Anti-SLC34A2 monoclonal antibody and its use

Abstract

Antibodies or fragments thereof having binding specificity for human solute carrier 34A2 (SLC34A2) protein are provided. These antibodies can bind to SLC34A2 with high affinity, mediate antibody-dependent cellular cytotoxicity (ADCC), and effectively induce endocytosis. Methods and uses for treating cancer are also provided. In one embodiment, an antibody or fragment thereof having specificity for human SLC34A2 protein is provided. In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region comprising heavy chain complementarity determining regions CDRH1, CDRH2 and CDRH3, and a light chain variable region comprising light chain complementarity determining regions CDRL1, CDRL2 and CDRL3.

Description

Background Art

[0001] Background Cell membrane transporter proteins, such as transporters belonging to the glucose transporter GLUT, ATP-binding cassette transporter ABC, and solute carrier transporter SLC family, are frequently upregulated on the surface of cancer cells compared to adjacent normal cells. High levels of transporters are found in a wide range of solid tumors and are correlated with poor survival. One potential molecular tumor marker could be the sodium-dependent phosphate transporter NaPi2b encoded by the SLC34A2 (solute carrier 34A2) gene.

[0002] SLC34A2 encodes the type II Na / Pi cotransporter (NaPi2b), a multiple transmembrane sodium-dependent phosphate transporter that causes transcellular inorganic phosphate uptake. NaPi2b is very abundant in the brush border membrane of the small intestine, where it is involved in the transcellular flux of inorganic phosphate across the apical membrane of epithelial cells. Altered expression of the sodium-dependent phosphate transporter NaPi2b has been reported in ovarian cancer, lung cancer, gastric cancer, thyroid cancer, and other cancers.

Summary of the Invention

Means for Solving the Problems

[0003] Currently, NaPi2b is the target of therapeutic antibodies XMT-1536 and RG-7599 that are in clinical trials for the treatment of ovarian and lung cancer. There is a need to develop more effective and safe clinical candidates that target this protein.

[0004] Summary Anti-SLC34A2 antibodies have been discovered herein that have high binding affinity for human SLC34A2 protein and are efficient in mediating antibody-dependent cellular cytotoxicity (ADCC) and inducing cytotoxicity. Compared to two benchmark antibodies, XMT-1536 and RG-7599, the antibodies discovered in the present invention showed higher overall binding affinity, much improved cross-reactivity, more potent ADCC and cytotoxicity. Therefore, these antibodies can be suitably used for the treatment of diseases such as cancer.

[0005] In one embodiment, an antibody or a fragment thereof having specificity for human SLC34A2 protein is provided. In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region comprising heavy chain complementarity determining regions CDRH1, CDRH2 and CDRH3, and a light chain variable region comprising light chain complementarity determining regions CDRL1, CDRL2 and CDRL3.

[0006] In some embodiments, a polynucleotide encoding the antibody or fragment, and a composition comprising the antibody or fragment thereof and a pharmaceutically acceptable carrier are also provided.

[0007] Methods and uses for the treatment of diseases and conditions are also provided. In one embodiment, a method for treating cancer in a patient in need of treatment for cancer, the method comprising administering to the patient an antibody or fragment thereof of the present disclosure is provided. BRIEF DESCRIPTION OF THE DRAWINGS

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Mode for Carrying Out the Invention

[0028] Detailed Description Definitions Note that the term "a" or "an" entity refers to one or more of such entities; for example, "an antibody" is understood to represent one or more antibodies. Thus, the terms "a" (or "an"), "one or more", and "at least one" can be used interchangeably herein.

[0029] As used herein, the term "polypeptide" is intended to include both the singular form "polypeptide" and the plural form "polypeptides", and refers to a molecule composed of monomers (amino acids) linearly linked by amide bonds (also known as peptide bonds). The term "polypeptide" refers to any chain (singular or plural) of two or more amino acids and does not refer to a product of a specific length. Thus, peptides, dipeptides, tripeptides, oligopeptides, "proteins", "amino acid chains", or any other term used to refer to a chain (singular or plural) of two or more amino acids are included within the definition of "polypeptide", and the term "polypeptide" can be used in place of or interchangeably with any of these terms. The term "polypeptide" is also intended to refer to the product of post-expression modification of a polypeptide, including, without limitation, glycosylation, acetylation, phosphorylation, amidation, derivatization by known protecting / blocking groups, proteolytic cleavage, or modification by non-naturally occurring amino acids. A polypeptide can be derived from a natural biological source or produced by recombinant techniques, but is not necessarily translated from a specified nucleic acid sequence. It can be produced in any manner, including by chemical synthesis.

[0030] "Homology" or "identity" or "similarity" refers to sequence similarity between two polypeptides or between two nucleic acid molecules. Homology can be determined by comparing the positions in each sequence that can be aligned for comparison purposes. If a position in the sequences being compared is occupied by the same base or amino acid, the molecules are homologous at that position. The degree of homology between sequences is a function of the number of matching positions or homologous positions shared by the sequences. "Unrelated" or "non-homologous" sequences share less than 40% identity, preferably less than 25% identity, with one of the sequences of the present disclosure.

[0031] A polynucleotide or polynucleotide region (or polypeptide or polypeptide region) has a certain percentage (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99%) of "sequence identity" to another sequence means that, when aligned, that percentage of bases (or amino acids) is the same when comparing the two sequences.

[0032] The term "equivalent nucleic acid or polynucleotide" refers to a nucleic acid having a nucleotide sequence with a certain degree of homology or sequence identity to the nucleotide sequence of a nucleic acid or its complement. Homologs of double-stranded nucleic acids are intended to include nucleic acids having a nucleotide sequence with a certain degree of homology to the nucleic acid or its complement. In one embodiment, a homolog of a nucleic acid can hybridize to the nucleic acid or its complement. Similarly, "equivalent polypeptide" refers to a polypeptide having a certain degree of homology or sequence identity to the amino acid sequence of a reference polypeptide. In some embodiments, the sequence identity is at least about 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99%. In some embodiments, an equivalent polypeptide or polynucleotide has 1, 2, 3, 4 or 5 additions, deletions, substitutions and combinations thereof compared to the reference polypeptide or polynucleotide. In some embodiments, an equivalent sequence retains the activity (e.g., epitope binding) or structure (e.g., salt bridge) of the reference sequence.

[0033] As used herein, "antibody" or "antigen-binding polypeptide" refers to a polypeptide or polypeptide complex that specifically recognizes and binds to an antigen. An antibody can be the whole antibody and any of its antigen-binding fragments or single chains. Thus, the term "antibody" includes any protein or peptide containing a molecule that includes at least a portion of an immunoglobulin molecule having the biological activity of binding to an antigen. Examples of such include, but are not limited to, the complementarity determining regions (CDRs) of the heavy or light chains or the ligand-binding portion thereof, the variable regions of the heavy or light chains, the constant regions of the heavy or light chains, the framework (FR) regions, or any portion thereof, or at least one portion of a binding protein.

[0034] The terms "antibody fragment" or "antigen-binding fragment" as used herein are parts of an antibody such as F(ab’)2, F(ab)2, Fab’, Fab, Fv, scFv and the like. Regardless of structure, an antibody fragment binds to the same antigen recognized by the intact antibody. The term "antibody fragment" includes aptamers, spiegelmers and diabodies. The term "antibody fragment" also includes any synthetic or genetically engineered protein that acts like an antibody by binding to a specific antigen to form a complex.

[0035] "Single-chain variable fragment" or "scFv" refers to a fusion protein of the variable region (V H ) of the heavy chain of an immunoglobulin and the variable region (V L ) of the light chain. In some embodiments, the regions are connected by a short linker peptide of 10 to about 25 amino acids. The linker can be rich in glycine for flexibility and rich in serine or threonine for solubility, and can connect the N-terminus of V H to the C-terminus of V L or vice versa. This protein retains the specificity of the original immunoglobulin despite removal of the constant regions and introduction of the linker. scFv molecules are known in the art and are described, for example, in U.S. Patent No. 5,892,019.

[0036] The term antibody encompasses a wide variety of biochemically distinguishable classes of polypeptides. One of ordinary skill in the art will recognize that heavy chains are classified as gamma, mu, alpha, delta, or epsilon (γ, μ, α, δ, ε), and some of these have subclasses (e.g., γ1-γ4). It is the nature of this chain that determines the "class" of the antibody as IgG, IgM, IgA, IgD, or IgE, respectively. Immunoglobulin subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgG5, etc., are well-characterized and are known to confer functional specialization. Modified versions of each of these classes and isotypes are readily distinguishable to one of ordinary skill in the art in view of the present disclosure and are thus within the scope of the present disclosure. All immunoglobulin classes are clearly within the scope of the present disclosure, and the following description will generally be directed to the IgG class of immunoglobulin molecules. With respect to IgG, a standard immunoglobulin molecule comprises two identical light chain polypeptides having a molecular weight of approximately 23,000 daltons, and two identical heavy chain polypeptides having a molecular weight of 53,000-70,000. These four chains are typically linked by disulfide bonds to form a "Y" configuration, in which the light chains flank the heavy chains such that they extend from the mouth of the "Y" to the variable region.

