cell, and proteins that have been subjected to at least one purification or isolation step. "Isolated" refers to molecules that are substantially free of other cellular material and / or chemicals and includes molecules isolated to greater degrees of purity, e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% purity.

[0037] "Modulate" refers to either an enhanced or decreased ability of the test molecule to mediate a greater or lesser response (i.e., a downstream effect) when compared to a control or vehicle-mediated response.

[0038] "Natural killer cells" and "NK cells" are used interchangeably and synonymously herein. NK cells are cells that express CD16 + CD56 + and / or CD57 + TCR - NK cells are differentiated lymphocytes with a phenotype characterized by the ability to bind to and kill cells that fail to express "self" MHC / HLA antigens through the activation of specific cytolytic enzymes, to kill tumor cells or other diseased cells that express ligands for NK-activating receptors, and to release protein molecules called cytokines that stimulate or inhibit the immune response.

[0039] "Specifically binds," "specific binding," "specifically binding," or "binds" refers to a proteinaceous molecule binding to an antigen or an epitope within an antigen with higher affinity than its affinity for other antigens. Typically, a proteinaceous molecule binds to an antigen with an affinity of about 1 x 10 -7 M or less, e.g., about 5 × 10 -8 M or less, approximately 1×10 -8 Approximately 1×10 below M -9 M or less, approximately 1×10 -10 M or less, approximately 1×10 -11 M or less, or about 1 x 10 -12 M, 1 x 10 -13 M, 1 x 10 -14 M, 1 x 10 -15 The equilibrium dissociation constant (K D ) binds to an antigen or an epitope within an antigen, typically D is the K for binding to nonspecific antigens (e.g., BSA, casein) D In the context of the prostate neoantigens described herein, "specific binding" refers to the binding of a proteinaceous molecule to a prostate neoantigen without detectably binding to the wild-type protein of which the prostate neoantigen is a mutant.

[0040] "Tumor cells" or "cancer cells" refer to cancerous, precancerous, or transformed cells that have spontaneously or induced phenotypic changes, either in vivo, ex vivo, or in tissue culture. These changes do not necessarily involve the incorporation of new genetic material. Transformation can occur through infection with a transforming virus and the incorporation of new genomic nucleic acid, through the incorporation of exogenous nucleic acid, or through spontaneous or subsequent exposure to carcinogens, which result in mutations in endogenous genes. Transformation / cancer is exemplified by in vitro, in vivo, and ex vivo morphological changes, cellular immortalization, aberrant growth control, formation of foci, proliferation, malignant lesions, modulation of tumor-specific marker levels, invasiveness, and tumor growth in suitable animal hosts, such as nude mice.

[0041] The term "chimeric antigen receptor" or "CAR," as used herein, is defined as a cell surface receptor comprising an extracellular target binding domain, a transmembrane domain, and an intracellular signaling domain, all in a combination not naturally found together in a single protein. This particularly includes receptors in which the extracellular domain and intracellular signaling domain are not naturally found together in a single receptor protein. The chimeric antigen receptors of the present invention are intended primarily for use in lymphocytes, such as T cells and natural killer (NK) cells.

[0042] The terms "T cells" and "T lymphocytes" are interchangeable and are used synonymously herein. As used herein, T cells include thymocytes, naive T lymphocytes, immature T lymphocytes, mature T lymphocytes, resting T lymphocytes, or activated T lymphocytes. T cells can be T helper (Th) cells, e.g., T helper 1 (Th1) or T helper 2 (Th2) cells. T cells can be helper T cells (HTLs, CD4+ T cells), CD4+ T cells, cytotoxic T cells (CTLs, CD8+ T cells), tumor-infiltrating cytotoxic T cells (TILs, CD8+ T cells), CD4+CD8+ T cells, or any other subset of T cells. Other exemplary populations of T cells suitable for use in certain embodiments include naive T cells and memory T cells. Also included are "NKT cells," which refer to a special population of T cells that express not only the semi-invariant αβ T cell receptor but also various molecular markers typically associated with NK cells, such as NK1.1. NKT cells include NK1.1+ and NK1.1-, as well as CD4+, CD4-, CD8+, and CD8 cells. The TCR of NKT cells is unique in that it recognizes glycolipid antigens presented by the MHC I-like molecule CD Id. NKT cells can have either protective or harmful effects due to their ability to produce cytokines that promote either inflammation or immune tolerance. Also included are "gamma delta T cells (γδ T cells)," which refer to a special population of a small subset of T cells that possess a distinct TCR on their surface, distinct from the majority of T cells, whose TCR is composed of two glycoprotein chains, designated α and β-TCR chains. The TCR in γδ T cells is composed of a γ chain and a δ chain. γδ T cells can play a role in immune surveillance and immune regulation, and have been found to be an important source of IL-17 and to induce vigorous CD8+ cytotoxic T cell responses. Also included are "regulatory T cells" or "Tregs," which refer to T cells that suppress aberrant or excessive immune responses and play a role in immune tolerance. Tregs are typically Foxp3-positive CD4+ T cells, but can also include Foxp3-negative regulatory T cells, which are IL-10-producing CD4+ T cells.

[0043] As used herein, the term "antigen" refers to any molecule (e.g., a protein, peptide, polysaccharide, glycoprotein, glycolipid, nucleic acid, portion thereof, or combination thereof) capable of being bound by a T cell receptor. An antigen can also elicit an immune response. Examples of immune responses may involve, but are not limited to, antibody production, or activation of specific immunologically competent cells, or both. Those skilled in the art will understand that an antigen need not necessarily be encoded by a "gene." It is readily apparent that antigens may be synthetically produced or derived from biological samples, or may be macromolecules other than polypeptides. Such biological samples may include, but are not limited to, tissue samples, tumor samples, cells, or other biological components, organisms, protein / antigen subunits, killed or inactivated whole cells, or fluids containing lysates.

[0044] The term "antibody" refers to monoclonal antibodies, multispecific antibodies, human antibodies, humanized antibodies, chimeric antibodies, single-chain Fvs (scFvs), single-chain antibodies, Fab fragments, F(ab') fragments, disulfide-linked Fvs (sdFvs), intrabodies, minibodies, diabodies, and anti-idiotypic (anti-Id) antibodies (including, for example, anti-Id antibodies against antigen-specific TCRs), and epitope-binding portions of any of the above. The terms "antibody" and "antibody" also refer to covalent diabodies, such as those described in U.S. Patent Application Publication No. 2007 / 0004909, and Ig-DARTS, such as those disclosed in U.S. Patent Application Publication No. 2009 / 0060910. Antibodies useful as TCR-binding molecules include immunoglobulin molecules and immunologically active fragments of immunoglobulin molecules, i.e., molecules that contain an antigen-binding site. Immunoglobulin molecules can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgM1, IgM2, IgA1, and IgA2), or subclass.

[0045] The term "host cell" refers to any cell containing heterologous nucleic acid. The heterologous nucleic acid can be a vector (e.g., an expression vector). For example, a host cell can be a cell from any organism that is selected, modified, transformed, grown, used, or manipulated in any way for the cellular production of a substance, e.g., the cellular expression of a gene, DNA or RNA sequence, protein, or enzyme. An appropriate host can be determined. For example, a host cell can be selected based on the vector backbone and the desired result. By way of example, a plasmid or cosmid can be introduced into a prokaryotic host cell to replicate some types of vectors. Bacterial cells, such as, but not limited to, DH5α, JM109, and KCB, SURE® competent cells, and SOLOPACK Gold cells can be used as host cells for vector replication and / or expression. Additionally, bacterial cells, such as E. coli LE392, can be used as host cells for phage viruses. Eukaryotic cells that can be used as host cells include, but are not limited to, yeast (e.g., YPH499, YPH500, and YPH501), insect, and mammalian cells. Examples of mammalian eukaryotic host cells for replication and / or expression of vectors include, but are not limited to, HeLa, NIH3T3, Jurkat, 293, COS, CHO, Saos, and PC12.

[0046] The terms "express" and "expression" mean to allow or cause the information of a gene or DNA sequence to be produced, e.g., to produce a protein by activating cellular functions involved in transcription and translation of the corresponding gene or DNA sequence. A DNA sequence is expressed in or by a cell to form an "expression product," such as a protein. The expression product itself, e.g., the resulting protein, can also be said to be "expressed" by the cell. Expression products can be characterized as intracellular, extracellular, or transmembrane.

[0047] The term "transfection" refers to the introduction of "foreign" (i.e., exogenous or extracellular) nucleic acid into a cell using recombinant DNA technology. The term "genetic modification" refers to the introduction of a "foreign" (i.e., exogenous or extracellular) gene, DNA, or RNA sequence into a host cell such that the host cell expresses the introduced gene or sequence to produce a desired substance, usually a protein or enzyme encoded by the introduced gene or sequence. The introduced gene or sequence may also be referred to as a "cloned" or "foreign" gene or sequence and may include regulatory or control sequences operably linked to the polynucleotide encoding the chimeric antigen receptor, such as start, stop, promoter, signal, secretion, or other sequences used by the cell's genetic machinery. The gene or sequence may also include nonfunctional sequences with no known function. A host cell that receives and expresses introduced DNA or RNA is "genetically engineered." The DNA or RNA introduced into a host cell can be from any source, including cells of the same genus or species as the host cell, or from a different genus or species.

[0048] The term "transduction" refers to the introduction of foreign nucleic acid into cells using a viral vector.

[0049] The term "control element" refers to any cis-acting genetic element that controls some aspect of the expression of a nucleic acid sequence. In some embodiments, the term "promoter" essentially includes the minimal sequences necessary to initiate transcription. In some embodiments, the term "promoter" includes sequences for initiating transcription and, in addition, also includes sequences that can upregulate or downregulate transcription, commonly referred to as "enhancer elements" and "repressor elements," respectively.

[0050] As used herein, "operably linked" and similar phrases, when used with reference to nucleic acids or amino acids, refer to the operably linking of nucleic acid or amino acid sequences, respectively, that are placed in a functional relationship with each other. For example, operably linked promoters, enhancer elements, open reading frames, 5' and 3' UTRs, and terminator sequences result in the correct production of a nucleic acid molecule (e.g., RNA). In some embodiments, operably linked nucleic acid elements result in the transcription of the open reading frame and ultimately the production of a polypeptide (i.e., expression of the open reading frame). As another example, an operably linked peptide refers to one in which functional domains are positioned at an appropriate distance from each other to confer the intended function of each domain.

