Anti-NME antibody and method of treating cancer or cancer metastasis

Abstract

To provide anti-NME family member antibodies useful for treating or preventing diseases associated with the expression of NME family members, such as cancer, in a subject.SOLUTION: The present invention provides an NME7 specific antibody or fragment thereof that binds to the NME7 B3 peptide with a specific amino acid sequence.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] 1. Field of the Invention The present invention relates to NME protein, peptides derived from NME protein, antibodies generated from those peptides, or antibodies or antibody fragments selected for their ability to bind to said peptides. The present invention also relates to the treatment or prevention of diseases associated with expression of NME in patients. [Background technology]

[0002] 2. Background and current technology NDPK (nucleoside diphosphate protein kinase) proteins are a family of proteins grouped together because they all contain an NDPK domain. The first NME proteins discovered, formerly called NM23 proteins, were NM23-H1 and NM23-H2. For decades, it was unclear whether they induce or inhibit hematopoietic cell differentiation. We previously reported that NM23-H1, when dimerized, binds to MUC1. * It binds to growth factor receptors and prevents differentiation, but at higher concentrations of NM23-H1, it forms hexamers, which are involved in the differentiation of MUC1. * We found that NM23 does not bind to MUC1, which is overexpressed in most cancers, and induces differentiation. NM23 was called a metastasis suppressor when it was found to be underexpressed in certain highly aggressive cancers. We previously showed that NM23-H1 dimers bind to MUC1, which is overexpressed in most cancers. * They found that NM23 binds to the extracellular domain of growth factor receptors, dimerizing them, and that such binding promotes cancer cell proliferation. Conversely, at higher concentrations, NM23 inhibits the activity of MUC1. *They form tetramers and hexamers that do not bind to ATP and do not promote tumorigenesis. To date, their functions have not been elucidated, but more NME family proteins (NME1-10) have been discovered recently. NME7 is a newly discovered NME family protein, but its NDPK domain, unlike other NME family members, does not have enzymatic activity. NME7 is not expressed at all or at very low levels in adult tissues. Summary of the Invention [Problem to be solved by the invention]

[0003] The present invention relates to a method for treating or preventing cancer in a patient, comprising administering to the subject an antibody raised against an NME family member. The NME family may be the NME7 family. The antibody may bind to NME7. The antibody may bind to NME7. AB or NME7 AB The antibody may bind to a protein similar to NME7-X1. The antibody may inhibit binding between NME7 and its cognate binding partner, the cognate binding partner being MUC1. * The cognate binding partner may be MUC1 * The antibody may be a PSMGFR portion of the extracellular domain. In one embodiment, the antibody may be generated or selected for its ability to bind to a peptide selected from those listed in Figures 6-9 (SEQ ID NOS: 88-145). Preferably, the peptide may be selected from those listed in Figure 9 (SEQ ID NOS: 141-145).

[0004] The peptides may be highly homologous to the peptides listed in Figures 6-9 (SEQ ID NOS: 88-145) or may have up to 7, up to 6, up to 5, up to 4, up to 3, up to 2, or up to 1 amino acid residue added or deleted at the N-terminus or C-terminus. In one embodiment, the antibody is directed against NME7 but not against NME1. ABor may be selected for its ability to bind to NME7-X1. The antibody may be polyclonal, monoclonal, bivalent, monovalent, bispecific, an antibody fragment containing a variable domain, or an antibody mimetic. The antibody may be a human antibody or a humanized antibody. The antibody may be a single-chain scFv.

[0005] In another aspect, the present invention provides NME7 AB The present invention relates to a method of treating or preventing cancer in a subject, comprising administering to the subject a peptide highly homologous or identical to a region thereof. The peptide may be at least 80% homologous to one or more of the peptides listed in Figure 6. The peptide may be at least 80% homologous to one or more of the peptides listed in Figure 7. The peptide may be at least 80% homologous to one or more of the peptides listed in Figure 8. The peptide may be at least 80% homologous to one or more of the peptides listed in Figure 9. The peptide may be selected from the peptides listed in Figures 6-9 (SEQ ID NOs: 88-145). The peptide may be selected from those listed in Figure 9 (SEQ ID NOs: 141-145). Alternatively, the peptides can be highly homologous to the peptides listed in Figures 6-9 (SEQ ID NOS: 88-145), or have up to 7, up to 6, up to 5, up to 4, up to 3, up to 2, or up to 1 amino acid residue added or deleted at the N-terminus or C-terminus thereof. The peptides can be connected to another peptide via a spacer or linker.

[0006] In another aspect, the present invention relates to a chimeric antigen receptor (CAR) for the treatment or prevention of cancer, wherein the target extracellular portion of the CAR comprises at least a peptide fragment of a member of the NME family. The NME family can be the NME7 family. The member of the NME7 family can be NME7. Alternatively, the member of the NME7 family can be NME7. AB or NME7 ABThe NME7 family member may be an NME7-like protein. The NME7 family member may also be NME7-X1. The targeting extracellular portion of the CAR may comprise one of the peptides listed in Figures 6-9 (SEQ ID NOS: 88-145). The peptide may be selected from those listed in Figure 9 (SEQ ID NOS: 141-145). The peptide may include peptides that are highly homologous to, or have up to seven, up to six, up to five, up to four, up to three, up to two, or up to one amino acid residue added or deleted at the N- or C-terminus of, a peptide listed in Figures 6-9 (SEQ ID NOS: 88-145). The peptide may be connected to another peptide via a spacer or linker.

[0007] In yet another aspect, the present invention relates to a method for treating or preventing cancer or cancer metastasis, comprising engineering a chimeric antigen receptor according to claim 3 into an immune system cell and administering the cell to a subject in need thereof.

[0008] In another aspect, the present invention relates to a chimeric antigen receptor (CAR) for the treatment or prevention of cancer, wherein the target extracellular portion of the chimeric antigen receptor is NME7. AB , NME7 AB The antibody portion may be a single chain scFv, or may be a portion of a human or humanized antibody.

[0009] In yet another aspect, the present invention relates to a method of vaccinating a human against cancer or metastatic cancer, comprising immunizing the human with a peptide fragment of a member of the NME family. The NME family may be the NME7 family. The member of the NME7 family may be NME7 or NME7b. The member of the NME7 family may be NME7. AB or NME7 ABThe NME7 family member may be NME7-X1. The immunizing peptide may be one peptide from the peptides listed in Figures 6-9 (SEQ ID NOS: 88-145). Preferably, the peptide may be selected from those listed in Figure 9 (SEQ ID NOS: 141-145). The immunizing peptide may include peptides highly homologous to the peptides listed in Figures 6-9 (SEQ ID NOS: 88-145) or having up to seven, up to six, up to five, up to four, up to three, up to two, or up to one amino acid residue added or deleted at the N- or C-terminus thereof. The immunizing peptide may be connected to another peptide by a spacer or linker.

[0010] In another aspect, the present invention provides NME7, NME7b, NME7 AB The present invention relates to a method for treating or preventing cancer in a subject, comprising administering to the subject a nucleic acid that inhibits the expression of NME7, NME7-X1-like protein, or NME7-X1-like protein. AB The nucleic acid may be an antisense nucleic acid that inhibits the expression of NME7, NME7-like proteins, or NME7-X1. AB The inhibitory RNA may be an inhibitory RNA, siRNA, RNAi or shRNA that inhibits the expression of NME7-X1-like protein or NME7-X1.

[0011] In another aspect, the present invention provides NME7, NME7b, NME7 AB The present invention relates to a method for treating or preventing cancer in a subject, comprising administering to the subject a nucleic acid that has been gene-edited to suppress expression of NME7, NME7b, NME7-like proteins, or NME7-X1. AB Genetically edited nucleic acids that suppress expression of NME7, NME7b, NME7-like proteins, or NME7-X1 can be inserted into cells that can then be administered to the patient. AB The genetically edited nucleic acid that suppresses expression of the NME7-X1-like protein or NME7-X1 can be inserted into cells using a viral vector, which can be a lentiviral system.

[0012] In another aspect, the present invention provides NME7 AB , NME7b, NME7 AB The present invention also relates to a method of proliferating cancer cells comprising contacting the cells with an NME7-like protein, or an NME7-X1, 2i, or 5i. AB , NME7b, NME7 AB culturing the cells in a medium containing a human NME7-like protein or NME7-X1, 2i or 5i; AB , NME7b, NME7 AB expressing an NME7-like protein, or NME7-X1, or NME7 AB , NME7b, NME7 AB The method includes growing cells in an animal to which a NME7-X1-like protein or NME7-X1 is administered. The cancer cells can be breast, prostate, ovarian, colorectal, pancreatic, liver, melanoma, or brain cancer cells. Drug candidates can be tested on the cells. Drug efficacy can be evaluated by comparing cancer growth to a no-drug control, or comparing expression levels of metastasis markers or stem cell markers to a no-drug control, or comparing the ability of resulting cells from low cell copy number compared to a no-drug control to form tumors in animals, and determining the efficacy of the candidate drug for treating cancer or metastasis. The cells are obtained from a patient to be evaluated for cancer treatment, and a drug likely to be effective for that patient is selected based on results using the above-described method. The cells are not obtained from a patient to be evaluated for cancer treatment, but a drug likely to be effective for that patient is selected based on results using the above-described method.

[0013] In another aspect, the present invention relates to a method for generating antibodies or antibody-like molecules from peptides or peptidomimetics having sequences derived from the sequence of NME. The NME can be NME7. The peptide can be used as an immunogen to generate antibodies or antibody-like molecules. The peptide can be administered to animals to generate anti-NME7 antibodies. The peptide can be administered to humans to generate anti-NME7 antibodies. The peptide can have a sequence listed in Figures 6-9 (SEQ ID NOS: 88-145). Preferably, the peptide can be selected from those listed in Figure 9 (SEQ ID NOS: 141-145). The peptide can include peptides highly homologous to the peptides listed in Figures 6-9 (SEQ ID NOS: 88-145) or having up to 7, up to 6, up to 5, up to 4, up to 3, up to 2, or up to 1 amino acid residue added or deleted at the N- or C-terminus thereof.