[0037] The antibodies, antigen-binding polypeptides, variants or derivatives of the present disclosure include, but are not limited to, polyclonal, monoclonal, multispecific, human, humanized, primatized or chimeric antibodies, single-chain antibodies, epitope-binding fragments such as Fab, Fab’ and F(ab’)2, Fd, Fv, single-chain Fv (scFv), single-chain antibodies, disulfide-linked Fv (sdFv), fragments containing either the VK or VH domain, fragments produced by a Fab expression library, and anti-idiotype (anti-Id) antibodies (including, for example, anti-Id antibodies against the LIGHT antibodies disclosed herein). The immunoglobulin or antibody molecules of the present disclosure can be of any type (e.g., IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl and IgA2) or subclass of immunoglobulin molecule.

[0038] The light chains are classified as either kappa or lambda (Κ, λ). Each heavy chain class can be associated with either a kappa or lambda light chain. Generally, when an immunoglobulin is produced by any of a hybridoma, B cell or genetically engineered host cell, the light and heavy chains are covalently linked to each other, and the "tail" portions of the two heavy chains are linked to each other by covalent disulfide linkages or non-covalent linkages. In the heavy chain, the amino acid sequence proceeds from the N-terminus at the fork-shaped end of the Y configuration to the C-terminus at the base of each chain.

[0039] Both the light and heavy chains are divided into regions of structural and functional homology. The terms "constant" and "variable" are used functionally. In this regard, it will be appreciated that the variable domains of both the light (VK) and heavy (VH) chain portions determine antigen recognition and specificity. Conversely, the constant domains of the light chain (CK) and heavy chain (CH1, CH2 or CH3) confer important biological properties such as secretion, transplacental mobility, Fc receptor binding, complement binding and the like. By convention, the numbering of the constant region domains increases as one moves more distally from the antigen-binding site or amino terminus of the antibody. The N-terminal portion is the variable region and the C-terminal portion is the constant region; the CH3 and CK domains actually contain the carboxy termini of the heavy and light chains, respectively.

[0040] As indicated above, the variable region enables the antibody to selectively recognize and specifically bind to an epitope on an antigen. That is, the VK and VH domains of the antibody, or a subset of the complementarity-determining regions (CDRs), combine to form the variable region that defines the three-dimensional antigen-binding site. This quaternary antibody structure forms the antigen-binding sites present at the ends of each arm of the Y. More specifically, the antigen-binding site is defined by three CDRs (i.e., CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2 and CDR-L3) in each of the VH and VK chains. In some examples, such as certain immunoglobulin molecules derived from camelid species or engineered based on camelid immunoglobulins, the full-length immunoglobulin molecule can consist of only heavy chains without light chains. See, for example, Hamers-Casterman et al., Nature 363:446-448 (1993).

[0041] In naturally occurring antibodies, the six "complementary determining regions" or "CDRs" present in each antigen-binding domain are short contiguous sequences of amino acids that specifically position themselves to form the antigen-binding domain when the antibody assumes its three-dimensional conformation in an aqueous environment. The remaining amino acids in the antigen-binding domain, referred to as the "framework" regions, exhibit lower intermolecular variability. The framework regions mostly assume the form of a β-sheet conformation, and the CDRs form loops that connect the β-sheet structures and, in some cases, form part of them. Thus, the framework regions act to form a scaffold that provides for the positioning of the CDRs in the correct orientation by means of interchain, non-covalent interactions. The antigen-binding domain formed by the positioned CDRs defines a surface that is complementary to the epitope on the immunoreactive antigen. This complementary surface facilitates the non-covalent binding of the antibody to its cognate epitope. Since the amino acids that make up each of the CDR and framework regions are precisely defined, one of ordinary skill in the art can readily identify any given heavy-chain variable region or light-chain variable region (see "Sequences of Proteins of Immunological Interest," Kabat, E., et al., U.S. Department of Health and Human Services, (1983); and Chothia and Lesk, J. MoI. Biol., 196:901-917 (1987)).

[0042] When there are two or more definitions of a term in use and / or accepted within the relevant art, the definition of the term as used herein is intended to include all such meanings unless the contrary is clearly stated herein. A specific example is the use of the term "complementary determining region" ("CDR") to describe non-contiguous antigen-binding sites found within the variable regions of both heavy and light chain polypeptides. This particular region is described by Kabat et al., U.S. Dept. of Health and Human Services, "Sequences of Proteins of Immunological Interest" (1983) and Chothia et al., J. Mol. Biol. 196:901-917 (1987), which are hereby incorporated by reference in their entirety. The CDR definitions according to Kabat and Chothia include overlapping or subsets of amino acid residues when compared to each other. Nevertheless, the application of any definition referring to the CDR of an antibody or its variant is intended to be within the scope of the terms defined and used herein. The appropriate amino acid residues encompassing the CDR as defined by each of the references cited above are shown in the table below for comparison. The exact residue numbers encompassing a particular CDR will vary depending on the sequence and size of the CDR. One of ordinary skill in the art can routinely determine which residues constitute a particular CDR given the amino acid sequence of the variable region of the antibody.

Table 21

[0043] Kabat et al. also defined a numbering scheme for variable domain sequences that is applicable to any antibody. One of ordinary skill in the art can unambiguously assign this "Kabat numbering" scheme to any variable domain sequence without any reliance on experimental data other than the sequence itself. As used herein, "Kabat numbering" refers to the numbering scheme set forth by Kabat et al., U.S. Dept. of Health and Human Services, "Sequence of Proteins of Immunological Interest" (1983).

[0044] In addition to the above table, the Kabat numbering scheme represents the CDR regions as follows: CDR-H1 begins approximately at amino acid 31 (i.e., approximately 9 residues after the first cysteine residue), contains approximately 5-7 amino acids, and ends at the next tryptophan residue. CDR-H2 begins at the 15th residue after the end of CDR-H1, contains approximately 16-19 amino acids, and ends at the next arginine or lysine residue. CDR-H3 begins at approximately the 33rd amino acid residue after the end of CDR-H2; contains 3-25 amino acids; and ends with the sequence W-G-X-G, where X is any amino acid. CDR-L1 begins approximately at residue 24 (i.e., following the cysteine residue); contains approximately 10-17 residues; and ends at the next tryptophan residue. CDR-L2 begins at approximately the 16th residue after the end of CDR-L1 and contains approximately 7 residues. CDR-L3 begins at approximately the 33rd residue after the end of CDR-L2 (i.e., following the cysteine residue); contains approximately 7-11 residues; and ends with the sequence F or W-G-X-G, where X is any amino acid.

[0045] The antibodies disclosed herein can be derived from any animal origin, including birds and mammals. Preferably, the antibodies are human, mouse, donkey, rabbit, goat, guinea pig, camel, llama, horse or chicken antibodies. In another embodiment, the variable region can be of chondricthoid origin (e.g., shark-derived).

[0046] As used herein, the term "heavy chain constant region" includes an amino acid sequence derived from an immunoglobulin heavy chain. A polypeptide comprising a heavy chain constant region comprises at least one of the following: a CH1 domain, a hinge (e.g., upper, middle and / or lower hinge region) domain, a CH2 domain, a CH3 domain, or variants or fragments thereof. For example, an antigen-binding polypeptide for use in the present disclosure can comprise a polypeptide chain comprising a CH1 domain; a polypeptide chain comprising a CH1 domain, at least a portion of a hinge domain and a CH2 domain; a polypeptide chain comprising a CH1 domain and a CH3 domain; a polypeptide chain comprising a CH1 domain, at least a portion of a hinge domain and a CH3 domain, or a polypeptide chain comprising a CH1 domain, at least a portion of a hinge domain, a CH2 domain and a CH3 domain. In another embodiment, the polypeptide of the present disclosure comprises a polypeptide chain comprising a CH3 domain. Further, an antibody for use in the present disclosure can lack at least a portion of the CH2 domain (e.g., all or part of the CH2 domain). As shown above, one of ordinary skill in the art will understand that the heavy chain constant region can be modified such that the amino acid sequence varies from that of a naturally occurring immunoglobulin molecule.

[0047] The heavy chain constant region of the antibodies disclosed herein can be derived from different immunoglobulin molecules. For example, the heavy chain constant region of a polypeptide can comprise a CH1 domain derived from an IgG l molecule and a hinge region derived from an IgG3 molecule. In another example, the heavy chain constant region is an IgG lIt can include a hinge region that is partially derived from a molecule and partially derived from an IgG3 molecule. In another example, the heavy chain portion can include a chimeric hinge that is partially derived from an IgG l molecule and partially derived from an IgG4 molecule.

[0048] As used herein, the term "light chain constant region" includes an amino acid sequence derived from an antibody light chain. Preferably, the light chain constant region includes at least one of a constant kappa domain or a constant lambda domain.

[0049] A "light chain-heavy chain pair" refers to a collection of a light chain and a heavy chain that can form a dimer via a disulfide bond between the CL domain of the light chain and the CH1 domain of the heavy chain.