[0051] "Enhance" or "promote" or "increase" or "expand" or "improve" generally refers to the ability of the compositions contemplated herein to produce, induce, or cause a greater physiological response (i.e., a downstream effect) compared to the response caused by either a vehicle or a control molecule / composition. Measurable physiological responses may include, among other things, T cell expansion, activation, effector function, increased persistence, and / or increased cancer cell killing capacity, as will be apparent from an understanding of the art and the present disclosure. In certain embodiments, an "increased" or "enhanced" amount may be a "statistically significant" amount and may include an increase of 1.1, 1.2, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, or more (e.g., 500, 1000 fold) (including all integers and decimal points therebetween, and greater than 1, e.g., 1.5, 1.6, 1.7, 1.8, etc.) over the response produced by a vehicle or control composition.

[0052] "Decrease" or "lower" or "lesser" or "reduce" or "attenuate" generally refers to the ability of a composition contemplated herein to produce, induce, or cause a physiological response (i.e., a downstream effect) that is less than the response caused by either a vehicle or a control molecule / composition. In certain embodiments, a "decreased" or "reduced" amount may be a "statistically significant" amount and may include a 1.1, 1.2, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, or more (e.g., 500, 1000-fold) decrease (including all integers and decimal points therebetween, and greater than 1, e.g., 1.5, 1.6, 1.7, 1.8, etc.) of the response caused by the vehicle, a control composition (reference response), or a response in a particular cell lineage.

[0053] The term "effective" as applied to a dose or amount refers to that amount of a compound or pharmaceutical composition sufficient to produce the desired activity upon administration to a subject in need thereof. It should be noted that when administering a combination of active ingredients, the effective amount of the combination may or may not include the amount of each ingredient that would be effective when administered individually. The exact amount required will vary from subject to subject, depending on the species, age, and general condition, the severity of the condition being treated, the particular drug used, the mode of administration, and the like.

[0054] The phrase "pharmaceutically acceptable" as used in connection with the compositions described herein refers to molecular entities and other components of such compositions that are physiologically tolerable and do not normally produce adverse reactions when administered to mammals (e.g., humans). Preferably, the term "pharmaceutically acceptable" means approved by a federal or state regulatory agency or listed in the United States Pharmacopoeia or other generally recognized pharmacopeia for use in mammals, more specifically humans.

[0055] The term "protein," as used herein, encompasses all types of modified proteins, including, but not limited to, natural and synthetic proteins, including protein fragments of all lengths, fusion proteins, and glycoproteins, as well as all other types of modified proteins (e.g., proteins resulting from phosphorylation, acetylation, myristoylation, palmitoylation, glycosylation, oxidation, formylation, amidation, polyglutamylation, ADP-ribosylation, pegylation, biotinylation, etc.).

[0056] The terms "nucleic acid," "nucleotide," and "polynucleotide" encompass both DNA and RNA unless otherwise specified. "Nucleic acid sequence" or "nucleotide sequence" refers to a nucleic acid sequence that encodes an amino acid; these terms may also refer to a nucleic acid sequence that includes portions encoding any amino acids added as a cloning artifact, including any amino acids encoded by linkers.

[0057] The term "carrier" refers to a diluent, adjuvant, excipient, or vehicle with which a compound is administered. Such pharmaceutical carriers may be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, etc. Water or aqueous solutions, saline, and aqueous dextrose and glycerin solutions are preferably used as carriers, particularly for injectable solutions. Alternatively, the carrier may be a solid dosage form carrier, including, but not limited to, one or more of a binder (for compressed pills), a glidant, an encapsulating agent, a flavoring agent, and a coloring agent. Examples of suitable pharmaceutical carriers are described in "Remington's Pharmaceutical Sciences" by EW Martin.

[0058] The term "about" or "approximately" includes within a statistically meaningful range of values. Such a range can be within one order of magnitude, preferably within 50%, more preferably within 20%, even more preferably within 10%, and even more preferably within 5% of a given value or range. The allowable variation encompassed by the term "about" or "approximately" will depend on the specific system under study and can be readily understood by one of ordinary skill in the art.

[0059] "Kallikrein-related peptidase 2," "hK2," "KLK2," or "klk2" refers to a known protein also known as kallikrein-2, granular kallikrein 2, or HK2. hK2 is produced as a preproprotein, which is cleaved during proteolysis to generate the active protease. All hK2 isoforms and variants are encompassed by "hK2." The amino acid sequences of various isoforms can be found under GenBank accession numbers NP_005542.1, NP_001002231.1, and NP_001243009. The amino acid sequence of full-length human hK2 is shown in SEQ ID NO: 47. The sequence includes the signal peptide (residues 1-18) and propeptide region (residues 19-24).

[0060] The term "KL2B413" refers to any antibody, antigen-binding portion thereof, or any other protein containing variable regions derived from KL2B413 VH (SEQ ID NO: 41) and KL2B413 VL (SEQ ID NO: 42), including a CAR. In certain embodiments, an anti-idiotype antibody of the present disclosure specifically binds to a protein comprising the VH domain set forth in SEQ ID NO: 41 and / or the VL domain set forth in SEQ ID NO: 42. In certain embodiments, an anti-idiotype antibody of the present disclosure specifically binds to a protein comprising the three CDRs of the VH domain set forth in SEQ ID NO: 41 and the three CDRs of the VL domain set forth in SEQ ID NO: 42.

[0061] The term "KL2B513" refers to an scFv fusion protein derived from KL2B413 having variable regions in a VL-VH orientation. KL2B513 may be referred to interchangeably as KL2B413-LH-scFv.

[0062] The term "KL2B610" refers to an scFv fusion protein derived from KL2B413 with the variable regions in a VH-VL orientation. KL2B610 may be interchangeably referred to as KL2B413-HL-scFv.

[0063] The term "A002B39" refers to a chimeric mAb with human VH / VL and mouse IgG2a / k that targets KL2B413. A002B39 may be referred to interchangeably as A002M39.

[0064] The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting. As used herein, the articles "a," "an," and "the" are to be understood as including plural referents unless the context clearly indicates otherwise.

[0065] The present disclosure further provides variants, e.g., functional variants, of the antibodies, nucleic acids, polypeptides, and proteins described herein. A "variant" refers to a polypeptide or polynucleotide that differs from a reference polypeptide or reference polynucleotide by one or more modifications, e.g., substitutions, insertions, or deletions. As used herein, the term "functional variant" refers to an antibody, polypeptide, or protein that has substantial or significant sequence identity or similarity with a parent antibody, polypeptide, or protein, and the functional variant retains the biological activity of the antibody, polypeptide, or protein from which it is variant. Functional variants include, for example, variants of an antibody, polypeptide, or protein described herein (parent antibody, polypeptide, or protein) that retain the ability to recognize target cells to a similar, equal, or greater extent than the parent antibody, polypeptide, or protein. With respect to a parent antibody, polypeptide, or protein, a functional variant can be, for example, at least about 30%, about 40%, about 50%, about 60%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or more identical in amino acid sequence to the parent antibody, polypeptide, or protein.

[0066] As used herein, the structure of a polypeptide is defined based on its percent sequence identity with a listed reference sequence (with a given SEQ ID NO). In this context, the percent sequence identity between two amino acid sequences can be determined by comparing the two sequences aligned in an optimal manner, and the compared amino acid sequence may contain additions or deletions relative to the reference sequence for optimal alignment between the two sequences. The percent identity is calculated by determining the number of identical positions where the amino acid residues between the two sequences are identical, dividing this number by the total number of positions within the comparison window, and multiplying the result by 100 to obtain the percent identity between the two sequences. Typically, the comparison window corresponds to the entire length of the sequences being compared. For example, the BLAST program, "BLAST 2 sequences" (Tatusova et al., "Blast 2 sequences—a new tool for comparing protein and nucleotide sequences," FEMS Microbiol Lett. 174:247-250), available at http: / / www.ncbi.nlm.nih.gov / gorf / bl2.html, can be used, using the default parameters (specifically, the parameters "open gap penalty": 5 and "extension gap penalty": 2; the selected matrix is, for example, the matrix "BLOSUM 62" proposed by the program), and the percentage of identity between the two sequences being compared is calculated directly by the program. Determining the sequence identity of a query sequence to a reference sequence is within the capabilities of those skilled in the art and can be performed using commercially available analysis software such as BLAST (trademark).

[0067] A functional variant may, for example, comprise the amino acid sequence of a parent antibody, polypeptide, or protein with at least one conservative amino acid substitution. In another embodiment, a functional variant may comprise the amino acid sequence of a parent antibody, polypeptide, or protein with at least one non-conservative amino acid substitution. In this case, the non-conservative amino acid substitution may not suppress or inhibit the biological activity of the functional variant. The non-conservative amino acid substitution may improve the biological activity of the functional variant, such that the biological activity of the functional variant is enhanced compared to the parent antibody, polypeptide, or protein.

[0068] The amino acid substitutions in the antibodies of the present invention may be conservative amino acid substitutions. Conservative amino acid substitutions are known in the art and include amino acid substitutions in which one amino acid having particular physical and / or chemical properties is replaced with another amino acid having the same or similar chemical or physical properties. For example, conservative amino acid substitutions may be an acidic amino acid (e.g., Asp or Glu) substituted with another acidic amino acid, an amino acid having a nonpolar side chain substituted with another amino acid having a nonpolar side chain (e.g., Ala, Gly, Val, Ile, Leu, Met, Phe, Pro, Trp, Val, etc.), a basic amino acid (Lys, Arg, etc.) substituted with another basic amino acid, an amino acid having a polar side chain substituted with another amino acid having a polar side chain (Asn, Cys, Gln, Ser, Thr, Tyr, etc.), etc.

[0069] The antibodies, polypeptides, and proteins (including functional portions and functional variants of the invention) of the present embodiments can contain synthetic amino acids in place of one or more naturally occurring amino acids. Such synthetic amino acids are known in the art and include, for example, aminocyclohexanecarboxylic acid, norleucine, α-amino n-decanoic acid, homoserine, S-acetylaminomethyl-cysteine, trans-3- and trans-4-hydroxyproline, 4-aminophenylalanine, 4-nitrophenylalanine, α-(2-amino-2-norbornane)-carboxylic acid, α,γ-diaminobutyric acid, α,β-diaminopropionic acid, homophenylalanine, 4-chlorophenylalanine, 4-carboxyphenylalanine, β-phenylserine, β- Hydroxyphenylalanine, phenylglycine, α-naphthylalanine, cyclohexylalanine, cyclohexylglycine, N'-benzyl-N'-methyl-lysine, N',N'-dibenzyl-lysine, 6-hydroxylysine, ornithine, α-aminocyclopentanecarboxylic acid, α-aminocyclohexanecarboxylic acid, α-aminocycloheptanecarboxylic acid, indoline-2-carboxylic acid, 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid, aminomalonic acid, aminomalonic acid monoamide, and α-tert-butylglycine.