[0014] In another aspect, the present invention relates to a method for detecting the presence or progression of cancer, comprising the steps of: 1) obtaining a sample from a patient who has cancer or is at risk of developing cancer; 2) subjecting the sample to an assay capable of detecting or measuring levels of an NME7 family member or levels of a nucleic acid encoding an NME7 family member; 3) comparing the level of the measured member of the NME7 family, or a nucleic acid encoding a member of the NME7 family, in the test sample with the level in a control patient or control cell; 4) determining that the level of an NME7 family member, or a nucleic acid encoding an NME7 family member, is elevated compared to a control; and 5) concluding that the cancer is progressing if the donor of the test sample is afflicted with cancer or if the control to which the test sample is compared is from a donor previously diagnosed with cancer. In this method, detection of an NME7 family member in the circulatory system or in tissues can be an indicator of cancer in the patient. NME7 family members include NME7, NME7b, NME7-X1, or NME7. AB It may be a protein similar to the .

[0015] In yet another aspect, the present invention provides a method for detecting an NME7 family member or MUC1 in a patient. * detecting the presence of an NME7 family member or MUC1 * Anti-NME7 or anti-MUC1 in patients who express * The method includes administering an antibody or antibodies to a patient having an NME7 family member selected from the group consisting of NME7, NME7b, NME7-X1, and NME7. AB It may be a protein similar to the .

[0016] In yet another aspect, the present invention relates to a method for treating or preventing cancer, comprising: 1) obtaining a sample from a patient who has cancer or is at risk of developing cancer or is at risk of developing metastatic cancer; 2) measuring the amount of an NME7 family member or a nucleic acid encoding an NME7 family member, wherein the measured level is significantly higher than that measured in a control sample; 3) determining that the patient has cancer or has developed more aggressive or metastatic cancer; 4) administering to the patient an effective amount of a therapeutic agent that inhibits expression of an NME7 family member, inhibits cleavage of NME7, or inhibits binding of NME7 to its target. The target of NME7 family members is MUC1 * The target of NME7 family members may be MUC1 *The NME7 family member may be NME7, NME7b, NME7-X1, or NME7. AB It may be a protein similar to the .

[0017] In any of the above methods relating to cancer, the cancer may include breast, prostate, ovarian, colorectal, pancreatic, liver, melanoma or brain cancer.

[0018] In one aspect, the present invention relates to an NME7-specific antibody or fragment thereof that binds to the NME7 B3 peptide of SEQ ID NO: 145 or SEQ ID NO: 169. The antibody can be a monoclonal antibody or a bivalent, monovalent, Fab, or single-chain variable fragment. The antibody can be linked to an antibody-drug conjugate. The drug can be linked to a toxin or protoxin.

[0019] The present invention also relates to an isolated nucleic acid encoding the antibody.

[0020] The present invention also relates to an isolated hybridoma expressing the above-described monoclonal antibody. AB Alternatively, the antibody may specifically bind to NME7-X1 but not to NME1. AB and MUC1 * Interaction between the extracellular domains, or NME7-X1 and MUC1 * Alternatively, the antibody may disrupt the interaction between the extracellular domains of NME7. AB and PSMGFR, or between NME7-X1 and PSMGFR. AB and N-10, or the bond between NME7-X1 and N-10.

[0021] In another embodiment, the antibody is NME7 AB and MUC1 * Interaction between the extracellular domains, or NME7-X1 and MUC1 * May not disrupt interactions between extracellular domains. ABAlternatively, NME7-X1 binds to the N-10 peptide (SEQ ID NO: 170) but not to the C-10 peptide (SEQ ID NO: 171). In particular, the antibody can be 5A1, 4A3, or 5D4.

[0022] The antibody comprises an amino acid sequence in the heavy chain variable region comprising: YTFTNYGMN (SEQ ID NO: 439) in the CDR1 region; WINTYTGEPTYVDDFKG (SEQ ID NO: 440) in the CDR2 region; LRGIRPGPLAY (SEQ ID NO: 441) in the CDR3 region; and An amino acid sequence in the light chain variable region comprising: SASSSVSYMN (SEQ ID NO: 444) in the CDR1 region; GISNLAS (SEQ ID NO: 445) in the CDR2 region; QQRSSYPPT (SEQ ID NO: 446) in the CDR3 region may include:

[0023] In another embodiment, the antibody comprises an amino acid sequence in the heavy chain variable region comprising: NTFTEYTMH (SEQ ID NO: 429) in the CDR1 region; GFNPNNGVTNYNQKFKG (SEQ ID NO: 430) in the CDR2 region; RYYHSTYVFYFDS (SEQ ID NO: 431) in the CDR3 region; and An amino acid sequence in the light chain variable region comprising: SASQGISNYLN (SEQ ID NO: 434) in the CDR1 region; YTSSLHS (SEQ ID NO: 435) in the CDR2 region; QQYSKLPYT (SEQ ID NO: 436) in the CDR3 region may include:

[0024] In another embodiment, the antibody comprises an amino acid sequence in the heavy chain variable region comprising: NTFTEYTMH (SEQ ID NO: 388) in the CDR1 region; GFNPNNGVTNYNQKFKG (SEQ ID NO: 389) in the CDR2 region; RYYHSLYVFYFDY (SEQ ID NO: 390) in the CDR3 region; and An amino acid sequence in the light chain variable region comprising: ITSTDIDDDMN (SEQ ID NO: 393) in the CDR1 region; EGNTLRP (SEQ ID NO: 394) in the CDR2 region; LQSDNLPLT (SEQ ID NO: 395) in the CDR3 region may include:

[0025] The antibody can be a human antibody or a humanized antibody or an engineered antibody mimetic. The antibody may be a non-human antibody, for example a murine or camelid antibody.

[0026] The present invention also relates to a method for administering to a patient for the prevention or treatment of cancer, the method comprising administering to the patient a composition comprising the above-described antibody.

[0027] The present invention also relates to a method for preventing or treating cancer metastasis in a patient, the method comprising administering to the patient a composition comprising the above-described antibody.

[0028] The present invention also relates to a method for diagnosing cancer or cancer metastasis, comprising contacting a patient sample and a normal sample with the antibody and comparing the results obtained from both samples, wherein the presence of positive binding to the antibody in the patient sample indicates the presence of cancer or cancer metastasis in the patient. The antibody may be linked to an imaging agent. The patient sample may be blood, body fluids, tissue, or circulating cells in vitro, including intraoperatively, or in vivo.

[0029] The present invention also provides an anti-NME7 AB The present invention relates to cells engineered to express an antibody or fragment thereof. The cells may be immune cells, such as T cells or NK cells, or stem or progenitor cells, preferably stem or progenitor cells that subsequently differentiate into T cells.

[0030] The cells may contain a chimeric antigen receptor (CAR) that recognizes a tumor-associated antigen. Expression of the anti-NME7 antibody may be inducible. AB The nucleic acid encoding the antibody can be inserted within a Foxp3 enhancer or promoter. AB The antibody can be present in an NFAT induction system. An NFATc1 response element can be inserted upstream of the antibody sequence inserted into the enhancer or promoter region.

[0031] anti-NME7 AB The antibody or fragment thereof may bind to the NME7 B3 peptide or AB or NME7-X1, MUC1 * The binding of the extracellular domain to PSMGFR may be disrupted.

[0032] The CAR can recognize a tumor-associated antigen and an anti-NME7 antibody. The tumor-associated antigen is MUC1. * It could be.

[0033] The present invention also provides NME7, listed in Figures 6-9, as an immunogenicity-inducing moiety. AB The present invention relates to an anti-cancer vaccine comprising a composition comprising one or more peptides derived from, or having at least 80%, 85%, 90%, 95%, or 97% sequence identity thereto. The peptides can be peptides of SEQ ID NOs: 141-145, or peptides having at least 80%, 85%, 90%, 95%, or 97% sequence identity thereto. The peptides can be SEQ ID NO: 145, or peptides having 80%, 85%, 90%, 95%, or 97% sequence identity thereto.

[0034] In another aspect, the present invention relates to a BiTE comprising the above-described antibody.

[0035] In yet another aspect, the present invention provides an anti-NME7 AB For methods of generating antibodies, cysteine ​​residues in the NME7 B3 peptide are mutated to avoid disulfide bonds.

[0036] In yet another aspect, the present invention provides a method for the production of NME7 AB or a method for generating cells with enhanced metastatic potential, comprising culturing the cells with NME7-X1.

[0037] The present invention also provides NME7 AB or cells engineered to express NME7-X1, NME7 AB or NME7-X1-expressing transgenic animals, AB Alternatively, NME7-X1 can be human, and NME7 AB Alternatively, expression of NME7-X1 can be inducible.

[0038] The patent or application file contains at least one color drawing. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee. The present invention may be more fully understood from the detailed description given below and the accompanying drawings, which are given for purposes of illustration only and are not intended to be limiting of the invention. [Brief explanation of the drawings]

[0039] [Figure 1] 1 shows a graph of HRP signal from an ELISA sandwich assay demonstrating that NME7AB dimerizes the MUC1* extracellular domain peptide. [Figure 2] 1 shows a graph of RT-PCR measurement of gene expression of stem cell markers and cancer stem cell markers for T47D cancer cells after culture in conventional or NME7-containing medium. Cells that became non-adherent (floating) were analyzed separately from those that remained adherent. [Figure 3]1 shows a graph of RT-PCR measurements of gene expression of various stem and putative cancer stem cell markers for DU145 prostate cancer cells. Cells were cultured in either conventional medium or medium containing NME1 dimer ("NM23") or NME7 (NME7AB). Rho kinase inhibitors were not used because cells remained adherent by passage 2. [Figure 4] 1 is a graph of RT-PCR measurements of metastatic and pluripotent stem cell markers showing that 2i inhibitors (GSK3beta and MEK inhibitors), previously shown to revert stem cells to a more naive state, also induce cancer cells into a more metastatic state, although to a lesser extent than NME7AB. [Figure 5] Sequence alignment between human NME1 and human NME7-A or -B domains. [Figure 6-1] Listed are immunogenic peptides derived from human NME7 that have low sequence identity to NME1 and were selected for their ability to generate therapeutic anti-NME7 antibodies for the treatment or prevention of cancer. [Figure 6-2] Listed are immunogenic peptides derived from human NME7 that have low sequence identity to NME1 and were selected for their ability to generate therapeutic anti-NME7 antibodies for the treatment or prevention of cancer. [Figure 7] Listed are immunogenic peptides derived from human NME7 that may be important for structural integrity or for binding to MUC1* and that were selected for their ability to generate therapeutic anti-NME7 antibodies for the treatment or prevention of cancer. [Figure 8] Listed are immunogenic peptides derived from human NME1 that may be important for structural integrity or for binding to MUC1* and that were selected for their ability to generate therapeutic anti-NME7 antibodies for the treatment or prevention of cancer. [Figure 9]Listed below are immunogenic peptides derived from human NME7, selected for their low sequence identity to NME1 and for their homology to bacterial NME1 proteins, which have been linked to cancer. These peptides are more preferred for their ability to generate therapeutic anti-NME7 antibodies for the treatment or prevention of cancer. The peptide shown in this figure contains an additional covalently linked cysteine ​​at the C-terminus. [Figure 10] 10A and 10B show graphs of an ELISA assay in which NME7-AB (FIG. 10A) or NME1 (FIG. 10B) was adsorbed to a plate, and anti-NME7 antibodies generated by NME7 peptides A1, A2, B1, B2, and B3 were tested for their ability to bind to NME7 but not NME1. C20 is an anti-NME1 antibody. [Figure 11] 1 shows a graph of an ELISA assay in which generated anti-NME7 antibodies were tested for their ability to inhibit the binding of NME7-AB, but not NME1, to surface-immobilized MUC1* peptide. [Figure 12]