[0050] As previously indicated, the subunit structure and three-dimensional configuration of the constant regions of various immunoglobulin classes are well known. As used herein, the term "VH domain" includes the amino-terminal variable domain of an immunoglobulin heavy chain, and the term "CH1 domain" includes the first (most amino-terminal) constant region domain of an immunoglobulin heavy chain. The CH1 domain is adjacent to the VH domain and is amino-terminal to the hinge region of the immunoglobulin heavy chain molecule.

[0051] As used herein, the term "CH2 domain" includes, for example, the portion of the heavy chain molecule that extends from approximately residue 244 to residue 360 of an antibody using a conventional numbering scheme (residues 244-360, Kabat numbering system; and residues 231-340, EU numbering system; see Kabat et al., U.S. Dept. of Health and Human Services, "Sequences of Proteins of Immunological Interest" (1983)). The CH2 domain is unique in that it does not pair closely with another domain. Rather, two N-linked branched carbohydrate chains are inserted between the two CH2 domains of an intact native IgG molecule. It has also been well demonstrated that the CH3 domain extends from the CH2 domain to the C-terminus of the IgG molecule and contains approximately 108 residues.

[0052] As used herein, the term "hinge region" includes the portion of the heavy chain molecule that links the CH1 domain to the CH2 domain. This hinge region contains approximately 25 residues and is flexible, thus allowing the two N-terminal antigen-binding regions to move independently. The hinge region can be subdivided into three distinct domains: the upper, middle, and lower hinge domains (Roux et al., J. Immunol 161:4083 (1998)).

[0053] As used herein, the term "disulfide bond" includes a covalent bond formed between two sulfur atoms. The amino acid cysteine contains a thiol group that can form a disulfide bond or bridge with a second thiol group. In most naturally occurring IgG molecules, the CH1 and CK regions are linked by disulfide bonds, and the two heavy chains are linked by two disulfide bonds at positions corresponding to 239 and 242 (positions 226 or 229, EU numbering system) using the Kabat numbering system.

[0054] As used herein, the term "chimeric antibody" is meant to include any antibody in which the immunoreactive region or site is obtained from or derived from a first species and the constant region (which may be intact, partial or modified according to the present disclosure) is obtained from a second species. In certain embodiments, the target binding region or site is derived from a non-human source (e.g., mouse or primate) and the constant region is human.

[0055] As used herein, "percent humanization" is calculated by determining the number of framework amino acid differences (i.e., non-CDR differences) between the humanized domain and the germline domain, subtracting this number from the total number of amino acids, then dividing this by the total number of amino acids and multiplying by 100.

[0056] "Specifically binds" or "has specificity for" generally means that an antibody binds to an epitope through its antigen-binding domain and that the binding requires a degree of complementarity between the antigen-binding domain and the epitope. By this definition, an antibody is said to "specifically bind" to an epitope if it binds to the epitope through its antigen-binding domain more readily than it binds to a random, unrelated epitope. The term "specificity" is used herein to denote the relative affinity of a particular antibody for a particular epitope. For example, antibody "A" may be considered to have higher specificity for a given epitope than antibody "B", or it may be said that antibody "A" binds to epitope "C" with higher specificity than to related epitope "D".

[0057] As used herein, the term "treating" or "treatment" refers to both therapeutic treatment and prophylactic or preventive measures, with the goal of preventing or slowing down (reducing) an undesired physiological change or disorder, such as the progression of cancer. Beneficial or desired clinical outcomes include, but are not limited to, alleviation of symptoms, diminishment of the extent of a disease, stabilized (i.e., not worsening) disease state, delay or slowing of disease progression, amelioration or palliation of a disease state, and remission (whether partial or complete). "Treatment" can also mean an extension of survival as compared to expected survival if not receiving treatment. Those in need of treatment include those already having a condition or disorder, those having a tendency to have a condition or disorder, or those in whom a condition or disorder is to be prevented.

[0058] As used herein, the terms "subject" or "individual" or "animal" or "patient" or "mammal" mean any subject for which a diagnosis, prognosis or therapy is desired, particularly a mammalian subject. Mammalian subjects include humans, laboratory animals, domestic animals and zoo, sport animals or pet animals such as dogs, cats, guinea pigs, rabbits, rats, mice, horses, cattle, cows, etc.

[0059] As used herein, phrases such as "to a patient in need of treatment" or "a subject in need of treatment" include subjects such as mammalian subjects who would benefit from administration of an antibody or composition of the present disclosure used, for example, for detection, diagnostic procedures and / or treatment.

[0060] Anti-SLC34A2 antibody The present disclosure provides anti-SLC34A2 antibodies and fragments thereof that have high affinity for human SLC34A2 protein, are potent in mediating ADCC and cytotoxicity, and can effectively induce endocytosis.

[0061] In addition, these antibodies were extensively tested compared to two reference antibodies, XMT-1536 and RG-7599, in clinical development. The new antibodies were superior to both reference antibodies in terms of overall binding affinity, cross-reactivity, ADCC, and cytotoxicity. Thus, these antibodies are suitable agents for the treatment of various diseases characterized by overexpressed SLC34A2, such as cancer.

[0062] Accordingly, in accordance with one embodiment of the present disclosure, antibodies and antigen-binding fragments thereof that can bind to SLC34A2 are provided. Examples of antibodies include the mouse antibodies listed in Table 1 (e.g., 30-H9(B8)F9, 62-G10(E4)G8, 63-A11(D5)B4, 91-C9F1, 7-F4C4, 94-B10A8, 94-E9B12, 67-C9G11, 90-G10H7, 11-F6H8, 84-F2D10, 26-C5D2, 7-G10E11, 25-B11B9, 62-C10(B3)D2, 55-E5(B6)D12, 28-G3(C6)B9, 18-G1H4, 16-E5D10, 69-B11D8, 71-F7D4, 78-D1B10, 87-H12B7, 20-H2F1, and 95-H11A1D8), as well as the humanized antibodies in Tables 12-15. Also included are antibodies containing the same CDRs as those described herein. In some embodiments, the disclosed antibodies and fragments include antibodies and fragments that bind to the same epitope as those described herein, as well as antibodies and fragments that compete with those disclosed in the present invention in binding to SLC34A2.

[0063] In accordance with one embodiment of the present disclosure, antibodies or fragments thereof comprising heavy and light chain variable domains having the CDR regions disclosed herein, and biological equivalents thereof are provided.

[0064] In one embodiment, the CDR is the CDR of 30-H9(B8)F9 or its humanized counterpart as exemplified in Table 12A. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 51, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH2 comprises the amino acid sequence of SEQ ID NO: 52 or 57, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH3 comprises the amino acid sequence of SEQ ID NO: 53, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL1 comprises the amino acid sequence of SEQ ID NO: 54, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL2 comprises the amino acid sequence of SEQ ID NO: 55, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; and CDRL3 comprises the amino acid sequence of SEQ ID NO: 56, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof.

[0065] As shown in the experimental examples, CDRH2 (SEQ ID NO: 57) of 30-H9(B8)F9 contains an NG dipeptide that may undergo post-translational modification (PTM). Therefore, an N=>S mutation was created to prevent such PTM, and the mutated CDRH2 is referred to as a PTM risk avoidance CDR. However, the experimental data demonstrated that the PTM risk avoidance version was as effective as the original version.

[0066] In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 51, CDRH2 comprises the amino acid sequence of SEQ ID NO: 52, CDRH3 comprises the amino acid sequence of SEQ ID NO: 53, CDRL1 comprises the amino acid sequence of SEQ ID NO: 54, CDRL2 comprises the amino acid sequence of SEQ ID NO: 55, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 56. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 51, CDRH2 comprises the amino acid sequence of SEQ ID NO: 57, CDRH3 comprises the amino acid sequence of SEQ ID NO: 53, CDRL1 comprises the amino acid sequence of SEQ ID NO: 54, CDRL2 comprises the amino acid sequence of SEQ ID NO: 55, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 56.

[0067] In some embodiments, antibodies and fragments thereof that contain the same CDRs as 30-H9(B8)F9 or its humanized counterpart are also provided. In some embodiments, the disclosed antibodies and fragments include antibodies and fragments that bind to the same epitope as 30-H9(B8)F9 or its humanized counterpart, and antibodies and fragments that compete with any of them in binding to SLC34A2.

[0068] In some embodiments, the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1 (mouse or chimeric) and 58-61 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 1 (mouse or chimeric) and 58-61 (humanized).

[0069] In some embodiments, the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2 (mouse or chimeric) and 62-65 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 2 (mouse or chimeric) and 62-65 (humanized).

[0070] In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 59, and the light chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 62-65. In some embodiments, the heavy chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 58-61, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 62. In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 59, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 62.

[0071] In one embodiment, the CDRs are the CDRs of 62-G10(E4)G8 or its humanized counterpart as exemplified in Table 13A. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 66, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH2 comprises the amino acid sequence of SEQ ID NO: 67 or 72, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH3 comprises the amino acid sequence of SEQ ID NO: 68, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL1 comprises the amino acid sequence of SEQ ID NO: 69, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL2 comprises the amino acid sequence of SEQ ID NO: 70, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL3 comprises the amino acid sequence of SEQ ID NO: 71, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof.