[0070] The antibodies, polypeptides, and proteins (including functional portions and functional variants) of the present embodiments can be post-translationally modified. They can be glycosylated, esterified, N-acylated, amidated, carboxylated, phosphorylated, esterified, cyclized, for example, via disulfide bridges, or converted into acid addition salts. In some embodiments, they are dimerized or polymerized, or conjugated.

[0071] Antibodies, polypeptides, and / or proteins (including functional portions and functional variants thereof) of embodiments of the present invention can be obtained by methods known in the art. Suitable methods for de novo synthesis of polypeptides and proteins are described in references such as Chan et al., Fmoc Solid Phase Peptide Synthesis, Oxford University Press, Oxford, United Kingdom, 2000; Peptide and Protein Drug Analysis, ed. Reid, R., Marcel Dekker, Inc., 2000; and Epitope Mapping, ed. Westwood et al., Oxford University Press, Oxford, United Kingdom, 2001. Polypeptides and proteins can also be produced recombinantly using the nucleic acids described herein using standard recombinant methods. See, for example, Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd ed., Cold Spring Harbor Press, Cold Spring Harbor, NY 2001, and Ausubel et al., Current Protocols in Molecular Biology, Greene Publishing Associates and John Wiley & Sons, NY, 1994. Furthermore, portions of the antibodies, polypeptides, and proteins (including functional portions and functional variants thereof) of the present invention can be isolated and / or purified from sources such as plants, bacteria, insects, mammals, etc. Methods of isolation and purification are known in the art. Alternatively, the antibodies, polypeptides, and / or proteins (including functional portions and functional variants thereof) described herein can be commercially synthesized. In this regard, antibodies, polypeptides, and proteins can be synthesized, recombinant, isolated, and / or purified.

[0072] Methods and Uses of the Disclosure The present disclosure provides anti-idiotype antibodies and antigen-binding portions thereof, e.g., antibodies or antigen-binding portions thereof, that specifically bind to KL2B413-containing proteins. For example, anti-idiotype antibodies can contain an amino acid sequence complementary to a portion of the KL2B413 antibody to promote specific binding. The present disclosure also provides nucleic acids encoding anti-idiotype antibodies and antigen-binding portions thereof, methods for producing anti-idiotype antibodies and antigen-binding portions thereof, methods for detecting KL2B413 using anti-idiotype antibodies and antigen-binding portions thereof, and kits containing anti-idiotype antibodies and antigen-binding portions thereof. For example, anti-idiotype antibodies can be included in kits with other reagents and used to determine whether a given biological sample contains a KL2B413 antibody or a fragment thereof expressed on the surface of T cells, for example, in a CAR.

[0073] Methods for testing antibodies for their ability to bind to any functional portion of KL2B413 are known in the art and include any antibody-antigen binding assay, such as radioimmunoassay (RIA), Western blot, enzyme-linked immunosorbent assay (ELISA), immunoprecipitation, and competitive inhibition assay.

[0074] Suitable methods for producing antibodies are known in the art, e.g., standard hybridoma techniques are described in, e.g., Kohler and Milstein, Eur. J. Immunol., 5, 511-519 (1976), Harlow and Lane (eds.), Antibodies: A Laboratory Manual, CSH Press (1988), and CA Janeway et al. (eds.), Immunobiology, 5th Ed., Garland Publishing, New York, NY (2001). Alternatively, other methods are known in the art, such as the EBV-hybridoma method (Haskard and Archer, J. Immunol. Methods, 74(2), 361-67 (1984), and Roder et al., Methods Enzymol., 121, 140-67 (1986)), and bacteriophage vector expression systems (see, e.g., Huse et al., Science, 246, 1275-81 (1989)). Additionally, methods for producing antibodies in non-human animals are described, for example, in U.S. Pat. Nos. 5,545,806, 5,569,825, and 5,714,352, and U.S. Patent Application Publication No. 2002 / 0197266(A1)).

[0075] Antibodies can also be generated using phage display. In this regard, phage libraries encoding antibody antigen-binding variable (V) domains can be generated using standard molecular biology and recombinant DNA techniques (see, e.g., Sambrook et al., supra, and Ausubel et al., supra). Phage encoding variable regions with the desired specificity are selected for specific binding to the desired antigen, and complete or partial antibodies comprising the selected variable domains are reconstituted. Nucleic acid sequences encoding the reconstituted antibodies are introduced into a suitable cell line, such as myeloma cells used for hybridoma production, so that the cells secrete antibodies with the properties of monoclonal antibodies (see, e.g., Janeway et al., supra, Huse et al., supra, and U.S. Pat. No. 6,265,150).

[0076] In one aspect, the present disclosure provides an anti-idiotype antibody or antigen-binding portion thereof that specifically binds to a targeting antibody or CAR comprising KL2B413. For example, the anti-idiotype antibody or antigen-binding portion may specifically bind to one or more domains of a fragment antigen-binding region (Fab) comprising a VH and a VL. In some embodiments, the anti-idiotype antibody or antigen-binding portion comprises a VH domain having the amino acid sequence of SEQ ID NO: 45 and a VL domain having the amino acid sequence of SEQ ID NO: 46.

[0077] In other embodiments, the anti-idiotype antibody or antigen-binding portion is for use in detecting KL2B413 in a biological sample, the detection comprising (a) providing a biological sample, (b) contacting the biological sample with the anti-idiotype antibody or antigen-binding portion, and (c) detecting the anti-idiotype antibody or antigen-binding portion. For example, the anti-idiotype antibody can be added to any biological sample, including tissue samples, tumor samples, liquids containing cells or other biological components, organisms, protein / antigen subunits, killed or inactivated whole cells, or lysates. The anti-idiotype antibody can be contained in a solution containing pharmaceutically acceptable reagents, including, but not limited to, buffers, stabilizers, and / or polymers. The anti-idiotype antibody can be contacted with the biological sample by pipetting and / or mixing. The anti-idiotype antibody can then specifically bind to a KL2B413-containing protein, such as an antibody or its antigen-binding portion, in the biological sample. As an example, whether an anti-idiotype antibody has bound to KL2B413 can be determined by washing away unbound anti-idiotype antibody, leaving only the complexed anti-idiotype antibody. Continuing this example, the anti-idiotype antibody can contain a fluorophore that can be irradiated to give a signal proportional to the amount of KL2B413 in the biological sample. Detection of the binding complex of the anti-idiotype antibody to KL2B413 is further described below.

[0078] In another aspect, the present disclosure provides an anti-idiotype antibody or antigen-binding portion that specifically binds to an anti-KLK2 antibody, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4-7, a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-14, and a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 19-20, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 25-26, a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 29-30, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 33-34. The six CDRs can be selected according to any known method. The VH and VL CDRs determined according to the Kabat method, the AbM method, the Chothia method, and the contact method are shown in Table 3.

[0079] In certain embodiments, the present disclosure provides an anti-idiotype antibody or antigen-binding portion that specifically binds to an anti-KLK2 antibody, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having the amino acid sequence of SEQ ID NO: 4, a VH CDR2 having the amino acid sequence of SEQ ID NO: 11, and a VH CDR3 having the amino acid sequence of SEQ ID NO: 19, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having the amino acid sequence of SEQ ID NO: 25, a VL CDR2 having the amino acid sequence of SEQ ID NO: 29, and a VL CDR3 having the amino acid sequence of SEQ ID NO: 33.

[0080] In certain embodiments, the present disclosure provides an anti-idiotype antibody or antigen-binding portion that specifically binds to an anti-KLK2 antibody, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having the amino acid sequence of SEQ ID NO: 5, a VH CDR2 having the amino acid sequence of SEQ ID NO: 12, and a VH CDR3 having the amino acid sequence of SEQ ID NO: 19, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having the amino acid sequence of SEQ ID NO: 25, a VL CDR2 having the amino acid sequence of SEQ ID NO: 29, and a VL CDR3 having the amino acid sequence of SEQ ID NO: 33.

[0081] In certain embodiments, the present disclosure provides an anti-idiotype antibody or antigen-binding portion that specifically binds to an anti-KLK2 antibody, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having the amino acid sequence of SEQ ID NO: 6, a VH CDR2 having the amino acid sequence of SEQ ID NO: 13, and a VH CDR3 having the amino acid sequence of SEQ ID NO: 19, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having the amino acid sequence of SEQ ID NO: 25, a VL CDR2 having the amino acid sequence of SEQ ID NO: 29, and a VL CDR3 having the amino acid sequence of SEQ ID NO: 33.

[0082] In certain embodiments, the present disclosure provides an anti-idiotype antibody or antigen-binding portion that specifically binds to an anti-KLK2 antibody, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having the amino acid sequence of SEQ ID NO: 7, a VH CDR2 having the amino acid sequence of SEQ ID NO: 14, and a VH CDR3 having the amino acid sequence of SEQ ID NO: 20, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having the amino acid sequence of SEQ ID NO: 26, a VL CDR2 having the amino acid sequence of SEQ ID NO: 30, and a VL CDR3 having the amino acid sequence of SEQ ID NO: 34.

[0083] In some embodiments, an anti-idiotype antibody, or antigen-binding portion thereof, that specifically binds to an anti-KLK2 antibody comprises a VH domain having an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45, and a VL domain having an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 46. In some embodiments, an anti-idiotype antibody, or antigen-binding portion thereof, that specifically binds to an anti-KLK2 antibody comprises a heavy chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37, and further comprises a light chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 39.

[0084] In some embodiments, an anti-idiotype antibody, or antigen-binding portion thereof, that specifically binds to an anti-KLK2 antibody comprises a VH domain having the amino acid sequence of SEQ ID NO: 45 and a VL domain having the amino acid sequence of SEQ ID NO: 46. In some other embodiments, an anti-idiotype antibody, or antigen-binding portion thereof, that specifically binds to an anti-KLK2 antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 37, and further comprises a light chain comprising the amino acid sequence of SEQ ID NO: 39.

[0085] In some embodiments, the antigen-binding portion is selected from a Fab, F(ab')2, or scFv. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the chimeric antibody comprises a mouse IgG2a framework. In certain embodiments, the mouse IgG2a framework may comprise a mouse Ig heavy chain signal peptide from a mixed FVB / N, C57BL / 6J strain comprising the sequence MAWVWTLLFLMAAAQSIQA (SEQ ID NO: 48).