[0033] Figure 1 shows a graph of a cancer cell proliferation experiment in which breast cancer cells were grown in the presence or absence of NME7 antibodies or short peptides derived from NME7 used to generate or select the antibodies. Furthermore, antibodies generated by immunization with nearly the entire NME7AB peptide, amino acids 100-376, were shown to inhibit cancer cell proliferation. [Figure 13] 1 shows a graph of a cancer cell proliferation experiment in which breast cancer cells were grown in the presence or absence of the NME7 antibody or a combination of short peptides derived from NME7 used to generate or select the antibodies. Both antibodies inhibited cancer cell proliferation similarly to their immunizing NME7AB peptide. [Figure 14]This table shows the scientists' observations when cancer cells were grown in NME7-AB or 2i inhibitors, both of which can transform cancer cells to a more metastatic state, and in the presence or absence of NME7-derived peptides A1, A2, B1, B2, and B3. The NME7-AB peptide inhibited the transition of adherent cancer cells to suspension cells, and RT-PCR measurements showed increased expression of metastatic markers, particularly CXCR4. [Figure 15A] Figure 15A shows graphs of RT-PCR measurements of CXCR4 and other metastatic marker expression in T47D breast cancer cells grown in NME7AB or 2i inhibitor, which transforms cancer cells to a more metastatic state, and the inhibitory effect of anti-NME7 antibodies on metastatic transformation. Figure 15A is a PCR graph of CXCR4 expression in T47D cancer cells grown in NME7AB or 2i in the presence or absence of anti-NME7 antibodies. Figure 15B is a graph of RT-PCR measurements of CXCR4, CHD1, and SOX2 expression in T47D breast cancer cells grown in 2i inhibitor for 72 or 144 hours in the presence of the NME7AB immunizing peptide, demonstrating that the peptides themselves are inhibitory to metastatic transformation. Peptides A1, A2, and B1 used in inhibitory combos 2 and 3 in Figure 15A are also inhibitory as peptides. Peptide B3 is the immunizing peptide for antibody 61, which was the most inhibitory antibody tested in Figure 15A. FIG. 15C is a graph in which the Y axis of FIG. 15B is enlarged. [Figure 15B]Figure 15A shows graphs of RT-PCR measurements of CXCR4 and other metastatic marker expression in T47D breast cancer cells grown in NME7AB or 2i inhibitor, which transforms cancer cells to a more metastatic state, and the inhibitory effect of anti-NME7 antibodies on metastatic transformation. Figure 15A is a PCR graph of CXCR4 expression in T47D cancer cells grown in NME7AB or 2i in the presence or absence of anti-NME7 antibodies. Figure 15B is a graph of RT-PCR measurements of CXCR4, CHD1, and SOX2 expression in T47D breast cancer cells grown in 2i inhibitor for 72 or 144 hours in the presence of the NME7AB immunizing peptide, demonstrating that the peptides themselves are inhibitory to metastatic transformation. Peptides A1, A2, and B1 used in inhibitory combos 2 and 3 in Figure 15A are also inhibitory as peptides. Peptide B3 is the immunizing peptide for antibody 61, which was the most inhibitory antibody tested in Figure 15A. FIG. 15C is a graph in which the Y axis of FIG. 15B is enlarged. [Figure 15C] Figure 15A shows graphs of RT-PCR measurements of CXCR4 and other metastatic marker expression in T47D breast cancer cells grown in NME7AB or 2i inhibitor, which transforms cancer cells to a more metastatic state, and the inhibitory effect of anti-NME7 antibodies on metastatic transformation. Figure 15A is a PCR graph of CXCR4 expression in T47D cancer cells grown in NME7AB or 2i in the presence or absence of anti-NME7 antibodies. Figure 15B is a graph of RT-PCR measurements of CXCR4, CHD1, and SOX2 expression in T47D breast cancer cells grown in 2i inhibitor for 72 or 144 hours in the presence of the NME7AB immunizing peptide, demonstrating that the peptides themselves are inhibitory to metastatic transformation. Peptides A1, A2, and B1 used in inhibitory combos 2 and 3 in Figure 15A are also inhibitory as peptides. Peptide B3 is the immunizing peptide for antibody 61, which was the most inhibitory antibody tested in Figure 15A. FIG. 15C is a graph in which the Y axis of FIG. 15B is enlarged. [Figure 16]A table of threshold cycle numbers for control housekeeping genes and CXCR4 expression, as well as recorded RNA levels in samples used for RT-PCR measurement of CXCR4 in FIG. 31 is shown. [Figure 17] 1 shows a graph of RT-PCR measurements of NME7-X1 expression in a series of human stem cells and cancer cells. [Figure 18] Graph showing RT-PCR measurements of NME7, NME7a, NME7b, and NME7-X1 expression in a series of human stem cells and cancer cells. NME7a is full-length NME7, NME7b lacks a small portion of the DM10 domain, and NME7-X1 lacks all of the DM10 domain and a small portion of the N-terminus of the first NDPK A domain. The bar labeled NME7 indicates that primers detecting both NME7a and NME7b were used. [Figure 19A-C]Figure 19 shows photographs of Western blots in which various cancer cell lines were detected for expression of NME7 species using antibodies generated by immunization with NME7-derived peptides. Figure 19A shows a Western blot in which antibody 52, which binds to the A1 peptide, was used to detect the presence of full-length NME7, NME7AB, or NME7-X1 in a series of cells. Figure 19B shows a Western blot in which antibody 56, which binds to the B1 peptide, was used to detect the presence of full-length NME7, NME7AB, or NME7-X1 in a series of cells. Figure 19C shows a Western blot in which antibody 61, which binds to the B3 peptide, was used to detect the presence of full-length NME7, NME7AB, or NME7-X1 in a series of cells. Figure 19D shows a Western blot in which commercially available polyclonal antibody H278, raised against both NME7 A and B, was used to detect the presence of full-length NME7 in a series of cells. As the figure shows, antibody H278 also recognizes NME1. Figure 19E shows a gel published on a website for the commercially available anti-NME7 antibody B9, demonstrating that it binds to a species with the apparent molecular weight of full-length NME7. Figure 19F shows a Western blot in which anti-NME7 antibody B9 was used to detect a gel loaded with NME1 alone. As can be seen, antibody B9 recognizes NME1 as well as full-length NME7. This is not surprising because, like antibody H278, B9 was raised against both the A and B domains of NME7, and the A domain of NME1 is highly homologous to the A domain of NME7AB. [Figure 19D-F]Figure 19 shows photographs of Western blots in which various cancer cell lines were detected for expression of NME7 species using antibodies generated by immunization with NME7-derived peptides. Figure 19A shows a Western blot in which antibody 52, which binds to the A1 peptide, was used to detect the presence of full-length NME7, NME7AB, or NME7-X1 in a series of cells. Figure 19B shows a Western blot in which antibody 56, which binds to the B1 peptide, was used to detect the presence of full-length NME7, NME7AB, or NME7-X1 in a series of cells. Figure 19C shows a Western blot in which antibody 61, which binds to the B3 peptide, was used to detect the presence of full-length NME7, NME7AB, or NME7-X1 in a series of cells. Figure 19D shows a Western blot in which commercially available polyclonal antibody H278, raised against both NME7 A and B, was used to detect the presence of full-length NME7 in a series of cells. As the figure shows, antibody H278 also recognizes NME1. Figure 19E shows a gel published on a website for the commercially available anti-NME7 antibody B9, demonstrating that it binds to a species with the apparent molecular weight of full-length NME7. Figure 19F shows a Western blot in which anti-NME7 antibody B9 was used to detect a gel loaded with NME1 alone. As can be seen, antibody B9 recognizes NME1 as well as full-length NME7. This is not surprising because, like antibody H278, B9 was raised against both the A and B domains of NME7, and the A domain of NME1 is highly homologous to the A domain of NME7AB. [Figure 20A-B]Figure 20 shows graphs of RT-PCR measurements of metastatic markers in cancer cells after culturing in serum-free medium containing NME7AB compared to standard medium. Figure 20A shows that SK-OV3, a MUC1-positive ovarian cancer cell line, increased expression of metastatic markers CXCR4, CDH1 (also known as E-cadherin), SOX2, and NME7-X1. Figure 20B shows that OV-90, a MUC1-negative ovarian cancer cell line, increased expression of metastatic markers CXCR4 and NME7-X1. Figure 20C shows that MDA-MB, a breast cancer cell line expressing minimal levels of MUC1, increased expression of metastatic markers CDH1 (also known as E-cadherin) and SOX2. [Figure 20C] Figure 20 shows graphs of RT-PCR measurements of metastatic markers in cancer cells after culturing in serum-free medium containing NME7AB compared to standard medium. Figure 20A shows that SK-OV3, a MUC1-positive ovarian cancer cell line, increased expression of metastatic markers CXCR4, CDH1 (also known as E-cadherin), SOX2, and NME7-X1. Figure 20B shows that OV-90, a MUC1-negative ovarian cancer cell line, increased expression of metastatic markers CXCR4 and NME7-X1. Figure 20C shows that MDA-MB, a breast cancer cell line expressing minimal levels of MUC1, increased expression of metastatic markers CDH1 (also known as E-cadherin) and SOX2. [Figure 21A-C]Photographs and descriptions of Western blots of analyzed cancer cell lines are shown. For the Western blots in Figures 21A and 21B, all cancer samples were normalized so that they were loaded onto the gel at a concentration of 40 μg / mL. In Figure 21A, the expression of full-length MUC1 in various cancer cell lines is detected using the anti-tandem repeat monoclonal antibody VU4H5. In Figure 21B, various cancer cell lines are detected for the expression of truncated MUC1* using a polyclonal anti-PSMGFR antibody. Figure 21C is a description of the analyzed cancer cell lines. Figure 21D shows that HER2-positive BT474 breast cancer cells, labeled "BT474 (parental cells)," express little to no MUC1 or MUC1* until they acquire resistance to Herceptin and other chemotherapy drugs (labeled "BTRes1" in the figure). Parental cells were made resistant to Herceptin, Taxol, doxorubicin, and cyclophosphamide by culturing the cells in sublethal levels of Herceptin. Figure 21D shows that HER2 expression levels remained unchanged, while MUC1* expression dramatically increased as the cells acquired resistance to Herceptin. Figure 21E shows a graph of the proliferation of parental BT474 cells compared to drug-resistant metastatic cells in response to treatment with Herceptin in the presence or absence of anti-MUC1* Fab. As can be seen, BT474 parental cells exhibit a Herceptin concentration-dependent decrease in cell proliferation, while two Herceptin-resistant cell lines, BTRes1 and BTRes2, do not exhibit a decrease in cancer cell proliferation in response to treatment with Herceptin. However, when treated with anti-MUC1* Fab, the resistant cell lines exhibit a Herceptin concentration-dependent decrease in cancer cell proliferation. FIG. 21F shows a graph of percent cell death of parental BT474 cells compared to drug-resistant BTRes1 in response to treatment with taxol in the presence or absence of anti-MUC1* Fab. [Figure 21D-F]Photographs and descriptions of Western blots of analyzed cancer cell lines are shown. For the Western blots in Figures 21A and 21B, all cancer samples were normalized so that they were loaded onto the gel at a concentration of 40 μg / mL. In Figure 21A, the expression of full-length MUC1 in various cancer cell lines is detected using the anti-tandem repeat monoclonal antibody VU4H5. In Figure 21B, various cancer cell lines are detected for the expression of truncated MUC1* using a polyclonal anti-PSMGFR antibody. Figure 21C is a description of the analyzed cancer cell lines. Figure 21D shows that HER2-positive BT474 breast cancer cells, labeled "BT474 (parental cells)," express little to no MUC1 or MUC1* until they acquire resistance to Herceptin and other chemotherapy drugs (labeled "BTRes1" in the figure). Parental cells were made resistant to Herceptin, Taxol, doxorubicin, and cyclophosphamide by culturing the cells in sublethal levels of Herceptin. Figure 21D shows that HER2 expression levels remained unchanged, while MUC1* expression dramatically increased as the cells acquired resistance to Herceptin. Figure 21E shows a graph of the proliferation of parental BT474 cells