[0072] CDRH2, which is one of the CDRs, also contains the potential PTM site, NG, and thus was mutated to NA as a PTM risk avoidance version. Experimental data demonstrated that the PTM risk avoidance version was as effective as the original version.

[0073] In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 66, CDRH2 comprises the amino acid sequence of SEQ ID NO: 67, CDRH3 comprises the amino acid sequence of SEQ ID NO: 68, CDRL1 comprises the amino acid sequence of SEQ ID NO: 69, CDRL2 comprises the amino acid sequence of SEQ ID NO: 70, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 71. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 66, CDRH2 comprises the amino acid sequence of SEQ ID NO: 72, CDRH3 comprises the amino acid sequence of SEQ ID NO: 68, CDRL1 comprises the amino acid sequence of SEQ ID NO: 69, CDRL2 comprises the amino acid sequence of SEQ ID NO: 70, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 71.

[0074] In some embodiments, antibodies and fragments thereof are also provided that contain the same CDRs as 62-G10(E4)G8 or its humanized counterpart. In some embodiments, the disclosed antibodies and fragments include antibodies and fragments that bind to the same epitope as 62-G10(E4)G8 or its humanized counterpart, and antibodies and fragments that compete with any of them in binding to SLC34A2.

[0075] In some embodiments, the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 (mouse or chimeric) and 73 - 76 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 3 (mouse or chimeric) and 73 - 76 (humanized).

[0076] In some embodiments, the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 4 (mouse or chimeric) and 77 - 80 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 4 (mouse or chimeric) and 77 - 80 (humanized).

[0077] In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 76, and the light chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 77-80. In some embodiments, the heavy chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 73-76, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 79. In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 76, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 79.

[0078] In one embodiment, the CDRs are the CDRs of 63-A11(D5)B4 or its humanized counterpart as illustrated in Table 14A. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 81, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH2 comprises the amino acid sequence of SEQ ID NO: 82 or 87, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH3 comprises the amino acid sequence of SEQ ID NO: 83, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL1 comprises the amino acid sequence of SEQ ID NO: 84, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL2 comprises the amino acid sequence of SEQ ID NO: 85, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL3 comprises the amino acid sequence of SEQ ID NO: 86, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof.

[0079] CDRH2, which is one of the CDRs, also contains the potential PTM site, NG, and thus was mutated to SG as a PTM risk avoidance version. Experimental data demonstrated that the PTM risk avoidance version was as effective as the original version.

[0080] In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 81, CDRH2 comprises the amino acid sequence of SEQ ID NO: 82, CDRH3 comprises the amino acid sequence of SEQ ID NO: 83, CDRL1 comprises the amino acid sequence of SEQ ID NO: 84, CDRL2 comprises the amino acid sequence of SEQ ID NO: 85, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 86. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 81, CDRH2 comprises the amino acid sequence of SEQ ID NO: 87, CDRH3 comprises the amino acid sequence of SEQ ID NO: 83, CDRL1 comprises the amino acid sequence of SEQ ID NO: 84, CDRL2 comprises the amino acid sequence of SEQ ID NO: 85, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 86.

[0081] In some embodiments, antibodies and fragments thereof that contain the same CDRs as 63 - A11(D5)B4 or its humanized counterpart are also provided. In some embodiments, the disclosed antibodies and fragments include antibodies and fragments that bind to the same epitope as 63 - A11(D5)B4 or its humanized counterpart, and antibodies and fragments that compete with any of them in binding to SLC34A2.

[0082] In some embodiments, the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 5 (mouse or chimeric) and SEQ ID NOs: 88 - 91 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 5 (mouse or chimeric) and SEQ ID NOs: 88 - 91 (humanized).

[0083] In some embodiments, the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 6 (mouse or chimeric) and SEQ ID NOs: 92 - 95 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 6 (mouse or chimeric) and SEQ ID NOs: 92 - 95 (humanized).

[0084] In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 90, and the light chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 92-95. In some embodiments, the heavy chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 88-91, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 92. In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 90, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 92.

[0085] In one embodiment, the CDRs are the CDRs of 91-C9F1 or its humanized counterpart as illustrated in Table 15A. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 96, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH2 comprises the amino acid sequence of SEQ ID NO: 97 or 102, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRH3 comprises the amino acid sequence of SEQ ID NO: 98 or 103, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL1 comprises the amino acid sequence of SEQ ID NO: 99, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL2 comprises the amino acid sequence of SEQ ID NO: 100, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof; CDRL3 comprises the amino acid sequence of SEQ ID NO: 101, or a variant thereof having 1, 2, or 3 deletions, additions, substitutions, or combinations thereof.

[0086] Two of the CDRs, CDRH2 and CDRH3, also contain potential PTM sites, NG, and were thus mutated to ND and NA, respectively, as PTM risk avoidance versions. Experimental data demonstrated that the PTM risk avoidance versions were as effective as the original versions.

[0087] In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 96, CDRH2 comprises the amino acid sequence of SEQ ID NO: 97, CDRH3 comprises the amino acid sequence of SEQ ID NO: 98 or 103, CDRL1 comprises the amino acid sequence of SEQ ID NO: 99, CDRL2 comprises the amino acid sequence of SEQ ID NO: 100, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 101. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 96, CDRH2 comprises the amino acid sequence of SEQ ID NO: 97 or 102, CDRH3 comprises the amino acid sequence of SEQ ID NO: 98, CDRL1 comprises the amino acid sequence of SEQ ID NO: 99, CDRL2 comprises the amino acid sequence of SEQ ID NO: 100, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 101. In one embodiment, CDRH1 comprises the amino acid sequence of SEQ ID NO: 96, CDRH2 comprises the amino acid sequence of SEQ ID NO: 97, CDRH3 comprises the amino acid sequence of SEQ ID NO: 98, CDRL1 comprises the amino acid sequence of SEQ ID NO: 99, CDRL2 comprises the amino acid sequence of SEQ ID NO: 100, and CDRL3 comprises the amino acid sequence of SEQ ID NO: 101.

[0088] In some embodiments, antibodies and fragments thereof that contain the same CDRs as 91-C9F1 or its humanized counterpart are also provided. In some embodiments, the disclosed antibodies and fragments include antibodies and fragments that bind to the same epitope as 91-C9F1 or its humanized counterpart, and antibodies and fragments that compete with any of them in binding to SLC34A2.

[0089] In some embodiments, the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 7 (mouse or chimeric) and 104-107 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 7 (mouse or chimeric) and 104-107 (humanized).

[0090] In some embodiments, the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 8 (mouse or chimeric) and 108-111 (humanized), or a peptide having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 8 (mouse or chimeric) and 108-111 (humanized).

[0091] In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 105, and the light chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 108-111. In some embodiments, the heavy chain variable region comprises any one of the amino acid sequences of SEQ ID NOs: 104-107, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 110. In some embodiments, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 105, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 110.

[0092] It is recognized that the CDRs can be modified to include CDRs having 1, 2, or 3 amino acid additions, deletions, and / or substitutions. In some embodiments, the substitutions can be conservative substitutions.

[0093] "Conservative amino acid substitutions" are substitutions where an amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains are defined in the art and include basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, nonessential amino acid residues in an immunoglobulin polypeptide are preferably replaced with another amino acid residue from the same side chain family. In another embodiment, a string of amino acids can be replaced with a structurally similar string where the order and / or composition of the side chain family members is different.

[0094] Non-limiting examples of conservative amino acid substitutions are provided in the table below, where a similarity score of 0 or higher indicates a conservative substitution between two amino acids. Table A. Amino Acid Similarity Matrix [Table A] Table B. Conservative Amino Acid Substitutions [Table B-1] [Table B-2]

[0095] One skilled in the art will also understand that the antibodies disclosed herein can be modified such that the amino acid sequence varies from the naturally occurring binding polypeptide from which it is derived. For example, a polypeptide or amino acid sequence derived from a specified protein can be the same as the starting sequence, for example, it can have a certain percent identity to the starting sequence, for example, it can be 60%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to the starting sequence.

[0096] In certain embodiments, the antibody comprises an amino acid sequence or one or more moieties that do not normally associate with the antibody. Exemplary modifications are described in more detail below. For example, the antibodies of the present disclosure can comprise a flexible linker sequence or can be modified to add a functional moiety (e.g., PEG, drug, toxin or label).

[0097] The antibodies, variants or derivatives of the present disclosure include derivatives modified by covalent attachment of any type of molecule to the antibody such that the covalent attachment does not prevent the antibody from binding to the epitope. For example, by way of non-limiting example, the antibody can be modified by, for example, glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization with known protecting / blocking groups, proteolytic cleavage, conjugation to a cell ligand or other protein, etc. Any of a number of chemical modifications, including but not limited to specific chemical cleavage, acetylation, formylation, metabolic synthesis of tunicamycin, etc., can be carried out by known techniques. Moreover, the antibody can contain one or more non-classical amino acids.

[0098] An antibody can be detectably labeled by coupling it to a chemiluminescent compound. Next, the presence of the chemiluminescently tagged antigen-binding polypeptide is determined by detecting the presence of luminescence that occurs during the course of a chemical reaction. Examples of particularly useful chemiluminescent labeling compounds are luminol, isoluminol, theromatic acridinium esters, imidazole, acridinium salts, and oxalate esters.