[0086] In some other embodiments, the antibody is a fully human antibody. For example, the fully human antibody can be IgG, IgM, IgA, IgE, or IgD. In some embodiments, the anti-idiotype antibody or antigen-binding portion thereof is specific for KL2B413, which is within the antigen-binding domain of the extracellular portion of a chimeric antigen receptor (CAR). For example, a nucleic acid encoding KL2B413 can be introduced into T cells in vitro, at least a portion of which is then expressed in the extracellular portion of the CAR. The anti-idiotype antibody can then specifically bind to the extracellular portion of the CAR. In some embodiments, KL2B413 is an scFv, and the anti-idiotype antibody or antigen-binding portion specifically binds to an epitope in the scFv of the CAR. In some embodiments, KL2B413 specifically binds to KLK2. In some embodiments, the antibody or antigen-binding portion does not cross-react with other KLK2 antibodies or other KLK2-binding CARs. For example, to prevent false positives in an assay to determine whether KL2B413 is expressed on the extracellular portion of a CAR, the anti-idiotypic antibody can be specific for KL2B413 and may have no appreciable binding to its target KLK2 or other KLK2-targeting ligands that are not KL2B413. In some embodiments, the CAR has an amino acid sequence selected from the group consisting of SEQ ID NOs: 43-44.

[0087] In some embodiments, the present disclosure provides nucleic acids encoding the heavy chain, light chain, or both of an anti-idiotype antibody or antigen-binding portion. For example, the nucleic acid can be DNA, RNA, and any chemical modifications thereof (e.g., nucleoside modifications).

[0088] In another aspect, the present disclosure provides a nucleic acid encoding the heavy chain, light chain, or both of an anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to KL2B413, the nucleic acid comprising the nucleotide sequence of SEQ ID NO: 38, the nucleotide sequence of SEQ ID NO: 40, or both. In another aspect, the present disclosure provides a vector comprising the nucleic acid sequence. For example, the vector may be a self-replicating nucleic acid structure or may be integrated into the genome of a host cell into which it is introduced. In some embodiments, the vector is an expression vector. In another aspect, the present disclosure provides a host cell comprising the vector. In some embodiments, the host cell is a mammalian cell.

[0089] In another aspect, the present disclosure provides a method for producing an anti-idiotype antibody or antigen-binding portion thereof that specifically binds to KL2B413, the method comprising culturing host cells comprising nucleotide sequences encoding the heavy and light chains of the antibody or antigen-binding portion under conditions conducive to expression of the antibody or antigen-binding portion, and isolating the antibody or antigen-binding portion from the culture. For example, anti-idiotype antibodies can be produced by homogenous suspension culture in deep-tank stirred fermenters, perfusion tank systems, airlift reactors, and continuous culture systems. Anti-idiotype antibodies can be isolated from reaction and / or growth mixtures by physical or chemical separation procedures, including affinity separation using Protein A or G, size exclusion chromatography, and charge separation. In some embodiments, the host cells encode a vector comprising a nucleic acid encoding the anti-idiotype antibody or antigen-binding portion thereof.

[0090] In another aspect, the present disclosure provides a method for detecting KL2B413 in a biological sample, comprising: (a) preparing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion; and (c) detecting the anti-idiotype antibody or antigen-binding portion. For example, an anti-idiotype antibody may bind to KL2B413 expressed in the biological sample. The bound complex can be detected by any detection method, including both chemical and physical detection methods. For example, the detection method can be used to identify the mere presence of the antibody of interest in the biological sample, or to test whether the antibody of interest is present at a detectable level in the sample, or to quantify the amount of the antibody of interest in a sample and further compare antibody levels from different samples. For example, the detection method can be one or more of immunoprecipitation, immunocytochemistry, immunoblotting, and immunosorbent assay. Specifically, the immunosorbent assay can be an ELISA or ELISA-type assay, in which the biological sample contains a bound anti-idiotype antibody or fragment thereof.

[0091] In another aspect, the present disclosure provides a method for detecting expression of a chimeric antigen receptor (CAR) comprising KL2B413 in a biological sample, the method comprising: (a) providing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion; and (c) detecting the anti-idiotype antibody or antigen-binding portion, thereby detecting expression of the CAR.

[0092] In some embodiments, the antibody comprises a detectable label. In some embodiments, the method further comprises contacting the anti-idiotype antibody or antigen-binding portion with a detectable label prior to detecting the anti-idiotype antibody or antigen-binding portion. For example, the detectable label can be any chemical tag or moiety that associates with, binds to, or otherwise complexes with the anti-idiotype antibody and emits or otherwise provides a unique, identifiable signal. For example, the detectable label can be an isotopic marker, a colorimetric biosensor, a photochromic compound, a fluorescent label, a fluorogenic label, or an electrochemical sensor. Specific examples of fluorescent labels include green fluorescent protein, yellow fluorescent protein, blue fluorescent protein, fluorescein, rhodamine, coumarin, cyanine, phycoerythrin, and derivatives thereof.

[0093] In some embodiments, the biological sample is blood, serum, or urine. For example, the biological sample can be whole blood, serum, plasma, urine, feces, cerebrospinal fluid, ascites, etc. In some embodiments, the biological sample is fresh biological material, such as biological material collected at a given time for this analysis. The biological sample can also be biological material collected at another time during the patient's treatment for this or other purposes, or biological material using archived patient material. The biological sample can be freshly obtained or previously obtained, or if previously obtained, can be stored (e.g., at room temperature, refrigerated, or frozen) before use.

[0094] In some aspects, the present disclosure provides a kit for detecting KL2B413 in a biological sample, comprising: (a) an anti-idiotype antibody or antigen-binding portion; and (b) instructions for detecting the anti-idiotype antibody or antigen-binding portion. For example, the kit may comprise the anti-idiotype antibody or antigen-binding portion as a solid powder, a lyophilized powder, a liquid solution, or liquid components mixed to form a solution, or bound to a solid support. The kit may also include additional reagents, including stabilizers, buffers, and other pharmaceutically acceptable excipients, necessary to facilitate use of the kit in assaying a biological sample. The kit may also include instructions instructing the user on how to perform the assay.

[0095] In another aspect, the present disclosure provides a method for purifying KL2B413 from a sample, the method comprising: (a) providing a biological sample containing KL2B413; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion of the present disclosure; and (c) capturing the anti-idiotype antibody or antigen-binding portion, including a CAR or other protein containing KL2B413, thereby purifying KL2B413. For example, any separation method, including physical and chemical methods, can be used to capture the anti-idiotype antibody. Specifically, KL2B413 can be captured and isolated from cell culture medium, host cells, or both using techniques known in the art for purifying proteins, including ion exchange chromatography, gel filtration chromatography, ultrafiltration, electrophoresis, and immunoaffinity purification using an antibody specific for a particular epitope of the anti-idiotype antibody. In some embodiments, the anti-idiotype antibody is a fusion protein containing a domain that facilitates its purification. In certain embodiments, a purified KL2B413 composition is substantially isolated from non-KL2B413-containing proteins, hi some embodiments, a purified KL2B413 composition is 100% pure, 99% pure, 98% pure, 97% pure, 96% pure, 95% pure, or 90% pure or greater.

[0096] In another aspect, the disclosure provides a method of selecting CAR-T cells from a cell population, the method comprising: (a) providing a biological sample containing CAR-T cells; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion thereof; and (c) capturing the anti-idiotype antibody or antigen-binding portion thereof, thereby selecting the CAR-T cells. In some embodiments, the anti-idiotype antibody or antigen-binding portion thereof is specific for KL2B413.

[0097] Embodiment Specific non-limiting embodiments of the present invention are described in the following numbered paragraphs. 1. An anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to a target antibody, including KL2B413. 2. The anti-idiotype antibody or antigen-binding portion thereof of paragraph 1, wherein the target antibody or antigen-binding portion thereof comprises a VH domain having the amino acid sequence of SEQ ID NO: 41 and a VL domain having the amino acid sequence of SEQ ID NO: 42. 3. An anti-idiotype antibody or antigen-binding portion thereof that specifically binds to KL2B413, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 to 7, a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11 to 14, and a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 19 to 20, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 25 to 26, a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 29 to 30, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 33 to 34. 4. The anti-idiotype antibody or antigen-binding portion of any one of Items 1 to 3, wherein the VH domain has an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45, and the VL domain has an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 46. 5. The anti-idiotype antibody or antigen-binding portion of any one of Items 1 to 3, comprising a heavy chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37, and further comprising a light chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 39. 6. The anti-idiotype antibody or antigen-binding portion according to any one of items 1 to 3, wherein the VH domain has the amino acid sequence of SEQ ID NO: 45 and the VL domain has the amino acid sequence of SEQ ID NO: 46. 7. The anti-idiotype antibody or antigen-binding portion according to any one of Items 1 to 3, comprising a heavy chain comprising the amino acid sequence of SEQ ID NO: 37 and further comprising a light chain comprising the amino acid sequence of SEQ ID NO: 39. 8. The anti-idiotype antibody or antigen-binding portion of any one of items 1 to 3, wherein the antigen-binding portion is selected from Fab, F(ab')2, or scFv. 9. The anti-idiotype antibody or antigen-binding portion according to any one of items 1 to 3, wherein the antibody is a monoclonal antibody. 10. The anti-idiotype antibody or antigen-binding portion of any one of items 1 to 3, wherein the antibody is a chimeric antibody. 11. The anti-idiotype antibody or antigen-binding portion of paragraph 10, wherein the antibody comprises a murine IgG2a framework. 12. The anti-idiotype antibody or antigen-binding portion of any one of items 1 to 3, wherein the antibody is a fully human antibody. 13. A nucleic acid encoding the heavy chain, the light chain, or both of the anti-idiotype antibody or antigen-binding portion thereof according to any one of items 1 to 3. 14. A nucleic acid encoding the heavy chain, the light chain, or both of an anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to KL2B413, a) the nucleotide sequence of SEQ ID NO: 38; b) the nucleotide sequence of SEQ ID NO: 40; c) A nucleic acid comprising both a) and b). 15. A vector comprising the nucleic acid according to item 14. 16. The vector according to paragraph 15, wherein the vector is an expression vector. 17. A host cell comprising the vector according to paragraph 16. 18. The host cell of paragraph 17, wherein the cell is a mammalian cell. 19. A method for producing an anti-idiotype antibody or antigen-binding portion thereof that specifically binds to KL2B413, the method comprising culturing the host cell described in paragraph 17, which contains nucleotide sequences encoding the heavy and light chains of the antibody or antigen-binding portion, under conditions that allow expression of the antibody or antigen-binding portion, and isolating the antibody or antigen-binding portion from the culture. 20. A method for detecting KL2B413 in a biological sample, comprising: (a) preparing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in any one of items 1 to 3; and (c) detecting the anti-idiotype antibody or antigen-binding portion. 21. A method for detecting expression of a chimeric antigen receptor (CAR) comprising KL2B413 in a biological sample, the method comprising: (a) preparing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in any one of paragraphs 1 to 3; and (c) detecting the anti-idiotype antibody or antigen-binding portion, thereby detecting CAR expression. 22. The method of paragraph 20, wherein the antibody comprises a detectable label. 23. The method of paragraph 20, further comprising contacting the anti-idiotype antibody or antigen-binding portion with a detectable label prior to detecting the anti-idiotype antibody or antigen-binding portion. 24. The method according to item 20, wherein the biological sample is blood, serum or urine. 25. The anti-idiotype antibody or antigen-binding portion of any one of paragraphs 1 to 3, wherein KL2B413 is within the antigen-binding domain of the extracellular portion of a chimeric antigen receptor (CAR). 26. The anti-idiotype antibody or antigen-binding portion of clause 25, wherein KL2B413 is an scFv and the anti-idiotype antibody or antigen-binding portion specifically binds to an epitope in the scFv of the CAR. 27. The anti-idiotype antibody or antigen-binding portion of paragraph 25, wherein KL2B413 binds to KLK2. 28. The anti-idiotype antibody or antigen-binding portion of paragraph 25, wherein the antibody or antigen-binding portion does not cross-react with other KLK2 antibodies or other KLK2-binding CARs. 29. The anti-idiotype antibody or antigen-binding portion of paragraph 25, wherein the CAR has an amino acid sequence selected from the group consisting of SEQ ID NOs: 43-44. 30. A kit for detecting KL2B413 in a biological sample, comprising: (a) an anti-idiotype antibody or antigen-binding portion described in any one of items 1 to 3; and (b) instructions for detecting the anti-idiotype antibody or antigen-binding portion. 31. An anti-idiotype antibody or antigen-binding portion of any one of items 1 to 3 for use in detecting KL2B413 in a biological sample, wherein the detection comprises (a) providing a biological sample, (b) contacting the biological sample with the anti-idiotype antibody or antigen-binding portion, and (c) detecting the anti-idiotype antibody or antigen-binding portion. 32. A method for purifying KL2B413 from a sample, comprising: (a) preparing a biological sample containing KL2B413; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in any one of items 1 to 3; and (c) capturing the anti-idiotype antibody or antigen-binding portion, thereby purifying KL2B413. 33. A method for selecting CAR-T cells from a cell population, the method comprising: (a) providing a biological sample containing CAR-T cells; (b) contacting the biological sample with the anti-idiotype antibody or antigen-binding portion of any one of paragraphs 1 to 3; and (c) capturing the anti-idiotype antibody or antigen-binding portion, thereby selecting CAR-T cells. [Example]