compared to drug-resistant metastatic cells in response to treatment with Herceptin in the presence or absence of anti-MUC1* Fab. As can be seen, BT474 parental cells exhibit a Herceptin concentration-dependent decrease in cell proliferation, while two Herceptin-resistant cell lines, BTRes1 and BTRes2, do not exhibit a decrease in cancer cell proliferation in response to treatment with Herceptin. However, when treated with anti-MUC1* Fab, the resistant cell lines exhibit a Herceptin concentration-dependent decrease in cancer cell proliferation. FIG. 21F shows a graph of percent cell death of parental BT474 cells compared to drug-resistant BTRes1 in response to treatment with taxol in the presence or absence of anti-MUC1* Fab. [Figure 22]Photographs of Western blots from coimmunoprecipitation experiments are shown. T47D breast cancer cell extracts were incubated with Ab-5, an antibody against the cytoplasmic tail of MUC1, or a control antibody, IgG, and coimmunoprecipitated. The gels were blotted with two different commercially available anti-NME7 antibodies, B9 (Figure 22A) and CF7 (Figure 22B). Both gels show distinctive NME7 bands at approximately 33 kDa and 30 kDa. The gels were stripped and reprobed with anti-PSMGFR (Figures 22C and 22D), an antibody against the extracellular domain of MUC1*. This indicates that NME7 species and MUC1* interact with each other. Recombinant NME7-AB and recombinant NME7-X1 were mixed together, run on a gel, and then probed with anti-NME7 antibodies. We show that two unique NME7 species that occur naturally in breast cancer cells and interact with MUC1* are NME7-AB-like and NME7-X1 (FIG. 22E). [Figure 23] Western blot images of coimmunoprecipitation experiments are shown. Extracts from human induced pluripotent stem cells (iPS7 cells) or embryonic stem cells (HES3 cells) were incubated with Ab-5, an antibody against the cytoplasmic tail of MUC1, or a control antibody, IgG, and coimmunoprecipitated. The gel was blotted with the commercially available anti-NME7 antibody B9 (Figure 23A). Both cell types exhibit distinctive NME7 bands at approximately 33 kDa and 30 kDa. The gel was stripped and reprobed with anti-PSMGFR, an antibody against the extracellular domain of MUC1* (Figure 23B). This indicates that NME7 species and MUC1* interact with each other. Recombinant NME7-AB and recombinant NME7-X1 were mixed, run on a gel, and then probed with the anti-NME7 antibody. We show that two unique NME7 species that occur naturally in breast cancer cells and interact with MUC1* are NME7-AB-like and NME7-X1 (FIG. 23C). [Figure 24]Figure 1 shows a graph of an ELISA experiment assaying novel anti-NME7 antibodies for their ability to bind NME7-AB. NME7-AB is known to bind to the extracellular domain of MUC1*. The surface of a multiwell plate was coated with recombinant NME7-AB. Anti-NME7-AB antibodies were added separately to the wells. Standard washes were performed and visualization was performed by adding an HRP-conjugated secondary antibody. As can be seen, seven of the ten anti-NME7 antibodies strongly bound to NME7-AB. [Figure 25] Figure 1 shows a graph of an ELISA experiment assaying novel anti-NME7 antibodies for their ability to bind NME1, or preferably their inability to bind. The surface of a multiwell plate was coated with recombinant NME1-S120G dimer, which is known to bind to the MUC1* extracellular domain. Anti-NME7-AB antibody was added separately to the wells. Standard washes were performed and visualization was performed by adding an HRP-labeled secondary antibody. As can be seen, only one antibody showed only minimal binding to NME1. [Figure 26] A graph of an ELISA competitive inhibition assay is shown. NME7-AB / anti-NME7 antibody complexes were generated and then added to multiwell plates coated with the MUC1* extracellular domain peptide, PSMGFR. Recall that NME7-AB has two pseudo-identical domains, A and B, each capable of binding to the MUC1* extracellular domain. Antibodies that bind to the NME7 B3 peptide in the B domain do not bind to the NME7 A domain. Therefore, only partial inhibition of the NME7-AB / MUC1* interaction is expected. [Figure 27] Graph of ELISA displacement assay: NME7-AB first bound to surface-immobilized MUC1* extracellular domain peptide on the plate and then was cleaved by addition of anti-NME7 antibody. [Figure 28]Graph of ELISA displacement assay is shown. In this case, multiwell plates were coated with a truncated MUC1* peptide, N-10 (containing 10 N-terminal amino acids deleted from the PSMGFR sequence). NME7-AB is known to bind to the N-10 peptide. NME7-AB bound to the surface-immobilized N-10 peptide on the plate and was subsequently cleaved by the addition of anti-NME7 antibody. [Figure 29] This graph shows the amount of RNA present in samples of T47D breast cancer cells cultured in either their normal recommended medium, RMPI, serum-free medium containing only 4 nM (optimal) or 8 nM NME7-AB as a growth factor, or serum-free medium containing only 8 nM NME1 S120G dimer as a growth factor. Because NME1 is a homodimer and NME7-AB is a monomer consisting of two pseudo-identical domains, 8 nM NME1 is the molar equivalent of 4 nM NME7-AB. Cancer cells were cultured in the presence or absence of anti-NME7 B3 antibody. In this experiment, floating cells were separated from adherent cells and analyzed separately. Significant data indicate that the floating cells are cancer stem cells. An increase or decrease in the amount of RNA in a sample indicates an increased or decreased number of cells, respectively, in a given population. [Figure 30] This graph shows PCR measurements of the metastasis marker CXCR4 in T47D breast cancer cells cultured in either their recommended normal medium, RPMI, serum-free medium containing only 4 nM (optimal) or 8 nM NME7-AB as a growth factor, or serum-free medium containing only 8 nM NME1 S120G dimer as a growth factor. Because NME1 is a homodimer and NME7-AB is a monomer consisting of two pseudo-identical domains, 8 nM NME1 is the molar equivalent of 4 nM NME7-AB. Cancer cells were cultured in the presence or absence of anti-NME7 B3 antibody. In this experiment, suspension cells were separated from adherent cells and analyzed separately. Significant data indicate that suspension cells are cancer stem cells. As can be seen from the figure, growth in NME7-AB medium increased CXCR4 expression in the suspension cell population, while anti-NME7 B3 antibody reduced expression, indicating that anti-NME7 antibody reduced the generation of cancer stem cells. [Figure 31]This graph shows PCR measurements of stem cell and metastatic markers in T47D breast cancer cells cultured in either their recommended normal medium, RMPI, serum-free medium containing 4 nM (optimal) or 8 nM NME7AB as the only growth factor, or serum-free medium containing 8 nM NME1 S120G dimer as the only growth factor. Because NME1 is a homodimer and NME7-AB is a monomer consisting of two pseudo-identical domains, 8 nM NME1 is the molar equivalent of 4 nM NME7-AB. Cancer cells were cultured in the presence or absence of anti-NME7 B3 antibody. In this experiment, suspension cells were separated from adherent cells and analyzed separately. Significant data indicate that suspension cells are cancer stem cells. As can be seen from the figure, growth in NME7-AB medium increased SOX2 expression in the suspension cell population, while anti-NME7 B3 antibody reduced expression, indicating that anti-NME7 antibody reduced the generation of cancer stem cells. [Figure 32] This graph shows PCR measurements of stem cell marker and metastatic growth factor receptor MUC1 in T47D breast cancer cells cultured in either their normal recommended medium, RMPI, serum-free medium containing 4 nM (optimal) or 8 nM NME7-AB as the only growth factor, or serum-free medium containing 8 nM NME1 S120G dimer as the only growth factor. Because NME1 is a homodimer and NME7-AB is a monomer consisting of two pseudo-identical domains, 8 nM NME1 is the molar equivalent of 4 nM NME7-AB. Cancer cells were cultured in the presence or absence of anti-NME7 B3 antibody. In this experiment, floating cells were separated from adherent cells and analyzed separately. Significant data indicate that floating cells are cancer stem cells. As can be seen from the figure, growth in NME7-AB medium increased MUC1 expression in the suspension cell population, while anti-NME7 B3 antibody reduced expression, indicating that anti-NME7 antibody reduced the generation of cancer stem cells. [Figure 33]Figure 33 shows IVIS images of immune-compromised nu / nu mice 6 days after tail vein injection of cancer cells. Figure 33A shows an IVIS image of a mouse injected with 500,000 T47D-wt breast cancer cells. Figure 33B shows an IVIS image of a mouse injected with 10,000 T47D breast cancer cells grown in NME7-AB in minimal media for 10 days. Floating cells were collected. These floating cells are referred to herein as cancer stem cells (CSCs). As can be seen, mice injected with wild-type cancer cells show no signs of metastasis. However, mice injected with cancer stem cells, albeit with 50-fold fewer cells, show clear metastatic potential of the injected cancer cells. [Figure 34] Figure 34 shows IVIS images of immune-compromised nu / nu mice 10 days after tail vein injection of cancer cells. Figure 34A shows IVIS images of a mouse injected with 500,000 T47D-wt breast cancer cells. Figure 34B shows IVIS images of a mouse injected with 10,000 T47D-CSCs (cancer stem cells). Figure 34C shows IVIS images of a mouse injected with 10,000 T47D-CSCs (cancer stem cells) and then injected with anti-NME7 antibody on day 7. Figure 34D shows a handwritten record of IVIS measurements of emitted photons. As can be seen, the mice selected for treatment are more metastatic than comparable T47D-CSC mice. The efficacy of the first antibody injection may have been blocked by the injection of free NME7-AB on day 6. Control mice injected with 500,000 T47D-wt cells show somewhat weak photon emission that may be background or surviving cancer cells. [Figure 35]Figure 35 shows IVIS images of an immunocompromised nu / nu mouse 12 days after tail vein injection of cancer cells. Figure 35A shows an IVIS image of a mouse injected with 500,000 T47D-wt breast cancer cells. Figure 35B shows a mouse injected with 10,000 T47D-CSCs (cancer stem cells) and not treated with anti-NME7 antibody, which died from excessive tumor burden before IVIS images were taken. Figure 35C shows IVIS images of a mouse injected with 10,000 T47D-CSCs (cancer stem cells) and then injected with anti-NME7 antibody on days 7 and 10. As can be seen, the mouse treated with anti-NME7 antibody has cleared cancer metastases. The control mouse injected with 500,000 T47D-wt cells showed less shedding, which may indicate fewer cancer cells or background. [Figure 36] Figure 36 shows IVIS images of immune-compromised nu / nu mice 14 days after tail vein injection of cancer cells. Figure 36A shows IVIS images of a mouse injected with 500,000 T47D-wt breast cancer cells. Figure 36B shows IVIS images of a mouse injected with 10,000 T47D-CSCs (cancer stem cells) and anti-NME7 antibody on days 7, 10, and 12. As can be seen, the mice treated with anti-NME7 antibody are almost completely free of cancer metastases. Control mice injected with 500,000 T47D-wt cells show no photon emission. [Figure 37]Figures 37A, 37C, 37E, 37G, 37I, 37K, 37M, and 37O show IVIS images of a mouse injected with 500,000 T47D-wt cells via the tail vein on day 0. Figures 37B, 37D, 37F, 37H, 37J, 37L, 37N, and 37P show IVIS images of a mouse injected with 10,000 T47D cancer stem cells via the tail vein on day 0, followed by administration of anti-NME7 antibody from days 7 to 17, at which point treatment was discontinued, and then resumed on day 21. Figures 37Q, 37R, 37S, 37T, and 37U show magnified IVIS photographs of treated mice from day 17, when anti-NME7 antibody treatment was discontinued, to day 21, when antibody treatment was resumed, through day 26. Figure 37V shows the scale bar for IVIS measurements. As can be seen from this time course, cancer cells grown in NME7 readily metastasize, and such metastasis can be effectively treated, prevented, or ameliorated by treatment with antibodies that bind to NME7. [Figure 38] Figure 38 shows time course IVIS images from days 6 to 19 after tail vein injection of 500,000 T47D wild-type breast cancer cells or 10,000 T47D cancer stem cells into immune-compromised nu / nu mice. Figure 38A shows a mouse injected via the tail vein (iv), Figure 38B