[0099] An antibody is, 152It is also possible to detectably label using a fluorescent-emitting metal such as Eu or other lanthanide series equivalents. These metals can be attached to the antibody using a metal chelating group such as diethylenetriaminepentacetic acid (DTPA) or ethylenediaminetetraacetic acid (EDTA). Techniques for conjugating various moieties to antibodies are well known, see for example, Arnon et al., "Monoclonal Antibodies For Immunotargeting Of Drugs In Cancer Therapy", in Monoclonal Antibodies And Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss, Inc. (1985); Hellstrom et al., "Antibodies For Drug Delivery", in Controlled Drug Delivery (2nd Ed.), Robinson et al., (eds.), Marcel Dekker, Inc., pp. 623- 53 (1987); Thorpe, "Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review", in Monoclonal Antibodies '84: Biological And Clinical Applications, Pinchera et al. (eds.), pp. 475-506 (1985); "Analysis, Results, And Future Prospective Of The Therapeutic Use Of Radiolabeled Antibody In Cancer Therapy", in Monoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al. (eds.), and Academic Press pp. 303-16 (1985).

[0100] Bifunctional Molecules and Combinatorial Therapies SLC34A2 is overexpressed in tumor cells. As a tumor antigen targeting molecule, an antibody or antigen-binding fragment specific for SLC34A2 can be combined with a second antigen-binding fragment specific for immune cells or an antigen-binding fragment specific for an immune checkpoint to generate a combinatorial therapy or a bispecific antibody.

[0101] In some embodiments, the immune cells are selected from the group consisting of T cells, B cells, monocytes, macrophages, neutrophils, dendritic cells, phagocytes, natural killer cells, eosinophils, basophils, and mast cells. Molecules on immune cells that can be targeted include, for example, CCL1, CD3, CD16, CD19, CD28, and CD64. Other examples include PD-1, PD-L1, CTLA-4, LAG-3 (also known as CD223), CD28, CD122, 4-1BB (also known as CD137), TIM3, OX-40 or OX40L, CD40 or CD40L, LIGHT, ICOS / ICOSL, GITR / GITRL, TIGIT, CD27, VISTA, B7H3, B7H4, HEVM or BTLA (also known as CD272), killer cell immunoglobulin-like receptors (KIR), and CD47.

[0102] Different formats of bispecific antibodies are also provided. In some embodiments, each of the anti-SLC34A2 fragment and the second fragment is independently selected from a Fab fragment, a single-chain variable fragment (scFv), or a single-domain antibody. In some embodiments, the bispecific antibody further comprises an Fc fragment.

[0103] Also provided are bifunctional molecules that do not simply comprise an antibody or antigen-binding fragment. As the tumor antigen targeting molecule, an antibody or antigen-binding fragment specific for SLC34A2, such as those described herein, can be combined with an immune cytokine or ligand via a peptide linker, if desired. The immune cytokine or ligand to be linked includes, but is not limited to, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-10, IL-12, IL-13, IL-15, GM-CSF, TNF-α, CD40L, OX40L, CD27L, CD30L, 4-1BBL, LIGHT, and GITRL. Such bifunctional molecules can combine the immune checkpoint blockade effect with local tumor site immune modulation.

[0104] Polynucleotide encoding an antibody and method for preparing an antibody The present disclosure also provides an isolated polynucleotide or nucleic acid molecule encoding an antibody, variant, or derivative thereof of the present disclosure. The polynucleotide of the present disclosure can encode the entire heavy and light chain variable regions of an antigen-binding polypeptide, variant, or derivative thereof on the same polynucleotide molecule or separate polynucleotide molecules. Moreover, the polynucleotide of the present disclosure can encode portions of the heavy and light chain variable regions of an antigen-binding polypeptide, variant, or derivative thereof on the same polynucleotide molecule or separate polynucleotide molecules.

[0105] Methods of making antibodies are well known in the art and are described herein. In certain embodiments, both the variable and constant regions of the antigen-binding polypeptides of the present disclosure are fully human. Fully human antibodies are described in the art and can be made using the techniques described herein. For example, fully human antibodies against specific antigens can be prepared by administering the antigen to a transgenic animal that has been modified to produce such antibodies in response to antigen loading, but whose endogenous locus has been inactivated. Exemplary techniques that can be used to make such antibodies are described in U.S. Patent Nos. 6,150,584; 6,458,592; and 6,420,140, which are hereby incorporated by reference in their entirety.

[0106] Treatment methods As described herein, the antibodies, variants or derivatives of the present disclosure can be used in certain treatment and diagnostic methods.

[0107] The present disclosure further targets antibody-based therapies involving administering an antibody of the present disclosure to a patient such as an animal, mammal, and human to treat one or more of the disorders or conditions described herein. The therapeutic compounds of the present disclosure include, but are not limited to, the antibodies of the present disclosure (including variants and derivatives thereof described herein), and nucleic acids or polynucleotides encoding the antibodies of the present disclosure (including variants and derivatives thereof described herein).

[0108] The antibodies of the present disclosure can also be used to treat or inhibit cancer. In some embodiments, cancer cells in a patient express or overexpress SLC34A2. As presented above, SLC34A2 can be overexpressed in tumor cells, particularly in gastric, pancreatic, esophageal, ovarian, and lung tumors. Inhibition of SLC34A2 has been shown to be useful for the treatment of tumors.

[0109] Accordingly, in some embodiments, a method for treating cancer in a patient in need thereof is provided. The method, in one embodiment, requires administering to the patient an effective amount of an antibody of the present disclosure. In some embodiments, at least one of the cancer cells (e.g., stromal cells) in the patient overexpresses SLC34A2.

[0110] Cell therapies such as chimeric antigen receptor (CAR) T cell therapy are also provided in the present disclosure. Suitable cells that are contacted with (or alternatively engineered to express) the anti-SLC34A2 antibody of the present disclosure can be used. In some embodiments, the antibody is presented in a chimeric antigen receptor (CAR). Upon such contact or engineering, the cells can then be introduced into a cancer patient in need of treatment. The cancer patient can have any of the types of cancer disclosed herein. The cells (e.g., T cells) can be, for example, without limitation, tumor-infiltrating T lymphocytes, CD4+ T cells, CD8+ T cells, or combinations thereof.

[0111] In some embodiments, the cells are isolated from the cancer patient itself. In some embodiments, the cells are provided by a donor or from a cell bank. When the cells are isolated from the cancer patient, unwanted immune reactions can be minimized.

[0112] Non-limiting examples of cancer include bladder cancer, breast cancer, colorectal cancer, endometrial cancer, esophageal cancer, head and neck cancer, kidney cancer, leukemia, liver cancer, lung cancer, lymphoma, melanoma, pancreatic cancer, prostate cancer, and thyroid cancer. In some embodiments, the cancer is one or more of stomach, pancreatic, esophageal, ovarian, and lung cancers.

[0113] Additional diseases or conditions associated with increased cell survival that can be treated, prevented, diagnosed, and / or prognosed using the antibodies, variants, or derivatives thereof of the present disclosure include malignant diseases and related disorders, such as leukemia (including acute leukemia (e.g., acute lymphoblastic leukemia, acute myelogenous leukemia (including myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia)) and chronic leukemia (e.g., chronic myelogenous (granulocytic) leukemia and chronic lymphocytic leukemia)), polycythemia vera, lymphoma (e.g., Hodgkin's disease and non-Hodgkin's disease), multiple myeloma, Waldenström's macroglobulinemia, heavy chain disease, and solid tumors including but not limited to sarcoma and carcinoma, such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon cancer, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, cholangiocarcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, cervical cancer, testicular tumor, lung cancer, small cell lung cancer, bladder cancer, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma, melanoma, neuroblastoma, and retinoblastoma, including but not limited to progression and / or metastasis thereof.

[0114] The specific dosage and treatment regimen for any particular patient will depend on a variety of factors including the specific antibody, its variant or derivative being used, the patient's age, weight, general health, gender and diet, as well as the time of administration, rate of excretion, drug combination, and the severity of the particular disease being treated. The determination of such factors by a healthcare provider is within the skill of those in the art. The amount will also depend on the individual patient to be treated, the route of administration, the type of formulation, the characteristics of the compound being used, the severity of the disease, and the desired effect. The amount to be used can be determined by pharmacological and pharmacokinetic principles well known in the art.

[0115] The methods of administration of the antibody or variant include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural and oral routes. The antigen-binding polypeptide or composition can be administered by any convenient route, for example, by infusion or bolus injection, by absorption through epithelial or mucosal skin layers (e.g., oral mucosa, rectal and intestinal mucosa, etc.), and can be administered together with other biologically active agents. Thus, the pharmaceutical composition containing the antigen-binding polypeptide of the present disclosure can be administered orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powder, ointment, droplet or transdermal patch), bucally, or as an oral spray or nasal spray.

[0116] The term "parenteral" as used herein refers to a mode of administration including, but not limited to, intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous and intra-articular injection and infusion.