[0098] Example 1: Determination of KL2B513 or KL2B610 binding Fab As described in Shi et al., J Mol Biol 397:385-96, 2010, WO 2009 / 085462, and U.S. Patent Application Publication No. 2010 / 0021477, KL2B413-derived scFv fusion protein-binding Fabs were selected from two sets of de novo Fab-pIX phage display libraries. Two KL2B413-derived scFv fusion proteins, KL2B513 (VL-VH) and KL2B610 (VH-VL), exist, with different orientations of VH and VL within the scFv.

[0099] In phage selection using purified recombinant antigen, biotinylated KL2B513 or KL2B610 was used as "bait" to capture and immobilize phage binders. After several selection rounds, polyclonal phage ELISA using purified antigen was performed to detect specific enrichment in individual panning experiments. Phage collected from these panning experiments that showed enrichment of binders to KL2B513 or KL2B610 were expressed in E. coli for primary screening. Monoclonal Fab lysates were prepared from the enriched Fab library and screened by ELISA for binding to KL2B513 or KL2B610, but not to the similar negative control scFv fusion proteins GC5B734 or GCDB332. Selected Fabs were sequenced to identify unique Fab clones, and their V region genes were isolated. The unique Fab V regions were cloned into mammalian expression vectors to express chimeric mAbs with mouse IgG2a / mouse kappa constant regions. The chimeric mAbs were evaluated for specific binding to the scFv from KL2B413 expressed on SupT1 cells corresponding to the scFv in the KL2B513 or KL2B610 scFv fusion proteins, and binding kinetics were measured using SPR.

[0100] Phage panning Six separate panning experiments using separate V3.0 and V5.0 de novo Fab phage libraries were performed against biotinylated KL2B513 or KL2B610 according to standard protocols (Cheadle, EJ et al. Antibody Engineering. 907, 645-666 (2012). Briefly, the phage library and paramagnetic streptavidin (SA) beads were blocked with 50% Chemiblocker (Millipore catalog no. 2170) / 50% 1xTBST (Teknova catalog no. T0310) for 1 hour. The library was added to the SA beads to adsorb clones that nonspecifically bound to the beads. The SA beads were discarded, and the pre-adsorbed library was added to biotinylated KL2B513 or KL2B610 in the presence of 100 nM GP5B305. SA beads were added to form bead / antigen / phage complexes, and binders were recovered by washing with 1xTBST. After the final wash, phage were rescued by infection of log-phase TG1 E. coli cells (OD600nm = 0.4-0.6). Phage-infected TG1 cells were cultured in 150 mM PBS containing 75 μg / mL carbenicillin and 1% glucose. The phage were spread onto three LB / agar plates and grown overnight at 37°C. Phages were generated and subjected to additional panning rounds. To increase the selection pressure, the antigen concentration was decreased in each subsequent round at room temperature: R1 100 nM for 1 hour; R2 50 nM for 1 hour; R3 10 nM for 1 hour; and R4 5 nM for 1 hour.

[0101] Polyclonal phage ELISA Binder enrichment was determined from each panning experiment by polyclonal phage ELISA. Briefly, 100 μL of 20 nM non-biotinylated KL2B513 or KL2B610 diluted in 1x TBS (Teknova catalog no. T9530) was captured onto an NA-coated plate (Thermo catalog no. 15217). After incubation at 37°C for 1 hour, the plate was washed three times with 300 μL of 1x TBST. 300 μL of blocking buffer (50% Chemiblocker / 50% 1x TBST) was added to each well of the plate and incubated for 1 hour at room temperature. After blocking, the plate was washed three times with 300 μL of 1x TBST. 100 μL of polyclonal phage output from each panning round, diluted 1 / 100 in assay buffer, 10% Chemiblocker / 90% TBST, was added to an ELISA plate and incubated at room temperature for 1 hour to allow Fab displayed on the phage particles to bind to immobilized KL2B513 or KL2B610. After incubation, the plate was washed three times with 1x TBST. 100 μL of HRP-conjugated anti-M13(pVIII) antibody (GE Healthcare catalog no. 27942101), diluted 1:2500 in assay buffer, was added to the plate and incubated at room temperature. After 1 hour of incubation, the plate was washed six times with 300 μL of 1x TBST. 100 μL of prepared BM chemiluminescent ELISA substrate (Roche catalog no. 11582950001) was added to the plate. Chemiluminescence or relative light units (RLU) were measured using an Envision plate reader.

[0102] Figure 1 shows the results of polyclonal phage ELISA. Panning outputs from rounds 1 to 4 from 12 panning experiments were screened on NA-coated plates for binding to KL2B513 or KL2B610 and the negative control GCDB332. Panning outputs with specific enrichment were selected for monoclonal Fab production. A002B39 was derived from the APD316XP_19 panning experiment.

[0103] Fab production Plasmid DNA was isolated and purified from glycerol stocks of specific rounds of phage panning experiments identified to show enrichment of binders to KL2B513 or KL2B610, transformed into TG-1 E. coli cells, and grown overnight on LB / agar plates. The overnight cultures were used for (i) colony PCR and V-region sequencing, and (ii) starting cultures for Fab production. For Fab production, the overnight cultures were diluted 10-100 times in fresh medium and grown at 37°C for 5-6 hours. Fab production was induced by adding fresh medium containing IPTG, and the cultures were grown overnight at 30°C. The cultures were spun down, and the bacterial pellets were lysed using BugBuster™ (Millipore) to release the soluble Fab protein. The cell lysates were spun down, and the supernatants were used for Fab ELISA.

[0104] Primary screening Phage collected from panning experiments that showed enrichment of binders to KL2B513 or KL2B610 were expressed in E. coli for primary screening. MSD 384-well streptavidin plates (Meso Scale #L21SA-5) were blocked with 50 μL / well of SuperBlock T20 (TBS) blocking buffer (Thermo #37536) at room temperature for 30 minutes and then aspirated. 2.5 nM of biotinylated ScFv-Fc fusion antigen, target antigen KL2B513 or KL2B610, or counterscreening reagent GCDB734 was added to the blocked 384-well MSD plate and incubated on a shaker at room temperature for 30 minutes. The plate was washed twice with 80 μL / well of 1× phosphate-buffered saline with Tween detergent ("1×PBST"), and crude Fab lysates from E. coli expression in the 96-well plate were stamped in duplicate onto a 384-well assay plate and incubated for 1 hour at room temperature on a shaker. The assay plate was washed once with 80 μL / well of 1×PBST, and then 6 nM SULFO-TAG anti-human / NHP kappa antibody (Meso Scale #D20TF-6) was added at 20 μL / well, and the plate was incubated for 1 hour at room temperature on a shaker. The plate was washed once with 80 μL / well of 1× PBST, and 35 μL / well of 1× MSD Read Buffer T (Meso Scale #R92TC-1) was added to each well, and the plate was analyzed on an MSD Sector S600 plate reader. All liquid handling was performed on an Agilent Bravo system, and washing of 384-well plates was performed on a BioTek 405 Select plate washer.

[0105] Figure 2 shows the results of the primary screening. Monoclonal Fabs were screened on MSD plates for binding to KL2B513 or KL2B610. Clones with signals greater than the mean background signal plus three times the standard deviation in the KL2B513 or KL2B610 target binding assay and less than the mean background signal plus three times the standard deviation in the GCDB734 counter reagent binding assay were selected for sequencing. Clones identified by asterisks are the parent clones of the VH and VL of A002B39.

[0106] Example 2: Production of monoclonal antibodies against KL2B413-LH / HL-scFv (A002B39) Fab Selection Selected Fabs from the primary screen were sequenced to determine the V-region sequences and identify unique clones. Unique Fab V-regions were cloned into mammalian expression vectors and expressed as chimeric mAbs with mouse IgG2a / mouse kappa constant regions.

[0107] The variable regions of A002B39 were identified by phage display using a human Fab-pIX de novo library against the soluble scFv-Fc fusion proteins KL2B513 and KL2B610. These V regions were not subjected to any affinity maturation. The DNA sequences were obtained from a de novo Fab library without codon optimization.