shows a mouse injected intraperitoneally (ip), and Figure 38C shows a mouse injected subcutaneously (sc). [Figure 39] Figure 1 shows human lung tissue specimens stained with an anti-NME7 antibody that binds to the B3 peptide, demonstrating the lack of NME7 expression in normal tissues, increasing expression of NME7 with tumor grade and increased metastasis. [Figure 40] Figure 1 shows a human small intestine tissue specimen stained with an anti-NME7 antibody that binds to the B3 peptide, demonstrating the lack of NME7 expression in normal tissue, increasing expression of NME7 with tumor malignancy and increased metastasis. [Figure 41] Figure 1 shows a human colon tissue specimen stained with an anti-NME7 antibody that binds to the B3 peptide, demonstrating the lack of NME7 expression in normal tissue, increasing expression of NME7 with tumor malignancy and increased metastasis. [Figure 42] Figures 42A-42C show IVIS photographs of animals facing downward. Figures 42D-42F show IVIS photographs of animals facing upward. Figures 42A and 42D show control animals injected with phosphate-buffered saline. Figures 42B and 42E show a prophylactic model in which animals were injected with anti-NME7AB antibody 4A3 24 hours before injection of metastatic cancer cells, followed by 12 antibody injections approximately every other day for 22 days. Figures 42C and 42F show a recovery model in which animals were injected with anti-NME7AB antibody 4A3 24 hours after injection of metastatic cancer cells, followed by approximately every other day for a total of 11 injections of antibody over a 20-day period. [Figure 43] Figures 43A-43C show IVIS images of animals facing downward. Figures 43D-43F show IVIS images of animals facing upward. Figures 43A and 43D show control animals injected with phosphate-buffered saline. Figures 43B and 43E, 43C, and 43F show a prophylactic model in which animals were injected with anti-NME7AB antibody 24 hours before injection of metastatic cancer cells, followed by 12 antibody injections approximately every other day for 22 days. Images were acquired on day 27. [Figure 44]Figure 44A shows a photograph of a female nu / nu mouse injected into the tail vein with 10,000 luciferase-positive T47D metastatic breast cancer stem cells mixed with NME7AB to a final concentration of 32 nM on day 0. On days 1 and 2, the animals were further injected into the tail vein with 32 nM NME7AB, which showed an increase in metastasis. This is a system to demonstrate the recovery of existing metastases. On day 7, the animals were treated with individual anti-NME7AB antibodies: 8F9A5A1, 8F9A4A3, or 5F3A5D4. Figure 44A shows a control animal injected with phosphate-buffered saline. Figure 44B shows an animal treated with anti-NME7AB monoclonal antibody 8F9A5A1 (also known as 5A1). Figure 44C shows an animal treated with anti-NME7AB monoclonal antibody 8F9A4A3 (also known as 4A3). Figure 44D shows animals treated with anti-NME7AB monoclonal antibody 5F3A5D4 (also known as 5D4). Green arrows indicate the low dose (5-7 mg / kg) over the indicated time period, and red arrows indicate the high dose (15 mg / kg). As can be seen, animals treated with anti-NME7AB antibody have fewer metastases than control animals, even though many animals in the group to be treated with the antibody had more metastases before treatment. Higher concentrations of anti-NME7AB antibody are more effective than lower concentrations. For example, animals were treated with a high dose between days 11 and 17, and most treated animals had eliminated metastases by approximately day 17. However, one low dose of antibody resulted in a recurrence of metastases. Animals again responded to high-dose treatment by day 32. [Figure 45]Figure 45A shows a photograph of a female nu / nu mouse subcutaneously injected with 10,000 luciferase-positive T47D metastatic breast cancer stem cells mixed with NME7AB to a final concentration of 32 nM and then mixed 1:1 vol:vol with Matrigel in the right flank on day 0. Tumor engraftment was allowed to progress from day 0 to day 6. The animal was then treated intravenously with anti-NME7AB antibody via tail vein injection. Control animals were injected with PBS. Figure 45A shows an IVIS photograph of a control animal. Figure 45B shows an IVIS photograph of an animal injected via the tail vein with a cocktail of anti-NME7AB antibodies 5A1, 4A3, and 5D4 at a total concentration of 15 mg / kg. Antibody or PBS was administered four times between days 7 and 18. As can be seen, mice treated with anti-NME7AB antibody exhibit fewer metastases than the control group. In the treatment group, two of the five animals had primary tumors larger than those in the control group. This is likely because anti-NME7AB antibodies prevented cancer cells from spreading, so they remained concentrated in the primary tumor. In this experiment, PCR analysis showed that after 11 days in culture with NME7AB, T47D breast cancer cells upregulated CXCR4 by 109-fold, OCT4 by 2-fold, NANOG by 3.5-fold, and MUC1 by 2.7-fold. [Figure 46]Photographs of female nu / nu mice were subcutaneously injected in the right flank on day 0 with 10,000 luciferase-positive T47D metastatic breast cancer stem cells mixed with NME7AB to a final concentration of 32 nM and then mixed 1:1 vol:vol with Matrigel. Tumor engraftment was allowed to progress from day 0 to day 6. The animals were then treated intravenously with anti-NME7AB antibody via tail vein injection. Control animals were injected with PBS. On day 38, the animals were sacrificed, and their livers were harvested and analyzed by IVIS to detect cancer cells that had metastasized to the liver. Figures 46A and 46B show whole-body IVIS photographs of a control animal injected with PBS only. Figures 46C and 46D show whole-body IVIS photographs of a control animal injected with anti-NME7AB antibody 5A1. Figures 46E and 46F show whole-body IVIS photographs of a control animal injected with anti-NME7AB antibody 4A3. Figures 46G and 46H show whole-body IVIS photographs of control animals injected with anti-NME7AB antibody 5D4. Figures 46A, 46C, 46E, and 46G are IVIS photographs taken 7 days before treatment. Figures 46B, 46D, 46F, and 46H are IVIS photographs taken 31 days after anti-NME7AB antibody or mock treatment. As can be seen from the figures, animals in the PBS control group showed metastases (blue dots) in whole-body IVIS photographs, while animals treated with anti-NME7AB antibody showed no metastases. Figures 46I-46P show photographs and IVIS photographs of the liver and lungs taken from animals after sacrifice. Figures 46I, 46K, 46M, and 46O are normal photographs. Figures 46J, 46L, 46N, and 46P are IVIS photographs revealing metastatic cancer cells there. As can be seen from the figures, anti-NME7AB antibody significantly suppressed metastasis to the liver. The liver is a primary site of breast cancer metastasis. Figure 46Q is a bar graph of measured photons emitted and counted by an IVIS counter in livers taken from control versus treated animals. [Figure 47]Photographs of immunofluorescence experiments in which various cancer cell lines were stained to detect the presence of NME7AB are shown. Figure 47A shows T47D breast cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47B shows ZR-75-1 breast cancer cells (also known as 1500s) stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47C shows H1975 non-small cell lung cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47D shows H292 non-small cell lung cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47E shows HPAFII pancreatic cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47F shows DU145 prostate cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. As can be seen from the figures, all cancer cell lines we tested exhibit strong membranous staining for NME7AB. The monoclonal antibody used in these experiments was 5D4. In parallel, the same cell lines were stained with NME7AB antibodies 5A1 and 4A3 with identical results. [Figure 48] Photographs of immunofluorescence experiments in which various human lung cancer cell lines were stained to detect the presence of NME7AB are shown. Figures 48A-48C show H1975 non-small cell lung cancer cells, an adenocarcinoma, stained with various concentrations of the anti-NME7AB antibody 5D4. Figure 48A is an overlay of DAPI and anti-NME7AB staining. Figure 48B shows anti-NME7AB staining alone. Figure 48C is a magnified view of the overlay of DAPI and anti-NME7AB staining. Figures 48D-48F show H292 non-small cell lung cancer cells, a pulmonary mucoepidermoid carcinoma, stained with various concentrations of the anti-NME7AB antibody 5D4. Figure 48D is an overlay of DAPI and anti-NME7AB staining. Figure 48E shows anti-NME7AB staining alone. Figure 48F is a magnified view of the overlay of DAPI and anti-NME7AB staining. Figures 48G-48I show metastatic bronchoalveolar carcinoma, H358 non-small cell lung cancer cells, stained with various concentrations of anti-NME7AB antibody 5D4. Figure 48G is an overlay of DAPI and anti-NME7AB staining. Figure 48H shows anti-NME7AB staining alone. Figure 48I is a magnified view of the overlay of DAPI and anti-NME7AB staining. [Figure 49A-F]Figure 49 shows PCR graphs of cancer cell lines: breast cancer T47D, lung H1975, lung H358, and pancreatic HPAFII before and after culture in NME7AB. Figure 49A shows the breast cancer metastasis marker CXCR4 measured. Figure 49B shows the stem cell marker OCT4 measured. Figure 49C shows the metastasis marker ALDH1 measured. Figure 49D shows the stem cell marker SOX2 measured. Figure 49E shows the stem cell marker NANOG measured. Figure 49F shows the metastasis marker CDH1 (also known as E-cadherin) measured. Figure 49G shows the metastasis marker CD133 measured. Figure 49H shows the stem cell marker ZEB2 measured. Figure 49I shows the stem, cancer, and metastasis marker MUC1 measured. Floating cells (also known as tumorspheres) are able to grow independently of the scaffold and exhibit increased metastasis markers compared to adherent cells. Animals injected with cancer stem cells were treated with NME7AB-expanded suspension cells. As can be seen, markers of metastasis, stem cells, or epithelial-mesenchymal transition (EMT) increased after culture in NME7AB, indicating a shift to a more metastatic state. [Figure 49G-I]Figure 49 shows PCR graphs of cancer cell lines: breast cancer T47D, lung H1975, lung H358, and pancreatic HPAFII before and after culture in NME7AB. Figure 49A shows the breast cancer metastasis marker CXCR4 measured. Figure 49B shows the stem cell marker OCT4 measured. Figure 49C shows the metastasis marker ALDH1 measured. Figure 49D shows the stem cell marker SOX2 measured. Figure 49E shows the stem cell marker NANOG measured. Figure 49F shows the metastasis marker CDH1 (also known as E-cadherin) measured. Figure 49G shows the metastasis marker CD133 measured. Figure 49H shows the stem cell marker ZEB2 measured. Figure 49I shows the stem, cancer, and metastasis marker MUC1 measured. Floating cells (also known as tumorspheres) are able to grow independently of the scaffold and exhibit increased metastasis markers compared to adherent cells. Animals injected with cancer stem cells were treated with NME7AB-expanded suspension cells. As can be seen, markers of metastasis, stem cells, or epithelial-mesenchymal transition (EMT) increased after culture in NME7AB, indicating a shift to a more metastatic state. [Figure 50] Figures 50A and 50C show IVIS photographs of NSG mice injected with 10,000 cancer cells into the tail vein, either parental NCI-H358 cells or NCI-H358 cells, after 10 days in culture with NME7AB. Figures 50A and 50C show IVIS photographs of mice injected with NCI-H358 lung cancer cells grown in NME7AB for 10 days. Figures 50B and 50D show IVIS photographs of mice injected with parental NCI-H358 cells. Figures 50A and 50B show IVIS photographs of mice imaged facing downward. Figures 50C and 50D show IVIS photographs of mice imaged facing upward. As can be seen, NME7AB-expanded cells significantly enhanced metastatic potential. [Figure 51]PCR graphs of the MUC1-negative prostate cancer line PC3 before and after two or three passages of culture in dimeric NM23-H1 (also known as NME1) or NME7AB are shown. The graphs show the fold difference in stem cell, cancer cell, and metastatic markers. As can be seen, repeated culture in NME1 or NME7AB induces upregulation of stem, cancer, and metastatic markers, but also upregulates MUC1 expression by 5-8 fold. DETAILED DESCRIPTION OF THE INVENTION