[0117] Administration can be systemic or local. Additionally, it may be desirable to introduce the antibodies of the present disclosure into the central nervous system by any suitable route including intracerebroventricular and intrathecal injection; intracerebroventricular injection can be facilitated, for example, by an intracerebroventricular catheter attached to a reservoir such as an Ommaya reservoir. For example, pulmonary administration can also be used by use of an inhaler or nebulizer and formulation with an aerosolizing agent.

[0118] It may be desirable to administer the antigen-binding polypeptide or composition of the present disclosure locally to the area in need of treatment; this can be achieved, by way of example and not limitation, during surgery by local injection, for example, by topical application in combination with a wound dressing after surgery, by injection, using a catheter, using a suppository, or using an implant, said implant being of a porous, non-porous or gelatinous material comprising a membrane, for example, a sialastic membrane or fibers. Preferably, when administering a protein comprising an antibody of the present disclosure, care should be taken to use a material that does not absorb the protein.

[0119] The amount of the antibody of the present disclosure that will be effective in the treatment, inhibition and prevention of immune or malignant diseases, disorders or conditions can be determined by standard clinical techniques. Additionally, in vitro assays can be used, if necessary, to aid in the identification of the optimal dosage range. The exact dosage to be used in the formulation will also depend on the route of administration, and the severity of the disease, disorder or condition, and should be decided according to the judgment of the practitioner and the circumstances of each patient. The effective dosage may be extrapolated from a dosage-response curve derived from in vitro or animal model test systems.

[0120] As a general proposal, the dosage of the antigen-binding polypeptide of the present disclosure administered to a patient is typically between 0.1 mg and 100 mg per kg of the patient's body weight, between 0.1 mg and 20 mg per kg of the patient's body weight, or between 1 mg and 10 mg per kg of the patient's body weight. Generally, human antibodies have a longer half-life in the human body than antibodies from other species due to the immune response to foreign polypeptides. Thus, lower dosages of human antibodies and less frequent administrations are often possible. Furthermore, the dosage and frequency of administration of the antibodies of the present disclosure can be reduced, for example, by enhancing the uptake and tissue penetration (e.g., into the brain) of the antibody by modifications such as lipidation.

[0121] In additional embodiments, the compositions of the present disclosure are administered in combination with a cytokine. Cytokines that can be administered with the compositions of the present disclosure include, but are not limited to, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-10, IL-12, IL-13, IL-15, anti-CD40, CD40L, and TNF-α.

[0122] In additional embodiments, the compositions of the present disclosure are administered in combination with other therapeutic or prophylactic regimens, such as, for example, radiation therapy.

[0123] Diagnostic methods Overexpression of SLC34A2 has been observed in certain tumor samples, and patients having SLC34A2-overexpressing cells may be responsive to treatment with the anti-SLC34A2 antibodies of the present disclosure. Thus, the antibodies of the present disclosure can also be used for diagnostic and prognostic purposes.

[0124] Preferably, the sample containing cells can be obtained from a patient who can be a cancer patient or a patient who desires a diagnosis. The cells can be cells from tumor tissue or tumor blocks, blood samples, urine samples, or any sample derived from the patient. During the pre-treatment of the sample as required, the sample can be incubated with the antibody of the present disclosure under conditions that allow the antibody to interact with the SLC34A2 protein potentially present in the sample. To detect the presence of the SLC34A2 protein in the sample, methods such as ELISA can be used by utilizing anti-SLC34A2 antibodies.

[0125] The presence (optionally by amount or concentration) of the SLC34A2 protein in the sample can be used for cancer diagnosis, as an indicator of a patient's suitability for treatment with the antibody, or as an indicator of whether a patient has responded (or not responded) to cancer treatment. For prognostic prediction methods, the detection can be performed once, twice, or more often, at a certain stage, at the start of cancer treatment to indicate the progress of the treatment.

[0126] Composition The present disclosure also provides a pharmaceutical composition. Such a composition contains an effective amount of the antibody and an acceptable carrier. In some embodiments, the composition further contains a second anti-cancer agent (e.g., an immune checkpoint inhibitor).

[0127] In a specific embodiment, the term "pharmaceutically acceptable" means approved by a federal or state government regulatory authority or listed in the United States Pharmacopeia or other generally recognized pharmacopeias for use in animals, more particularly in humans. Further, a "pharmaceutically acceptable carrier" will generally be any type of non-toxic solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary.

[0128] The term "carrier" refers to a diluent, adjuvant, additive, or vehicle with which a therapeutic agent is administered. Such pharmaceutical carriers can 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 and the like. Water is a preferred carrier when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be used especially as liquid carriers for injectable solutions. Suitable pharmaceutical additives include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene glycol, water, ethanol and the like. If desired, the composition can also contain minor amounts of wetting or emulsifying agents or pH buffering agents such as, for example, acetates, citrates or phosphates. Antibacterial agents such as benzyl alcohol or methylparaben; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; and agents for the adjustment of tonicity such as sodium chloride or dextrose are also contemplated. These compositions can take the form of solutions, suspensions, emulsions, tablets, pills, capsules, powders, sustained release formulations and other forms. The compositions can be formulated as suppositories with traditional binders and carriers such as triglycerides. Oral formulations can contain standard carriers such as, for example, pharmaceutical grade mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate and the like. Examples of suitable pharmaceutical carriers are described in Remington's Pharmaceutical Sciences by E. W. Martin which is incorporated herein by reference. Such compositions will preferably contain a therapeutically effective amount of the antigen-binding polypeptide in a purified form, together with a suitable amount of carrier to provide a form for proper administration to a patient. The formulations should be suitable for the mode of administration. Parenteral preparations can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.

[0129] In certain embodiments, the composition is formulated as a pharmaceutical composition adapted for intravenous administration to humans according to conventional procedures. Typically, compositions for intravenous administration are solutions in sterile isotonic aqueous buffer. If desired, the composition can also contain solubilizing agents and local anesthetics, such as lidocaine, to relieve pain at the site of injection. Generally, the components are supplied either separately or mixed together as a dry lyophilized powder or water-free concentrate in a unit dosage form, e.g., in a sealed container such as an ampoule or sachet indicating the content of the active agent. If the composition is to be administered by infusion, it can be dispensed by an infusion bottle containing sterile pharmaceutical grade water or saline. If the composition is administered by injection, an ampoule of sterile water for injection or saline may be provided so that the components can be mixed prior to administration.

[0130] The compounds of the present disclosure can be formulated as the neutral or salt forms. Pharmaceutically acceptable salts include salts formed by anions such as those derived from hydrochloric acid, phosphoric acid, acetic acid, oxalic acid, tartaric acid, etc., and salts formed by cations such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxide, isopropylamine, triethylamine, 2-ethylaminoethanol, histidine, procaine, etc.

Examples

[0131] (Example 1) Generation of Mouse Monoclonal Antibodies Against Human SLC34A2 Human SLC34A2 protein was used to immunize different strains of mice, resulting in the generation of hybridomas. SLC34A2 positive binders were selected and subcloned. Thereafter, in vitro binding and functional screening were performed to identify lead antibodies with the highest binding affinity and the strongest functional efficacy.

[0132] The VH / VL sequences of 25 lead mouse antibodies are provided in Table 1 below. Table 1. VH / VL Sequences of Lead Mouse Antibodies [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4]

[0133] (Example 2) FACS Binding of Chimeric Ab to Human SLC34A2 In this example, cell-based binding of a chimeric antibody (hIgG1) prepared from a mouse antibody in human SLC34A2-expressing cells was evaluated using flow cytometry.

[0134] Briefly, OVCAR3 cells or HEK293 cells expressing human SLC34A2 were incubated with the chimeric antibody (from 200 nM, 4-fold dilution, 8 points). After incubation for 60 minutes at 4°C, the cells were washed twice with FACS buffer and then stained with a fluorescent-conjugated secondary antibody for 30 minutes at 4°C. Thereafter, the cells were washed twice and analyzed by flow cytometry.

[0135] The research results (Figures 1A - 1B and Figures 2A - 2B and Tables 2 - 3) showed that all chimeric antibodies, except for a slight decline of chimeric 62-C10(B3)D2 and 95-H11A1D8 at the highest concentration, could bind to human SLC34A2 with high affinity compared to the reference antibody XMT-1536. Table 2. Binding Affinity of Chimeric Antibodies to OVCAR3 Cells [Table 2] Table 3. Binding Affinity of Chimeric Antibodies to HEK293 Cells Expressing Human SLC34A2

Table 3-1

Table 3-2

[0136] Based on the results of the cell-based binding described above, some of the chimeric antibodies were selected for further cell-based binding in human SLC34A2-expressing cells using flow cytometry. RMG-1 cells were incubated with the selected chimeric antibodies (from 200 nM, 4-fold dilutions, 8 points). After incubation for 60 minutes at 4°C, the cells were washed twice with FACS buffer and then stained with a fluorescent-conjugated secondary antibody for 30 minutes at 4°C. Subsequently, the cells were washed twice and analyzed by flow cytometry.