[0108] V H and V L Cloning Two pcDNA3.1-derived mammalian expression vectors (vDR000368 and vDR000961) were used to generate single-gene constructs encoding the heavy chain (HC) or light chain (LC) of the chimeric mAb. Each vector contains a human cytomegalovirus (hCMV) promoter driving expression of the HC and LC, and both contain the ampicillin resistance gene (Amp®) to facilitate cloning. vDR000368 also contains unique HindIII and DraIII restriction enzyme sites for cloning, as well as the mouse IgG2a constant region. vDR000961 also contains unique HindIII and Tth111I restriction enzyme sites for cloning, as well as the mouse kappa constant region.

[0109] HC(V H ) or LC(V L A DNA fragment containing the variable region of the V-terminal fragment was synthesized and ligated into the HC vector vDR000368 and the LC vector vDR000961. The HC synthetic fragment contained a HindIII restriction enzyme site, a Kozak sequence, a signal peptide, and a V-terminal fragment. H and C H1 The LC synthetic fragment contained a DNA sequence encoding a portion of the nucleotide sequence ... L The constructs contained DNA sequences encoding the HC, LC, and kappa constant regions, as well as a Tth111I restriction cloning site. The final HC construct is PBD000105920 and the LC construct is PBD000105921. The two constructs were co-transfected into mammalian expression cell lines HEK293 Expi or CHO to generate A002B39.

[0110] Protein expression The HC construct PBD000105920 and the LC construct PBD000105921 were sequence verified prior to transfection. HEK Expi293™ cells (Thermo Catalog No. A14527) were grown in Expi293™ Expression Medium (Thermo Catalog No. A1435101). Cells were grown at 37°C with 125 RPM shaking in 8% CO2. Cells were transfected at 2.5 x 10 per mL using the Expi293™ Expression Kit (Thermo Catalog No. A14524). 6 1000 cells were transfected at 1000 ng / L. For each liter of transfected cells, 1 mg of total DNA was diluted in 25 mL of Opti-MEM (Thermo catalog number 319850620), and 2.6 mL of Expi293™ reagent was diluted in 25 mL of Opti-MEM and incubated at room temperature for 5 minutes. The diluted DNA and diluted Expi293 reagent were combined and incubated at room temperature for 20 minutes. The DNA complex was then added to the cells. The cells were placed in a shaking incubator overnight. The day after transfection, 5 mL of Enhancer 1 was diluted in 50 mL of Enhancer 2, and the total amount of both Enhancers was added to the cells. The transfected cells were returned to the incubator for 4 days before being harvested. The cells were removed by centrifugation at 4,500 g for 35 minutes and then filtered through a 0.2 μm filter before expression levels were confirmed.

[0111] Expression was quantified using an Octet instrument. Mouse IgG2 (Sigma catalog number M9144) was used as a standard. A Protein A biosensor was used. Samples and standards were diluted in spent Expi293 medium. The standard curve started at 100 μg / mL with two-fold dilutions. Samples were diluted 1:10. The standard curve was a linear point curve. Calculations were performed by Forte Biosystems software.

[0112] Example 3: Binding assay of anti-idiotypic antibody A002B39 Biacore binding assay using soluble proteins Anti-mouse Fc antibody directly immobilized on an M5 sensor chip. Library antibody (mouse IgG2a) diluted to 1 μg / mL to achieve approximately 20-50 RU. Antigen (scFV Fc fusion) was allowed to associate for 3 minutes at 500 nM-0.8 nM, 1:5 dilution. Dissociation was allowed for 30 minutes. Assay results (Table 2 below) show binding affinity for KLB513 similar to that seen with ProteOn.

[0113] Binding assays using soluble proteins by Proteon and Biacore Binding assays were performed using a ProteOn XPR36 system (BioRad). ProteOn GLC chips (BioRad, catalog number 176-5011) were coated with anti-mouse Fc antibody. Library antibodies (mouse IgG2a) were diluted from 0.25 μg / mL to 1 μg / mL to approximately 100-200 RL. Antigen (scFv-Fc fusion) was allowed to associate for 3 minutes at 1000 nM to 0.4 nM, 1:4 dilution, followed by 30 minutes of dissociation. The assay results (Table 1 below) show similar binding affinities of A002B39 to both KLB513 and KLB610.

[0114] [Table 1]

[0115] Anti-mouse Fc antibody directly immobilized on an M5 sensor chip. Library antibody (mouse IgG2a) was diluted to 1 μg / mL, approximately 40-80 RU. Antigen (scFV Fc fusion) was incubated at 500 nM-0.8 nM, 1:5 dilution, for 3 minutes. Dissociation was allowed for 30 minutes. Assay results (Table 2 below) show the binding affinity of A002B39 to KLB513.

[0116] [Table 2]

[0117] Cell binding assay ScFv-transfected SupT1-KL2B413 HL, SupT1-KL2B413 LH, or SupT1-CD9B337-HL cells were cultured in RMPI 1640, 10% FBS, 1% non-essential amino acids, 1 mM sodium pyruvate, 2 mM L-glutamine, 10 mM HEPES, and 0.1% bicarbonate. Cell culture supplements were from ThermoFisher Scientific (Gibco). mAbs were diluted to 6 micrograms / mL in staining buffer (BSA) (BD Pharmingen catalog number 554657). scFv-expressing SupT1 cells were labeled with fixable Live / Dead stain (Molecular Probes #L34974) and plated at 50,000 cells / well into a 384-well V-bottom microplate (Greiner Bio-One #781281).

[0118] Normalized mAb samples were added to the cell suspension at 20 μL / well with gentle mixing, and the cells were incubated on ice for 30 minutes. The cell suspension was diluted with 70 μL / well of ice-cold staining buffer (BSA), and the cells were pelleted at 400 × g for 5 minutes at 4°C. The supernatant was aspirated. The cell pellet was washed again with 70 μL / well of ice-cold staining buffer (BSA). 3 μg / mL of AF488 anti-mouse IgG (H+L)-specific goat F(ab')2 (Jackson ImmunoResearch #115-546-062) was added to the cell pellet at 40 μL / well with gentle mixing, and the cells were incubated on ice in the dark for 30 minutes. The cells were washed as described above and fixed with 40 μL / well of BD Cytofix (BD Pharmingen #554655) for 20 minutes on ice. Fixed cells were washed as described above, and the cell pellet was resuspended in 20 μL of staining buffer and analyzed on an iQue PLUS VBR flow cytometer. Cells were gated on the viable and singlet populations and analyzed for antibody binding in the BL1-H (AF488) channel. All liquid handling was performed on an Agilent Bravo system, and 384-well plate aspiration was performed on a BioTek 405 Select plate washer.

[0119] As shown in Figure 3, a number of candidate mAbs were screened for binding to scFv-transfected SupT1 cells. A002B39 (labeled A002M39) showed specific binding to both SupT1-KL2B413 HL and SupT1-KL2B413 LH cells, but not to the negative control SupT1-CD9B337-HL.

[0120] Example 4: Characterization of detection antibodies against KLK2 CAR KL2B413-LH (Protein ID#A002B39) on CAR+SupT1 cells Antibodies detecting the KLK2 CAR (KL2B413-LH) expressed on NK cells and T cells were identified from a panel of proteins derived from phage display screening. As discussed in the previous example, the proteins were first tested for binding to recombinant CAR protein, and potential binders were scaled up. The proteins were purified and tested for dose-dependent binding to SupT1 cells expressing KL2B413-LH, respectively, by flow cytometry. Specific binding to CAR was confirmed by competitive binding experiments with Fc-KL2B413-LH fusion protein and lack of binding to parental SupT1 cells. After selecting the best binders, the antibodies were directly conjugated to recombinant phycoerythrin ("PE") for use as CAR detection reagents. The antibodies were purified to a 1:1 PE:antibody ratio to enable receptor enumeration studies (number of CARs expressed on the cell surface).

[0121] purification The cell culture supernatant was loaded onto a MabSelect column and eluted with a low pH buffer, such as 100 mM sodium acetate, pH 3.0, followed by buffer exchange into 1x SSC, 8.5% sucrose, pH 7.0, using a Sephadex G-25 column. Protein-containing fractions were collected. After purification, the protein was QCed using SDS-PAGE, SEC-HPLC, and LC-MS.

[0122] Phycoerythrin labeling One microliter of Modifier Reagent was added to every 10 μL of antibody to be labeled and mixed gently. The antibody sample (with Modifier Reagent) was pipetted directly into lyophilized PE (Expedeon, catalog no. 703-0015), gently resuspended, and incubated at room temperature (20-25°C) in the dark for 1 hour. One microliter of Quencher Reagent was added to every 10 μL of antibody used and incubated for 30 minutes.

[0123] After labeling, the PE-antibody conjugate was purified on a size-exclusion column. Fractions were collected and analyzed by SEC-HPLC. Fractions containing only one antibody with one PE were pooled together and concentrated as needed. The final product was analyzed by SEC-HPLC.

[0124] Characterization of KLK2 CAR (KL2B413-LH) anti-idiotypic antibody A002B39 SupT1 cells expressing KL2B413LH were compared with parental CAR-SupT1 cells. Cells (100,000 cells / well) were stained with LIVE / DEAD Fixable Near-IR viability dye (Life Technologies L10119) and incubated with increasing concentrations of A002B39 for 30 minutes on ice. After incubation, samples were washed with BSA staining buffer (BD Biosciences #554657) and stained with PE goat anti-mouse IgG polyclonal antibody (Biolegend #405307) to detect bound antibody on live CAR+SupT1 cells. After incubation, washing, and fixation (Cytofix, BD Biosciences), samples were acquired on a 10-color FACSCanto II (BD Biosciences) flow cytometer. Analysis was performed using FlowJo, and the PE median fluorescence intensity of live SupT1 cells is plotted in Figure 4. As shown in Figure 4, dose-dependent binding of A002B39 to KL2B413-LH SupT1 cells was observed. No binding was detected in CAR-SupT1 parental cells.

[0125] A002B39 was conjugated to R-PE as described above and tested for binding to KL2B413-LH SupT1 in the presence or absence of 10 μg / mL KL2B413-LH scFv Fc fusion protein to assess the specificity of A002B39 binding to the KL2B413 CAR. KL2B413-LH SupT1 cells (100,000 cells / well) were stained with LIVE / DEAD Fixable Near-IR viability dye (Life Technologies L10119) and incubated with increasing concentrations of PE-A002B39 and 10 μg / mL KL2B413-LH scFv Fc for 30 minutes on ice. After incubation, washing, and fixation, samples were acquired on a 10-color FACSCanto II flow cytometer. Analysis was performed using FlowJo analysis software, and the PE median fluorescence intensity of viable SupT1 cells is plotted in Figure 5. As shown in Figure 5, the dose-dependent binding of PE-A002B39 to KL2B413-LH SupT1 cells is specific for KL2B413-LH CAR and can be blocked by 10 μg / mL of KL2B413-LH scFv-Fc.