[0040] definition In this application, "a" and "an" are used to refer to both singular and plural objects.

[0041] As used herein, "about" or "substantially" generally provides leeway from being limited to an exact number. For example, when used in the context of the length of a polypeptide sequence, "about" or "essentially" indicates that the polypeptide is not limited to the stated number of amino acids. A small number of amino acids added or removed from the N-terminus or C-terminus may be included as long as functional activity, such as binding activity, exists.

[0042] As used herein, administration "in combination with" one or more further therapeutic agents includes simultaneous (concomitant) and consecutive administration in any order.

[0043] As used herein, "amino acid" and "amino acids" all refer to naturally occurring L-α-amino acids. This definition is intended to include norleucine, ornithine, and homocysteine.

[0044] As used herein, the term "amino acid sequence variant" generally refers to molecules that have some differences in their amino acid sequence compared to a reference (e.g., native sequence) polypeptide. The amino acid changes can be substitutions, insertions, deletions, or any desired combination of such changes in the native amino acid sequence.

[0045] Substitutional variants are those that have at least one amino acid residue in a native sequence removed and a different amino acid inserted in its place in the same position. The substitutions may be single, where only one amino acid in the molecule has been substituted, or they may be multiple, where more than one amino acid has been substituted in the same molecule.

[0046] Substitutes for an amino acid within the sequence may be selected from other members of the class to which the amino acid belongs. For example, nonpolar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, and methionine. Polar-neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine. Positively charged (basic) amino acids include arginine, lysine, and histidine. Negatively charged (acidic) amino acids include aspartic acid and glutamic acid. Also included within the scope of the invention are proteins or fragments or derivatives thereof that exhibit the same or similar biological activity, and derivatives that are differentially modified during or after translation, for example, by glycosylation, proteolytic cleavage, conjugation to antibody molecules or other cellular ligands, etc.

[0047] Insertional variants are those that have one or more amino acids inserted immediately adjacent to an amino acid at a particular position in the native amino acid sequence, where immediately adjacent to an amino acid means attached to either the α-carboxy or α-amino functionality of that amino acid.

[0048] Deletion variants have one or more amino acids removed from the native amino acid sequence. Typically, deletion variants have one or two amino acids deleted in a particular region of the molecule.

[0049] As used herein, "fragment" or "functional derivative" refers to biologically active amino acid sequence variants and fragments of the polypeptides of the invention, as well as derivatives obtained by reaction with organic derivatizing agents, post-translational modifications, derivatives containing non-proteinaceous polymers, and covalent modifications, including immunoadhesins.

[0050] As used herein, "carrier" includes pharmaceutically acceptable carriers, excipients, or stabilizers that are nontoxic to cells or mammals exposed thereto at the dosages and concentrations employed. Often, pharmaceutically acceptable carriers are pH-buffered aqueous solutions. Examples of pharmaceutically acceptable carriers include, but are not limited to, buffers such as phosphate, citric acid, and other organic acids; antioxidants, including ascorbic acid; low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum, albumin, gelatin, or immunoglobulins; hydrophilic polymers, such as polyvinylpyrrolidone; amino acids, such as glycine, glutamine, asparagine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrins; chelating agents, such as EDTA; sugar alcohols, such as mannitol or sorbitol; salt-forming counterions, such as sodium; and / or nonionic surfactants, such as TWEEN®, polyethylene glycol (PEG), and PLURONICS®.

[0051] As used herein, "pharmaceutically acceptable carriers and / or diluents" include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. The use of such media and agents for pharmaceutical active substances is known in the art. Except insofar as any conventional media or agent is incompatible with the active ingredient, its use in the therapeutic compositions is contemplated. Supplementary active ingredients can also be incorporated into the compositions.

[0052] For ease of administration and uniformity of dosage, it is particularly advantageous to formulate parenteral compositions in dosage unit form.As used herein, dosage unit form refers to a physically separate unit that is suitable as a unit dose for the subject to be treated, and each unit contains a predetermined amount of active substance calculated to produce desired therapeutic effect, and is incorporated into necessary pharmaceutical carrier.The specification for dosage unit form of the present invention is determined by and directly depends on (a) the specific characteristics of active substance and the specific therapeutic effect to be achieved, and (b) the inherent limitations in pharmaceutical technology, such as active substance for treating disease in living organisms with pathological conditions that cause physical health to be impaired.

[0053] The main active ingredient is formulated in an effective amount with a suitable pharmaceutically acceptable carrier in a dosage unit form for convenient and effective administration. The dosage unit form contains, for example, an amount of the main active ingredient ranging from 0.5 μg to about 2000 mg. Expressed as a percentage, the active compound is usually present in the carrier at about 0.5 μg / ml. In the case of compositions containing supplementary active ingredients, the dosage is determined by reference to the usual dose and method of administration of the ingredients.