[0137] The research results (Figure 3 and Table 4) indicate that the chimeric antibodies can bind to human SLC34A2 with high affinity compared to the reference antibody XMT-1536. Table 4. Binding affinity of the selected chimeric antibodies for RMG-1 cells

Table 4-1

Table 4-2

[0138] (Example 3) Cross-reactive binding of chimeric Abs to cynomolgus and rhesus SLC34A2 proteins In this example, cell-based binding of chimeric Abs in cynomolgus and rhesus SLC34A2-expressing cells was evaluated using flow cytometry.

[0139] Briefly, cynomolgus monkey SLC34A2-expressing HEK293 engineered cells (HEK293 / cynomolgus monkey_SLC34A2) were incubated with chimeric Ab (from 200 nM, 4-fold dilution, 8 points). After incubation for 60 minutes at 4°C, the cells were washed twice with FACS buffer and then stained with a fluorescent-conjugated secondary antibody at 4°C for 30 minutes. Thereafter, the cells were washed twice and analyzed by flow cytometry.

[0140] The research results (Figure 4 and Table 5) indicate that most of the chimeric Ab can bind to cynomolgus monkey SLC34A2 with high affinity. Table 5. Cross-reactive binding to cynomolgus monkey

Table 5

[0141] Similarly, the binding of the chimeric antibody to cynomolgus monkey SLC34A2 was measured, and the results are shown in Figure 5 and Table 6. The results indicate that most of the chimeric Ab can bind to cynomolgus monkey SLC34A2 with high affinity. Table 6. Cross-reactive binding to cynomolgus monkey

Table 6

[0142] (Example 4) Cross-reactive binding to rat SLC34A2 protein In this example, cell-based binding of chimeric Ab in rat SLC34A2-expressing cells was evaluated using flow cytometry.

[0143] Briefly, rat and mouse SLC34A2-expressing HEK293 engineered cells (HEK293 / rat_SLC34A2, HEK293 / mouse_SLC34A2) were incubated with chimeric Ab (10 ug / ml). After incubation for 60 minutes at 4°C, the cells were washed twice with FACS buffer and then stained with a fluorescent-conjugated secondary antibody for 30 minutes at 4°C. The cells were then washed twice and analyzed by flow cytometry.

[0144] The study results (Figures 6A - 6B) show that most of the chimeric Ab can bind to rodent SLC34A2 with high affinity.

[0145] (Example 5) ADCC against human SLC34A2-expressing cells This example evaluated the ability of chimeric antibodies to induce antibody-dependent cellular cytotoxicity activity.

[0146] The ADCC reporter bioassay used herein employs an alternative readout at an earlier time point in the activation of the ADCC MOA pathway: activation of gene transcription via the NFAT (nuclear factor of activated T cells) pathway in effector cells. Additionally, the ADCC reporter bioassay uses engineered Jurkat cells that stably express the FcγRIIIa receptor, the V158 (high affinity) variant, and an NFAT response element that drives the expression of firefly luciferase as effector cells. The antibody bioactivity in the ADCC MOA is quantified by luciferase produced as a result of NFAT pathway activation; luciferase activity in effector cells is quantified by luminescence readout. The signal is high and the assay background is low.

[0147] Serial dilutions of chimeric Ab were incubated with engineered Jurkat effector cells (ADCC bioassay effector cells) and ADCC bioassay target cells (expressing SLC34A2) for 6 hours at 37°C. Luciferase activity was quantified using ONE-Glo™ Luciferase Assay Buffer Reagent. The results are shown in FIGS. 7A - B and Table 7. As shown, all of the chimeric antibodies tested demonstrated potent ADCC-inducing activity. Table 7. ADCC of Chimeric Antibodies [Table 7-1] [Table 7-2]

[0148] (Example 6) Cytotoxic Effects on Human SLC34A2-Expressing Cells This example tested the cytotoxic effects of chimeric antibodies.

[0149] Human SLC34A2-expressing HEK293 engineered cells (HEK293 / H_SLC34A2) were seeded in 96-well plates. The cells were treated with chimeric Ab complexes at each concentration for 5 days. Cell viability was measured using CellTiter-Glo reagent. Luciferase activity was detected using Envison.

[0150] The results (FIGS. 8A - B and Table 8) indicate that all of these chimeric Abs have highly potent killing activity. Table 8. Cytotoxic Activity [Table 8]

[0151] (Example 7) Mutations to CDRs to Prevent Post-Translation Modifications Six of the mouse antibodies, 90G10H7, 55E5B6D12, 30H9B8F9, 62G10E4G8, 63A11D5B4, and 91C9F1, were selected for humanization and amino acid mutations to prevent potential post-translational modifications (PTMs), which are hereby referred to as PTM risk avoidance sequences.

[0152] The PTM risk avoidance VH chains were first tested for their impact on target binding. Cell-based binding of the PTM Abs in human SLC34A2-expressing cells was evaluated using flow cytometry. Briefly, HEK293 / H_SLC34A2 cells were incubated with the PTM Ab (from 200 nM, 4-fold dilution, 8 points). After incubation for 60 minutes at 4°C, the cells were washed twice with FACS buffer and then stained with a fluorescent-conjugated secondary antibody for 30 minutes at 4°C. Subsequently, the cells were washed twice and analyzed by flow cytometry.

[0153] The study results (Figures 9A - C and Table 9) indicate that the humanized / PTM risk avoidance antibodies can bind to human SLC34A2 with high affinity. Table 9. Binding to SLC34A2-expressing HEK293

Table 9-1

Table 9-2

[0154] Similarly, the binding of these humanized / PTM risk avoidance antibodies to OVCAR3 cells that endogenously express the protein was tested. The results are shown in Figure 10 and Table 10. Also, the results confirmed the binding affinity of these antibodies. Table 10. Binding to OVCAR3 cells

Table 10

[0155] The binding of these antibodies to rhesus SLC34A2 was also tested using the procedures described above. The results are shown in Figure 11 and Table 11, confirming the high affinity of these PTM risk avoidance sequences. Table 11. Binding to rhesus SLC34A2

Table 11-1

Table 11-2

[0156] (Example 8) Humanized antibody Based on the above results, four of the above antibodies, 30H9B8F9, 62G10E4G8, 63A11D5B4 and 91C9F1, were continued to be developed by humanization.

[0157] The amino acid sequences of the VH and VL of each mouse antibody were compared against a database of available human Ig gene sequences to find the human germline Ig gene sequences that best matched overall. Next, the CDRs of the mouse antibody were grafted onto the matched human sequences. cDNA was synthesized and used for the production of humanized antibodies. Next, certain back mutations from the mouse antibody were introduced back into the humanized antibody.

[0158] The amino acid sequences of the humanized antibodies are provided in Tables 12-15 below. Humanized and PTM risk avoidance sequences A. 30-H9(B8)F9 Table 12A. Humanized sequences

Table 12A-1

Table 12A-2

Table 12B

Table 15A

Table 15B-1

Table 15B-2

Table 15C

[0159] The binding of these humanized antibodies to OVCAR3 cells was evaluated as described above, and the results are shown in Figures 12A - D and Tables 16A - D. Table 16A. Binding of humanized antibodies to OVCAR3 - 30H9B8F9

Table 16A

Table 16B-1

Table 16B-2

Table 16C

Table 16D

[0160] In another experiment, cell-based binding of hu Abs in human SLC34A2-expressing cells was evaluated using flow cytometry. Briefly, HEK293 / H_SLC34A2, OVCAR3 and RMG-1 cells were incubated with hu Abs. After incubation for 60 minutes at 4°C, the cells were washed twice with FACS buffer and then stained with a fluorescent-conjugated secondary antibody for 30 minutes at 4°C. The cells were then washed twice and analyzed by flow cytometry.

[0161] The results of the study (Figures 13A-D and Tables 17A-D) indicate that the hu Abs are capable of binding to human SLC34A2 with high affinity. Table 17A. Binding to cell lines - 30H9B8F9 [Table 17A] Table 17B. Binding to cell lines - 62G10E4G8 [Table 17B] Table 17C. Binding to cell lines - 63A11D5B4 [Table 17C] Table 17D. Binding to cell lines - 91C9F1 [Table 17D]

[0162] In a further assay, these antibodies were evaluated for their binding affinity to cynomolgus / rat / mouse SLC34A2 using the method described above. As shown in Figures 14-16 and Table 18, these humanized antibodies retained high affinity. Table 18. Binding to cynomolgus SLC34A2 [Table 18]

[0163] (Example 8) ADCC of Humanized Antibody In this example, the antibody-dependent cellular cytotoxicity (ADCC) of a humanized antibody was measured.

[0164] The ADCC reporter bioassay uses an alternative readout at an earlier time point in the activation of the ADCC MOA pathway: activation of gene transcription via the NFAT (nuclear factor of activated T cells) pathway in effector cells. In addition, the ADCC reporter bioassay uses engineered Jurkat cells that stably express the FcγRIIIa receptor, the V158 (high-affinity) variant, and the NFAT response element that drives the expression of firefly luciferase as effector cells. The antibody bioactivity in the ADCC MOA is quantified by the luciferase produced as a result of NFAT pathway activation; the luciferase activity in the effector cells is quantified by a luminescence readout. The signal is high and the assay background is low.