[0126] To reflect the detection of CAR cells in clinical samples, KL2B413LH SupT1 cells were spiked into whole blood. As shown in Figure 3 below, KL2B413-LH CAR detection by PE-A002B39 was not inhibited in the presence of whole blood. KL2B413LH SupT1 cells were stained with LIVE / DEAD Fixable Near-IR viability dye, washed, and suspended in staining buffer or fresh whole blood (n=2). 100,000 KL2B413LH SupT1 cells were seeded at 50 μL / well and incubated on ice for 30 minutes with increasing concentrations of PE-A002B39. After incubation, washing, and fixation, samples were acquired on a 10-color FACSCanto II flow cytometer. Analysis was performed using FlowJo software, and the PE median fluorescence intensity of viable SupT1 cells is plotted in Figure 6. As shown in Figure 6, PE-A002B39 detection of KL2B413-LH-SupT1 cells spiked in whole blood.

[0127] Multiple lots of PE-A002B39 were compared to assess differences in binding to KL2B413LH-SupT1 cells between batches. KL2B413-LH SupT1 cells (200,000 cells / well) were stained with LIVE / DEAD Fixable Near-IR viability dye and incubated on ice for 30 minutes with increasing concentrations of PE-A002B39. After incubation, washing, and fixation, samples were acquired on a 10-color FACSCanto II flow cytometer. Analysis was performed using FlowJo analysis software, and the PE median fluorescence intensity of viable SupT1 cells is plotted in Figure 7. As shown in Figure 7, the two different lots of PE-A002B39 have similar binding profiles to KL2B413-LH-SupT1.

[0128] array

[0129] [Table 3-1]

[0130] [Table 3-2]

[0131] SEQ ID NO: 37: Heavy chain of A002B39 with muIgG2a (amino acids) EVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPGQGLEWMGGIIPIFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARVQWGLDYWGQGTL VTVSSAKTTAPSVYPLAPVCGDTTGSSVTLGCLVKGYFPEVTLTWNSGSLSSGVHTFPAVLQSDLYTLSSSVTVTSSTWPSQSITCNVAHPASSTKVDKKIEPRGPTIKPC PPCKCPAPNLLGGPSVFIFPPKIKDVLMISLSPIVTCVVVDVSEDDPDVQISWFVNNVEVHTAQTQTHREDYNSTLRVVSALPIQHQDWMSGKEFKCKVNNKDLPAPIERT ISKPKGSVRAPQVYVLPPPEEEMTKKQVTLTCMVTDFMPEDIYVEWTNNGKTELNYKNTEPVLDSDGSYFMYSKLRVEKKNWVERNSYSCSVVHEGLHNHHTTKSFSRTPGK

[0132] SEQ ID NO: 38: Heavy chain of A002B39 with muIgG2a (DNA)

[0133] SEQ ID NO: 39: Light chain of A002B39 with muKappa (amino acids) EIVLTQSPATLSLSPGERATLSCRASQSVDSALAWYQQKPGQAPRLLIYGASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRFNWPITFGQGTKVEIK RADAAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC

[0134] SEQ ID NO: 40: Light chain of A002B39 with muKappa (DNA) GAAATTGTGCTGACCCAGAGCCCGGCGACCCTGAGCCTGAGCCCGGGCGAACGCGCGACCCTTAGCTGCCGTGCAAGTCAGAGTGTGGACAGCGCGCTGGCGTGGTATCAGCAGAAACCGGGCCAGGCGCCGCGCCTGCTGATTTATGGTGCGAGCAACCGCGCGACCGGCATTCCGGCGCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGACCATTAGCAGCCTGGAACCGGAAGATTTTGCGGTGTATTATTGCCAGCAGCGTTTCAACTGGCCGATCACCTTTGGCCAGGGCACCAAAGTGGAAATTAAACGGGCTGATGCTGCACCGACTGTGTCCATCTTCCCACCATCCAGTGAGCAGTTAACATCTGGAGGTGCCTCAGTCGTGTGCTTCTTGAACAACTTCTACCCCAAAGACATCAATGTCAAGTGGAAGATTGATGGCAGTGAACGACAAAATGGCGTCCTGAACAGTTGGACTGATCAGGACAGCAAAGACAGCACCTACAGCATGAGCAGCACCCTCACGTTGACCAAGGACGAGTATGAACGACATAACAGCTATACCTGTGAGGCCACTCACAAGACATCAACTTCACCCATTGTCAAGAGCTTCAACAGGAATGAGTGT

[0135] Accession number 41: KL2B413 VH EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYWMTWVRQAPGKGLEWVANIKQDGSERYYVDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDQNYDILTGHYGMDVWGQGTTVTVSS

[0136] Accession number 42: KL2B413 VL EIVLTQSPSFLSASVGDRVTITCRASQGISSYLSWYQQKPGKAPKLLIYATSTLQSGVPSRFSGSGSGTEFTLTISSLQPEDFATYYCQQLNSYPRTFGQGTKVEIK

[0137] SEQ ID NO: 43 - CAR1 (KL2B413_HL; pBD000091628) EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYWMTWVRQAPGKGLEWVANIKQDGSERYYVDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDQNYDILTGHYGMDVWGQGT TVTVSSGGSEGKSSGSGSESKSTGGSEIVLTQSPSFLSASVGDRVTITCRASQGISSYLSWYQQKPGKAPKLLIYATSTLQSGVPSRFSGSGSGTEFTLTISSLQPEDFATYYCQQLNS YPRTFGQGTKVEIKTSTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEE EEGGCELRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR

[0138] SEQ ID NO: 44 - CAR2 (KL2B413_LH; pBD000091623) EIVLTQSPSFLSASVGDRVTITCRASQGISSYLSWYQQKPGKAPKLLIYATSTLQSGVPSRFSGSGSGTEFTLTISSLQPEDFATYYCQQLNSYPRTFGQGTKVEIKGGSEGKSSGSG SESKSTGGSEVQLVESGGGLVQPGGSLRLSCAASGFTFSSYWMTWVRQAPGKGLEWVANIKQDGSERYYVDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDQNYDILTGHYG MDVWGQGTTVTVSSTSTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEE EEGGCELRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR

[0139] SEQ ID NO: 45: Heavy chain variable domain of A002B39 (amino acids) EVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPGQGLEWMGGIIPIFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARVQWGLDYWGQGTLVTVSS

[0140] SEQ ID NO: 46: Light chain variable domain of A002B39 (amino acids) EIVLTQSPATLSLSPGERATLSCRASQSVDSALAWYQQKPGQAPRLLIYGASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRFNWPITFGQGTKVEIK

[0141] SEQ ID NO: 47 - Human kallikrein-2 sequence (signal sequence: amino acids 1-18) MWDLVLSIALSVGCTGAVPLIQSRIVGGWECEKHSQPWQVAVYSHGWAHCGGVLVHPQWVLTAAHCLKKNSQVWLGRHNLFEPEDTGQRVPVSHSFPHPLYNMSLLKHQSLRPDEDSSHDLMLLRLSEPA KITDVVKVLGLPTQEPALGTTCYASGWGSIEPEEFLRPRSLQCVSLHLLSNDMCARAYSEKVTEFMLCAGLWTGGKDTCGGDSGGPLVCNGVLQGITSWGPEPCALPEKPAVYTKVVHYRKWIKDTIAANP

[0142] Incorporation by Reference All publications and patent applications mentioned in this specification are herein incorporated by reference in their entirety to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

[0143] While specific embodiments of the subject disclosure have been discussed, the above specification is illustrative and not restrictive. Many variations of the present disclosure will become apparent to those skilled in the art upon consideration of this specification and the claims that follow. The full scope of the disclosure should be determined by reference to the claims, along with their full scope of equivalents, and the specification, along with such variations. Various embodiments of the present invention are described below. 1. An anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to a target antibody, including KL2B413. 2. The anti-idiotype antibody or antigen-binding portion thereof according to claim 1, wherein the target antibody or antigen-binding portion thereof comprises a VH domain having the amino acid sequence of SEQ ID NO: 41 and a VL domain having the amino acid sequence of SEQ ID NO: 42. 3. An anti-idiotype antibody or antigen-binding portion thereof that specifically binds to KL2B413, comprising a heavy chain variable (VH) domain comprising a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 to 7, a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11 to 14, and a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 19 to 20, and further comprising a light chain variable (VL) domain comprising a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 25 to 26, a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 29 to 30, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 33 to 34. 4. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein the VH domain has an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45 and the VL domain has an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 46. 5. An anti-idiotype antibody or antigen-binding portion according to claim 1, comprising a heavy chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37, and further comprising a light chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 39. 6. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein the VH domain has the amino acid sequence of SEQ ID NO: 45 and the VL domain has the amino acid sequence of SEQ ID NO: 46. 7. The anti-idiotype antibody or antigen-binding portion according to claim 1, comprising a heavy chain comprising the amino acid sequence of SEQ ID NO: 37, and further comprising a light chain comprising the amino acid sequence of SEQ ID NO: 39. 8. The antigen-binding portion is Fab, F(ab') 2 10. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein the anti-idiotype antibody or antigen-binding portion is selected from a IgG1A antibody, a IgG2A antibody, a IgG3A antibody, a IgG4A antibody, a IgG5A antibody, a IgG6A antibody, a IgG7A antibody, a IgG8A antibody, a IgG9A antibody, a IgG1A antibody, a IgG1B antibody, a IgG2C antibody, a IgG4A 9. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein the antibody is a monoclonal antibody. 10. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein the antibody is a chimeric antibody. 11. The anti-idiotype antibody or antigen-binding portion according to claim 10, wherein said antibody comprises a murine IgG2a framework. 12. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein said antibody is a fully human antibody. 13. A nucleic acid encoding the heavy chain, the light chain, or both, of an anti-idiotypic antibody or antigen-binding portion thereof according to 1 above. 14. A nucleic acid encoding the heavy chain, the light chain, or both of an anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to KL2B413, comprising: a) the nucleotide sequence of SEQ ID NO: 38; b) the nucleotide sequence of SEQ ID NO: 40; c) A nucleic acid comprising both a) and b). 15. A vector comprising the nucleic acid described in 14 above. 16. The vector according to claim 15, wherein the vector is an expression vector. 17. A host cell containing the vector described in 16 above. 18. The host cell according to claim 17, wherein the cell is a mammalian cell. 19. A method for producing an anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to KL2B413, comprising culturing the host cell described in 17 above, which contains nucleotide sequences encoding the heavy chain and the light chain of the antibody or antigen-binding portion, under conditions that allow expression of the antibody or antigen-binding portion, and isolating the antibody or antigen-binding portion from the culture. 20. A method for detecting KL2B413 in a biological sample, comprising: (a) preparing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in any one of 1 to 3 above; and (c) detecting the anti-idiotype antibody or antigen-binding portion. 21. A method for detecting the expression of a chimeric antigen receptor (CAR) comprising KL2B413 in a biological sample, the method comprising: (a) preparing a biological sample; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in any one of 1 to 3 above; and (c) detecting the anti-idiotype antibody or antigen-binding portion, thereby detecting the expression of the CAR. 22. The method of claim 20, wherein the antibody comprises a detectable label. 23. The method of claim 20, further comprising contacting the anti-idiotype antibody or the antigen-binding portion with a detectable label prior to detecting the anti-idiotype antibody or the antigen-binding portion. 24. The method according to claim 20, wherein the biological sample is blood, serum or urine. 25. The anti-idiotype antibody or antigen-binding portion according to claim 1, wherein KL2B413 is within the antigen-binding domain of the extracellular portion of a chimeric antigen receptor (CAR). 26. The anti-idiotype antibody or antigen-binding portion according to claim 25, wherein KL2B413 is an scFv and the anti-idiotype antibody or antigen-binding portion specifically binds to an epitope in the scFv of the CAR. 27. The anti-idiotype antibody or antigen-binding portion according to claim 25, wherein KL2B413 binds to KLK2. 28. The anti-idiotypic antibody or antigen-binding portion according to claim 25, wherein said antibody or said antigen-binding portion does not cross-react with other KLK2 antibodies or other KLK2-binding CARs. 29. The anti-idiotype antibody or antigen-binding portion according to claim 25, wherein the CAR has an amino acid sequence selected from the group consisting of SEQ ID NOs: 43 to 44. 30. A kit for detecting KL2B413 in a biological sample, comprising: (a) an anti-idiotype antibody or antigen-binding portion thereof described in 1 above; and (b) instructions for detecting the anti-idiotype antibody or antigen-binding portion thereof. 31. An anti-idiotype antibody or antigen-binding portion thereof according to claim 1, for use in detecting KL2B413 in a biological sample, wherein the detection comprises (a) preparing a biological sample, (b) contacting the biological sample with the anti-idiotype antibody or antigen-binding portion, and (c) detecting the anti-idiotype antibody or antigen-binding portion. 32. A method for purifying KL2B413 from a sample, comprising: (a) preparing a biological sample containing KL2B413; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in 1 above; and (c) capturing the anti-idiotype antibody or antigen-binding portion, thereby purifying KL2B413. 33. A method for selecting CAR-T cells from a cell population, comprising: (a) providing a biological sample containing CAR-T cells; (b) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion described in 1 above; and (c) capturing the anti-idiotype antibody or antigen-binding portion, thereby selecting CAR-T cells.