[0054] As used herein, the terms "vector," "polynucleotide vector," "construct," and "polynucleotide construct" are used interchangeably. The polynucleotide vectors of the present invention can be in any of several forms, including, but not limited to, RNA, DNA, RNA encapsulated in a retroviral coat, DNA encapsulated in an adenoviral coat, DNA packaged in another virus or virus-like form (such as herpes simplex, and adenovirus structures such as polynucleotides).

[0055] As used herein, a "host cell" includes an individual cell or cell culture that can be or has been a recipient of a vector of the invention. A host cell includes the progeny of a single host cell, and the progeny may not necessarily be completely identical (either morphologically or in total DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation and / or alteration.

[0056] As used herein, a "subject" is a vertebrate, preferably a mammal, more preferably a human.

[0057] As used herein, "mammal" for purposes of treatment refers to any animal classified as a mammal, including humans, farm animals, and zoo, sport or pet animals, dogs, cats, cows, horses, sheep, pigs, etc. Preferably, the mammal is a human.

[0058] As used herein, "treatment" refers to an approach for obtaining beneficial or desired clinical results. For purposes of the present invention, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, reduction in the extent of disease, stable disease (i.e., not worsening), delay or slowing of disease progression, improvement or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. "Treatment" can also mean prolonging survival as compared to expected survival in the absence of treatment. "Treatment" refers to both therapeutic and prophylactic or preventative treatment. Those in need of treatment include those already suffering from the disorder, as well as those in whom the disorder is to be prevented. "Alleviating" a disease means that the extent of the disease state and / or undesirable clinical symptoms are reduced and / or the time course of progression is slowed or prolonged as compared to the absence of treatment.

[0059] As used herein, the "A1" peptide, the "A2" peptide, the "B1" peptide, the "B2" peptide, and the "B3" peptide are peptides derived from human NME7. AB These antibodies refer to peptides that bind to human NME1 but do not bind (or bind very weakly) to human NME1. AB and NME7-X1, and are defined as follows: A1 is NME7A peptide 1 (A domain): MLSRKEALDFHVDHQS (SEQ ID NO: 141) A2 is NME7A peptide 2 (A domain): SGVARTDASES (SEQ ID NO: 142) B1 is NME7B peptide 1 (B domain): DAGFEISAMQMFNMDRVNVE (SEQ ID NO: 143) B2 is NME7B peptide 2 (B domain): EVYKGVVTEYHDMVTE (SEQ ID NO: 144) B3 is NME7B peptide 3 (B domain): AIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 145) Further, for clarity, NME7A (with a capital "A") refers to the subunit A portion of NME7; NME7a (with a lower case "a") refers to the full-length NME7 described elsewhere herein; and NME7B (with a capital "B") refers to the subunit B portion of NME7; and NME7b (with a lower case "b") refers to an NME7 species lacking a portion of the DM10 region, which is described elsewhere herein.

[0060] As used herein, the term "antibody-like" refers to a molecule that contains a portion of an antibody but can be engineered to contain a portion of an antibody that is not a naturally occurring antibody. Examples include, but are not limited to, CAR (chimeric antigen receptor) T cell technology and Ylanthia® technology. CAR technology uses an antibody epitope fused to a portion of a T cell so that the body's immune system is directed to attack a specific target protein or cell. Ylanthia® technology consists of an "antibody-like" library, a collection of synthetic human Fabs that are screened to bind to peptide epitopes derived from target proteins. Selected Fab regions can then be engineered into a scaffold or framework so that they resemble antibodies.

[0061] As used herein, "an effective amount of an agent to inhibit an NME family protein" refers to an amount of an agent effective in preventing the activating interaction between an NME family protein and its cognate receptor.

[0062] As used herein, "NME-derived fragment" refers to a fragment of NME or a peptide sequence that is highly homologous to a peptide sequence that is a fragment of NME.

[0063] As used herein, "MUC1 *The extracellular domain is primarily defined by the PSMGFR sequence (GTINVHDVETQFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 6)). The exact site of MUC1 cleavage depends on the enzyme that excises it, and since the cleavage enzyme varies depending on cell type, tissue type, or time in cellular evolution, MUC1 * The exact sequence of the extracellular domain may vary at the N-terminus.

[0064] As used herein, the term "PSMGFR" is an acronym for the primary sequence of the MUC1 growth factor receptor, defined as follows: GTINVHDVETQFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 6). In this regard, the "N-number" such as "N-10 PSMGFR" or simply "N-10," "N-15 PSMGFR" or simply "N-15," or "N-20 PSMGFR" or simply "N-20" refers to the number of amino acid residues deleted at the N-terminus of PSMGFR. Similarly, the "C-number" such as "C-10 PSMGFR" or simply "C-10," "C-15 PSMGFR" or simply "C-15," or "C-20 PSMGFR" or simply "C-20" refers to the number of amino acid residues deleted at the C-terminus of PSMGFR. Mixtures of deletions and additions are also possible. For example, N+20 / C-27 is a wild-type MUC1 protein with 20 amino acids added at the N-terminus of PSMGFR and 27 amino acids deleted from the C-terminus. * It refers to a peptide fragment of

[0065] As used herein, "MUC1 * The "extracellular domain of MUC1" refers to the extracellular portion of the MUC1 protein that lacks the tandem repeat domain. * is the cleavage product, and its MUC1 * The fragment consists of a short extracellular domain lacking tandem repeats, a transmembrane domain, and a cytoplasmic tail. The exact location of the cleavage site in MUC1 is unknown, possibly because it appears to be cleavable by more than one enzyme.* The extracellular domain of contains most of the PSMGFR sequence but may have an additional 10-20 N-terminal amino acids.

[0066] As used herein, "high homology" is considered to be at least 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 97% identity in the specified region of overlap between any two polypeptides.

[0067] As used herein, "NME family proteins," or "NME family member proteins," numbered 1-10, are proteins grouped together because they all contain at least one NDPK (nucleotide diphosphate kinase) domain. In some cases, the NDPK domain is not functional in that it can catalyze the conversion of ATP to ADP. NME proteins were previously known as NM23 proteins, numbered H1 and H2. Recently, as many as 10 NME family members have been identified. As used herein, the terms NM23 and NME are interchangeable. As used herein, the terms NME1, NME2, NME5, NME6, NME7, NME8, and NME9 refer to native proteins and NME variants. In some cases, these variants are more soluble, express better in E. coli, or are more soluble than the native sequence proteins. For example, NME7 as used herein has excellent commercial applicability because the variations allow for high-yield expression of soluble, properly folded protein in E. coli. AB It may refer to the naturally occurring protein, or a variant thereof, such as NME7. ABconsists primarily of the NME7 A and B domains but lacks most of the DM10 domain (SEQ ID NO: 39) at the N-terminus of the native protein. As referred to herein, "NME1" is interchangeable with "NM23-H1." It is also intended that the present invention is not limited by the exact sequence of the NME protein. The mutant NME1-S120G, also referred to as NM23-S120G, is used interchangeably throughout this application. The S120G and P96S mutants are preferred due to their propensity for dimer formation, and are referred to herein as NM23 dimers, NME1 dimers, or dimeric NME1 or dimeric NM23.

[0068] NME7, as referred to herein, is intended to mean native NME7, which has a molecular weight of approximately 42 kDa.

[0069] The "NME7 family" includes full-length NME7, as well as naturally occurring or artificially produced truncated forms having molecular weights of about 30 kDa, 33 kDa, or about 25 kDa; NME7b, NME7-X1, NME7 AB "NME7 family" also refers to "NME7 family members" or ... AB These may include "anti-cancer drug-like" proteins, which are proteins in the range of 30-33 kDa that are expressed in cancer cells.

[0070] As used herein, "agents that maintain stem cells in a naive state or revert primed stem cells to a naive state" refers to proteins, small molecules, or nucleic acids that, alone or in combination, maintain stem cells in a naive state that resembles cells of the inner cell mass of an embryo. Examples include, but are not limited to, human NME1 dimers, bacterial, fungal, yeast, viral, or parasitic NME proteins, including human NME proteins, particularly NME1, NME7, NME7-X1, NME7, NME7-X2, NME7-X3, NME7-X4, NME7-X5, NME7-X6, NME7-X7, NME7-X8, NME7-X9, NME7-X10, NME7-X11, NME7-X12, NME7-X13, NME7-X14, NME7-X15, NME7-X21, NME7-X3, NME7-X4, NME7-X16, NME7-X17, NME7-X18, NME7-X19, NME7-X21, NME7-X22, NME7-X31, NME7-X19, NME7-X19, NME7-X21 ...19, NME7-X21, NME7-X19, NME7-X21, NME7-X31, NME7-X19 AB These include nucleic acids such as those with high sequence identity to NME6, 2i (Silva J et al., 2008; Hanna et al., 2010), 5i (Theunissen TW et al., 2014), and siRNAs that suppress the expression of MBD3, CHD4 (Rais Y1 et al., 2013), BRD4, or JMJD6 (Liu W et al., 2013).

[0071] As used herein, the term "NME7 AB ", "NME7AB" and "MNE-AB" are used interchangeably.

[0072] As used herein, "agents that promote pluripotency" or "agents that revert somatic cells to a stem-like or cancer-like state" refer to proteins, small molecules, or nucleic acids that, alone or in combination, induce or suppress the expression of specific genes such that the gene signature changes to one that more closely resembles a stem cell or cancer cell. Examples include, but are not limited to, NME1 dimer, NME7, NME7-X1, NME7 AB , 2i, 5i; nucleic acids such as siRNA that suppress the expression of MBD3 or CHD4 or BRD4 or JMJD6; microbial NME proteins that have high sequence homology to human NME1, NME2, NME5, NME6, NME7, NME8 or NME9, preferably with a region containing the NDPK domain.

[0073] As used herein, with respect to an agent referred to as a "small molecule," it may be a synthetic chemical molecule or a chemical-based molecule having a molecular weight between 50 Da and 2000 Da, more preferably between 150 Da and 1000 Da, and even more preferably between 200 Da and 750 Da.

[0074] As used herein, with respect to an agent that is referred to as a "natural product," it can be a chemical molecule or a biological molecule, so long as the molecule occurs in nature.

[0075] As used herein, FGF, FGF-2 or bFGF refers to fibroblast growth factor (Xu RH et al, 2005; Xu C et al, 2005).

[0076] As used herein, a "Rho-associated kinase inhibitor" can be a small molecule, peptide, or protein (Rath N, et al, 2012). Rho kinase inhibitors are abbreviated here and elsewhere as ROCi, ROCKi, or Ri. The use of specific Rho kinase inhibitors is intended to be exemplary and can be substituted with any other Rho kinase inhibitor.