[0165] Serial dilutions of the hu Ab were incubated with engineered Jurkat effector cells (ADCC bioassay effector cells) and ADCC bioassay target cells (expressing SLC34A2) for 6 hours of induction at 37°C. Luciferase activity was quantified using the ONE-Glo™ luciferase assay buffer reagent.

[0166] The results are shown in Figure 17 and Table 19, confirming the potent ADCC-inducing activity of these humanized antibodies. Table 19. ADCC Activity [Table 19]

[0167] (Example 9) Testing of Selected Humanized Antibodies Based on previous results, four humanized antibodies, including 30-H9(B8)F9.p1.z5, 62-G10(E4)G8.p3.z15, 63-A11(D5)B4.p1.z9 and 91-C9F1.p1.z7, were selected for further development.

[0168] The candidate antibodies were further tested in comparison with benchmark antibodies, XMT-1536 and RG-7599. Figure 18 shows a comparison of binding to human SLC34A2 in different cell types. Figure 19 compares the antibodies for their cross-reactivity to SLC34A2 proteins from different species. Figures 20A - B compare their ADCC-inducing activities.

[0169] The above results are summarized in Table 20, which shows that the antibodies developed in the present invention were significantly superior to the benchmarks (XMT-1536 and RG-7599) in many aspects. Table 20. Summary of Comparison

Table 20

[0170] This disclosure should not be limited to the specific embodiments described, which are intended as one illustration of the individual aspects of this disclosure, but any compositions or methods that are functionally equivalent are within the scope of this disclosure. It will be apparent to those skilled in the art that various modifications and variations can be made to the methods and compositions of this disclosure without departing from the spirit or scope of the disclosure. Thus, this disclosure is intended to cover modifications and variations of this disclosure provided they fall within the scope of the appended claims and their equivalents.

[0171] All publications and patent applications mentioned herein are hereby incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

Claims

1. An antibody or fragment having binding specificity to the human solute transporter 34A2 (SLC34A2) protein, wherein the antibody or fragment comprises a heavy chain variable region including heavy chain complementarity determining regions CDRH1, CDRH2, and CDRH3, and a light chain variable region including light chain complementarity determining regions CDRL1, CDRL2, and CDRL3, (a) The CDRH1 comprises the amino acid sequence of SEQ ID NO: 51, The CDRH2 contains the amino acid sequence of SEQ ID NO: 52 or 57, The CDRH3 contains the amino acid sequence of Sequence ID No. 53, The aforementioned CDRL1 contains the amino acid sequence of SEQ ID NO: 54, The aforementioned CDRL2 contains the amino acid sequence of SEQ ID NO: 55, The CDRL3 contains the amino acid sequence of SEQ ID NO: 56, (b) The CDRH1 comprises the amino acid sequence of SEQ ID NO: 66, The CDRH2 contains the amino acid sequence of SEQ ID NO: 67 or 72, The CDRH3 contains the amino acid sequence of SEQ ID NO: 68, The CDRL1 contains the amino acid sequence of SEQ ID NO: 69, The aforementioned CDRL2 contains the amino acid sequence of SEQ ID NO: 70, The CDRL3 contains the amino acid sequence of SEQ ID NO: 71, (c) The CDRH1 comprises the amino acid sequence of SEQ ID NO: 81, The CDRH2 contains the amino acid sequence of SEQ ID NO: 82 or 87, The CDRH3 contains the amino acid sequence of Sequence ID No. 83, The CDRL1 contains the amino acid sequence of SEQ ID NO: 84, The aforementioned CDRL2 contains the amino acid sequence of SEQ ID NO: 85, The CDRL3 contains the amino acid sequence of SEQ ID NO: 86, or (d) The CDRH1 comprises the amino acid sequence of SEQ ID NO: 96, The CDRH2 contains the amino acid sequence of SEQ ID NO: 97 or 102, The CDRH3 contains the amino acid sequence of SEQ ID NO: 98 or 103, The CDRL1 contains the amino acid sequence of SEQ ID NO: 99, The CDRL2 contains the amino acid sequence of SEQ ID NO: 100, The CDRL3 contains the amino acid sequence of SEQ ID NO: 101, Antibodies or fragments thereof.

2. The antibody or fragment thereof according to claim 1, wherein CDRH1 comprises the amino acid sequence of SEQ ID NO: 51, CDRH2 comprises the amino acid sequence of SEQ ID NO: 52, CDRH3 comprises the amino acid sequence of SEQ ID NO: 53, CDRL1 comprises the amino acid sequence of SEQ ID NO: 54, CDRL2 comprises the amino acid sequence of SEQ ID NO: 55, and CDRL3 comprises the amino acid sequence of SEQ ID NO:

56.

3. The antibody or fragment thereof according to claim 2, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1 and 58-61, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2 and 62-65.

4. The antibody or fragment thereof according to claim 2, wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 59, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:

62.

5. The antibody or fragment thereof according to claim 1, wherein CDRH1 comprises the amino acid sequence of SEQ ID NO: 66, CDRH2 comprises the amino acid sequence of SEQ ID NO: 67 or 72, CDRH3 comprises the amino acid sequence of SEQ ID NO: 68, CDRL1 comprises the amino acid sequence of SEQ ID NO: 69, CDRL2 comprises the amino acid sequence of SEQ ID NO: 70, and CDRL3 comprises the amino acid sequence of SEQ ID NO:

71.

6. The antibody or fragment thereof according to claim 5, wherein the CDRH2 comprises the amino acid sequence of SEQ ID NO:

67.

7. The antibody or fragment thereof according to claim 5, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 and 73-76, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 4 and 77-80.

8. The antibody or fragment thereof according to claim 5, wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 76, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:

79.

9. The antibody or fragment thereof according to claim 1, wherein CDRH1 comprises the amino acid sequence of SEQ ID NO: 81, CDRH2 comprises the amino acid sequence of SEQ ID NO: 82 or 87, CDRH3 comprises the amino acid sequence of SEQ ID NO: 83, CDRL1 comprises the amino acid sequence of SEQ ID NO: 84, CDRL2 comprises the amino acid sequence of SEQ ID NO: 85, and CDRL3 comprises the amino acid sequence of SEQ ID NO:

86.

10. The antibody or fragment thereof according to claim 9, wherein the CDRH2 comprises the amino acid sequence of SEQ ID NO:

82.

11. The antibody or fragment thereof according to claim 9, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 5 and 88-91, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 6 and 92-95.

12. The antibody or fragment thereof according to claim 9, wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 90, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:

92.

13. The antibody or fragment thereof according to claim 1, wherein CDRH1 comprises the amino acid sequence of SEQ ID NO: 96, CDRH2 comprises the amino acid sequence of SEQ ID NO: 97 or 102, CDRH3 comprises the amino acid sequence of SEQ ID NO: 98 or 103, CDRL1 comprises the amino acid sequence of SEQ ID NO: 99, CDRL2 comprises the amino acid sequence of SEQ ID NO: 100, and CDRL3 comprises the amino acid sequence of SEQ ID NO:

101.

14. The antibody or fragment thereof according to claim 13, wherein CDRH2 comprises the amino acid sequence of SEQ ID NO: 97, and CDRH3 comprises the amino acid sequence of SEQ ID NO:

98.

15. The antibody or fragment thereof according to claim 13, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 7 and 104-107, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 8 and 108-111.

16. The antibody or fragment thereof according to claim 13, wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 105, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:

110.

17. A humanized antibody or fragment thereof according to claim 1.

18. An antibody or fragment thereof having binding specificity to the human solute transporter 34A2 (SLC34A2) protein, which binds to the same epitope in the SLC34A2 protein as the antibody or fragment thereof described in claim 1, or which competes with the antibody or fragment thereof described in claim 1 for binding to the SLC34A2 protein.

19. A composition comprising an antibody or fragment thereof according to any one of claims 1 to 18 and a pharmaceutically acceptable carrier.

20. One or more polynucleotides encoding an antibody or a fragment thereof according to any one of claims 1 to 18.

21. A cell comprising one or more polynucleotides as described in claim 20.

22. A composition for treating cancer in a patient requiring treatment for cancer, comprising an antibody or fragment thereof as described in any one of claims 1 to 18.

23. Use of an antibody or fragment thereof according to any one of claims 1 to 18 for the preparation of a pharmaceutical for treating cancer.

24. The composition according to claim 22, wherein the cancer is selected from the group consisting of bladder cancer, liver cancer, colon cancer, rectal cancer, endometrial cancer, leukemia, lymphoma, pancreatic cancer, small cell lung cancer, non-small cell lung cancer, breast cancer, urethral cancer, head and neck cancer, gastrointestinal cancer, stomach cancer, esophageal cancer, ovarian cancer, kidney cancer, melanoma, prostate cancer, and thyroid cancer.

25. The use according to claim 23, wherein the cancer is selected from the group consisting of bladder cancer, liver cancer, colon cancer, rectal cancer, endometrial cancer, leukemia, lymphoma, pancreatic cancer, small cell lung cancer, non-small cell lung cancer, breast cancer, urethral cancer, head and neck cancer, gastrointestinal cancer, stomach cancer, esophageal cancer, ovarian cancer, kidney cancer, melanoma, prostate cancer, and thyroid cancer.

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