Claims

1. An anti-idiotype antibody or antigen-binding portion thereof that specifically binds to a target antibody comprising KL2B413, wherein the target antibody or antigen-binding portion thereof comprises a VH domain having the amino acid sequence of SEQ ID NO: 41 and a VL domain having the amino acid sequence of SEQ ID NO:

42.

2. An anti-idiotype antibody or antigen-binding portion thereof that specifically binds to KL2B413, wherein the anti-idiotype antibody or antigen-binding portion thereof comprises a heavy chain variable (VH) domain and a light chain variable (VL) domain, wherein the VH and VL are as follows: (i) a VH CDR1 having an amino acid sequence consisting of SEQ ID NO: 4, a VH CDR2 having an amino acid sequence consisting of SEQ ID NO: 11, a VH CDR3 having an amino acid sequence consisting of SEQ ID NO: 19, a VL CDR1 having an amino acid sequence consisting of SEQ ID NO: 25, a VL CDR2 having an amino acid sequence consisting of SEQ ID NO: 29, and a VL CDR3 having an amino acid sequence consisting of SEQ ID NO: 33; (ii) a VH CDR1 having an amino acid sequence consisting of SEQ ID NO: 5, a VH CDR2 having an amino acid sequence consisting of SEQ ID NO: 12, a VH CDR3 having an amino acid sequence consisting of SEQ ID NO: 19, a VL CDR1 having an amino acid sequence consisting of SEQ ID NO: 25, a VL CDR2 having an amino acid sequence consisting of SEQ ID NO: 29, and a VL CDR3 having an amino acid sequence consisting of SEQ ID NO: 33; (iii) a VH CDR1 having an amino acid sequence consisting of SEQ ID NO: 6, a VH CDR2 having an amino acid sequence consisting of SEQ ID NO: 13, a VH CDR3 having an amino acid sequence consisting of SEQ ID NO: 20, a VL CDR1 having an amino acid sequence consisting of SEQ ID NO: 26, a VL CDR2 having an amino acid sequence consisting of SEQ ID NO: 30, and a VL CDR3 having an amino acid sequence consisting of SEQ ID NO: 34; or (iv) a VH CDR1 having an amino acid sequence consisting of SEQ ID NO: 7, a VH CDR2 having an amino acid sequence consisting of SEQ ID NO: 14, a VH CDR3 having an amino acid sequence consisting of SEQ ID NO: 20, a VL CDR1 having an amino acid sequence consisting of SEQ ID NO: 26, a VL CDR2 having an amino acid sequence consisting of SEQ ID NO: 30, and a VL CDR3 having an amino acid sequence consisting of SEQ ID NO: 34; An anti-idiotype antibody or an antigen-binding portion thereof, comprising:

3. 3. The anti-idiotype antibody or antigen-binding portion of claim 2, wherein the VH domain has an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 45, and the VL domain has an amino acid sequence having at least 90% sequence identity to SEQ ID NO:

46.

4. 3. The anti-idiotype antibody or antigen-binding portion of claim 2, comprising a heavy chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 37, and further comprising a light chain comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO:

39.

5. 2. The anti-idiotype antibody or antigen-binding portion of claim 1, wherein the VH domain has the amino acid sequence of SEQ ID NO: 45 and the VL domain has the amino acid sequence of SEQ ID NO:

46.

6. 2. The anti-idiotype antibody or antigen-binding portion of claim 1, comprising a heavy chain comprising the amino acid sequence of SEQ ID NO: 37 and further comprising a light chain comprising the amino acid sequence of SEQ ID NO:

39.

7. The antigen-binding portion is Fab, F(ab') ２ 2. The anti-idiotype antibody or antigen-binding portion of claim 1, which is selected from: a scFv;

8. The anti-idiotype antibody or antigen-binding portion of claim 1 , wherein the antibody is a monoclonal antibody.

9. The anti-idiotype antibody or antigen-binding portion of claim 1 , wherein the antibody is a chimeric antibody.

10. 10. The anti-idiotype antibody or antigen-binding portion of claim 9, wherein the antibody comprises a murine IgG2a framework.

11. The anti-idiotype antibody or antigen-binding portion of claim 1 , wherein the antibody is a fully human antibody.

12. A nucleic acid encoding the heavy and light chains of the anti-idiotype antibody or antigen-binding portion of claim 1.

13. A nucleic acid encoding the heavy chain and light chain of an anti-idiotype antibody or an antigen-binding portion thereof that specifically binds to KL2B413, a) the nucleotide sequence of SEQ ID NO: 38, and b) the nucleotide sequence of SEQ ID NO: 40; A nucleic acid comprising:

14. A vector comprising the nucleic acid of claim 13.

15. The vector of claim 14 , wherein the vector is an expression vector.

16. A host cell comprising the vector of claim 15.

17. The host cell of claim 16 , wherein the cell is a mammalian cell.

18. A method for producing an anti-idiotypic antibody or antigen-binding portion thereof that specifically binds to KL2B413, comprising culturing the host cell of claim 16, which comprises nucleotide sequences encoding the heavy and light chains of the antibody or antigen-binding portion, under conditions that allow expression of the antibody or antigen-binding portion, and isolating the antibody or antigen-binding portion from the culture.

19. A method for detecting KL2B413 in a biological sample, comprising: (a) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion thereof according to any one of claims 1 to 2; and (b) detecting the anti-idiotype antibody or antigen-binding portion thereof.

20. 1. A method for detecting expression of a chimeric antigen receptor (CAR) comprising KL2B413 in a biological sample, the method comprising: (a) contacting the biological sample with an anti-idiotype antibody or antigen-binding portion thereof according to any one of claims 1 to 2; and (b) detecting the anti-idiotype antibody or antigen-binding portion thereof, thereby detecting the expression of the CAR.

21. 20. The method of claim 19, wherein the antibody comprises a detectable label.

22. 20. The method of claim 19, further comprising contacting the anti-idiotype antibody or the antigen-binding portion with a detectable label prior to detecting the anti-idiotype antibody or the antigen-binding portion.

23. 20. The method of claim 19, wherein the biological sample is blood, serum, or urine.

24. 2. The anti-idiotype antibody or antigen-binding portion of claim 1, wherein KL2B413 is within the antigen-binding domain of the extracellular portion of a chimeric antigen receptor (CAR).

25. 25. The anti-idiotype antibody or antigen-binding portion of claim 24, wherein KL2B413 is an scFv and the anti-idiotype antibody or antigen-binding portion specifically binds to an epitope in the scFv of the CAR.

26. 25. The anti-idiotype antibody or antigen-binding portion of claim 24, wherein KL2B413 binds to KLK2.

27. 25. The anti-idiotype antibody or antigen-binding portion of claim 24, wherein the CAR has an amino acid sequence selected from the group consisting of SEQ ID NOs: 43-44.

28. A kit for detecting KL2B413 in a biological sample, comprising: (a) an anti-idiotype antibody or antigen-binding portion thereof described in claim 1; and (b) instructions for detecting the anti-idiotype antibody or antigen-binding portion thereof.

29. 2. The anti-idiotype antibody or antigen-binding portion of claim 1, for use in detecting KL2B413 in a biological sample, wherein the detection comprises: (a) contacting the biological sample with the anti-idiotype antibody or antigen-binding portion; and (b) detecting the anti-idiotype antibody or antigen-binding portion.

30. A method for purifying KL2B413 from a sample, comprising: (a) contacting a biological sample containing KL2B413 with an anti-idiotype antibody or antigen-binding portion described in claim 1; and (b) capturing the anti-idiotype antibody or antigen-binding portion, thereby purifying KL2B413.

31. 1. A method for selecting CAR-T cells from a cell population, comprising: (a) contacting a biological sample containing CAR-T cells with the anti-idiotype antibody or antigen-binding portion of claim 1; and (b) capturing the anti-idiotype antibody or antigen-binding portion, thereby selecting CAR-T cells.

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