[0077] As used herein, the term "cancer stem cells" or "tumor-initiating cells" refers to cancer cells that express levels of genes associated with more metastatic or aggressive cancers. "Cancer stem cells" or "tumor-initiating cells" also refer to cancer cells that, when transplanted into an animal, can give rise to tumors with very few cells. Cancer stem cells and tumor-initiating cells are often resistant to chemotherapeutic drugs.

[0078] As used herein, the terms "stem / cancer," "cancer-like," and "stem-like" refer to a state in which cells have acquired the properties of stem cells or cancer cells or share important elements of the gene expression characteristics of stem cells, cancer cells, or cancer stem cells. Stem-like cells can be somatic cells that have undergone induction toward a less mature state, such as increased expression of pluripotency genes. Stem-like cells also refer to cells that have undergone some degree of dedifferentiation or are in a metastable state in which they can transform into terminal differentiation. Cancer-like cells can be cancer cells that have not yet been fully characterized but that exhibit the morphology and characteristics of cancer cells, such as those that can grow anchorage-independently or give rise to tumors in animals.

[0079] As used herein, "spacers," or "linkers," of different lengths can be incorporated anywhere in the peptide. Spacer attachment is usually via an amide bond, although other functional groups are possible.

[0080] NME, NME7 and NME7 protein family We found that NME7 and NME7-X1 are highly expressed in early human stem cells and most cancer cells (Figures 17, 18, 19A-19F, 22, 23, 39, 40, 41, 47, and 48). Figure 17 shows a graph of RT-PCR measurement of NME7-X1 expression in a series of human stem cells and cancer cells. Figure 18 shows a graph of RT-PCR measurement of NME7, NME7a, NME7b, and NME7-X1 expression in a series of human stem cells and cancer cells. NME7a is full-length NME7, NME7b lacks a small portion of the DM10 domain, and NME7-X1 lacks all of the DM10 domain and a small portion of the N-terminus of the first NDPK A domain. The bar labeled NME7 indicates that primers detecting both NME7a and NME7b were used. Figures 19A-19F show photographs of Western blots in which expression of NME7 species in various cancer cell lines was detected using antibodies generated by immunization with short NME7-derived peptides. Figure 19A shows a Western blot detected with antibody #52 of the present invention, which binds to NME7-derived peptide A1. Figure 19B shows a Western blot detected with antibody #56 of the present invention, which binds to NME7-derived peptide B1. Figure 19C shows a Western blot detected with antibody #61 of the present invention, which binds to NME7-derived peptide B3. Figures 22A-22E show photographs of Western blots from coimmunoprecipitation experiments. T47D breast cancer cell extracts were incubated with Ab-5, an antibody against the cytoplasmic tail of MUC1, or a control antibody, IgG, and coimmunoprecipitated. Gels were blotted with two different commercially available anti-NME7 antibodies, B9 (Figure 22A) and CF7 (Figure 22B). Both gels show distinctive NME7 bands at approximately 33 kDa and 30 kDa. The gels were stripped and MUC1 * This was reprobed with anti-PSMGFR (Figure 22C and Figure 22D), an antibody against the extracellular domain of NME7 and MUC1. * Recombinant NME7 interacts with ABand recombinant NME7-X1 were mixed together, run on a gel, and then detected with an anti-NME7 antibody. * Two unique NME7 species interact with NME7 AB Figures 23A-23C show Western blot photographs of co-immunoprecipitation experiments. Human induced pluripotent stem cell (iPS7) cell extracts or embryonic stem cell (HES3) cell extracts were incubated with Ab-5, an antibody against the cytoplasmic tail of MUC1, or a control antibody, IgG, and co-immunoprecipitated. The gel was blotted with the commercially available anti-NME7 antibody B9 (Figure 23A). Both cell types show distinctive NME7 bands at approximately 33 kDa and 30 kDa. The gel was stripped and analyzed for MUC1. * This was reprobed with anti-PSMGFR (Figure 23B), an antibody against the extracellular domain of NME7 and MUC1. * Recombinant NME7-AB and recombinant NME7-X1 were mixed together, run on a gel, and then detected with an anti-NME7 antibody. * Figures 39A-39C show human lung tissue specimens stained with an anti-NME7 antibody that binds to the B3 peptide. This figure demonstrates the lack of NME7 expression in normal tissues, the increasing expression of NME7 with tumor grade, and the increased metastasis. Figures 40A-40C show human small intestine tissue specimens stained with an anti-NME7 antibody that binds to the B3 peptide. This figure demonstrates the lack of NME7 expression in normal tissues, the increasing expression of NME7 with tumor grade, and the increased metastasis. Figures 41A-41D show human colon tissue specimens stained with an anti-NME7 antibody that binds to the B3 peptide. This figure demonstrates the lack of NME7 expression in normal tissues, the increasing expression of NME7 with tumor grade, and the increased metastasis. Figures 47 and 48 show immunofluorescence photographs showing that NME7 is secreted by various cancer cell lines and binds to extracellular receptors on those cancer cell lines.

[0081] Furthermore, we demonstrate that, like NM23-H1, NME7 mediates the upregulation of MUC1 in both stem cells and cancer cells. * It has been demonstrated that they bind to and dimerize growth factor receptors. Figure 5 shows a sequence alignment of the NME1 and NME7 A and B domains.

[0082] We recently found that NME7 is a pristine form of NME1 (NM23-H1) expressed in very early embryonic stem cells. NME7 is either not expressed at all or expressed at very low levels in adult tissues. However, we found that NME7 is expressed at high levels in cancer cells and tissues, and even higher levels in metastatic cancer cells and tissues. A truncated form of NME7 may be a secreted form, allowing it to bind to and activate extracellular receptors. We detected full-length NME7, MW 42 kDa, as well as NME7 species of approximately 33 kDa and 30 kDa. The 33 kDa and 30 kDa species are secreted from cancer cells. Western blots detect full-length NME7 in cell lysates but smaller 30-33 kDa NME7 species in their conditioned medium. Western blots probed with either an antibody recognizing NME7 or an antibody recognizing only the DM10 domain indicate that the lower molecular weight NME7 species secreted into the conditioned medium lacks the DM10 domain. These data support the idea that the naturally occurring NME7 species have approximately the same molecular weight as those of the recombinant NME7 we generated. AB Consistent with the idea that they are comparable, both are secreted and both lack the 91 amino acids of the DM10 domain that may keep the protein intracellular.

[0083] We discovered a new NME7 isoform, NME7-X1, which is overexpressed in stem cells and cancer cells, particularly in prostate cancer (Figures 17, 18, 19, and 22). NME7-X1, with a molecular weight of approximately 30 kDa, contains amino acids 125-376 of NME7, whereas our recombinant NME7 ABNME7b, which has a molecular weight of approximately 33 kDa, spans amino acids 92-376 and therefore contains 33 more N-terminal amino acids. NME7b spans amino acids 37-376 and lacks only 37 amino acids of the DM10 domain, and is overexpressed in prostate cancer (Figure 18). We generated human recombinant NME7-X1, which is a truncated product or alternative isoform of the naturally occurring NME7. AB This indicates that the secreted NME7 species in cancer cells is 30 kDa, which has a molecular weight slightly smaller than the naturally occurring NME7 species of approximately 33 kDa, which is thought to be a "like" protein.

[0084] We tested a series of cancer cell lines and found that they expressed NME7 AB NME7-like proteins AB We found that IL-16 expresses high levels of NME7 and lower molecular weight species that can be cleaved, similar to alternative isoforms such as NME7-X1 or NME7-X1.

[0085] NM23-H1 (also known as NME1) must be a dimer, whereas NME7 must be a dimer. * It is a monomer with two binding domains in the extracellular domain. We generated recombinant human NME7 lacking the DM10 domain and characterized it as NME7. AB The sandwich ELISA binding assay was performed using recombinant NME7 AB However, MUC1 * We demonstrate that the extracellular domain simultaneously binds two PSMGFR peptides containing most or all of the PSMGFR sequence (Figure 1). In nanoparticle binding assays, NME7 binds to MUC1 * It has also been shown to be able to bind to the PSMGFR portion of the extracellular domain and dimerize it.

[0086] Agents that neutralize NME7, block its interaction with its binding partners, or suppress its expression are potential anti-cancer therapeutics. Such agents can be antibodies, small molecules, or nucleic acids. They can act directly on NME7, on molecules that regulate NME7 expression, or on enzymes that cleave NME7 into cancer-promoting forms.

[0087] We used recombinant NME7, as well as NM23-H1 dimers. AB We found that the NME7 and MUC1 monomers, which contain essentially the PSMGFR sequence, were fully capable of supporting pluripotent human stem cell proliferation in the absence of other growth factors, cytokines, or serum. * Competitive inhibition of the interaction between the extracellular domains of NME7 and MUC1 induces stem cell differentiation, and promotes stem cell proliferation and inhibits differentiation. * This shows that the interaction is

[0088] Next, we will also be looking at NME7 AB NME7 alone was shown to be fully capable of supporting human cancer cell growth. AB When added to normal cancer cell growth medium, it stimulates cancer cell proliferation, especially MUC1-positive and MUC1 * -Stimulated the proliferation of MUC1 positive cancer cells. * Inhibition of the interaction between MUC1 and NME7 suppressed cancer cell proliferation. * MUC1 by Fab * Blockade of growth factor receptors potently inhibited cancer cell proliferation. Similarly, antibodies that bind to NME7 inhibited cancer cell proliferation. In one example of cancer growth inhibition by anti-NME7 antibodies, polyclonal antibodies were generated by immunizing animals with a portion of NME7 ranging from amino acids 100 to 376 (Figures 12 and 13). However, we found that NME7 AB They found that antibodies generated from immunization with shorter peptides from NME7-X1 or from NME7-X1 also inhibited cancer growth. In particular, they found that antibodies against MUC1 and MUC1 * Inhibits the growth of NME7-positive cancers. Anti-NME7 antibodies of the invention inhibited the formation of non-adherent "floating" cells that can migrate from primary tumors and form metastatic tumorspheres (Figures 14, 16, 29). Anti-NME7 antibodies of the invention inhibited the upregulation of metastatic and stem cell markers, which are now also believed to be characteristic of metastasis (Figures 15, 30, 31, 32).

[0089] NME7 induces cancer metastasis The inventor is NME7 AB We further found that culturing cancer cells in minimal medium containing NME7 induces various cancer cells to transform into a more metastatic state. Evidence of this induced metastatic state includes a change from adherent to non-adherent, or "suspension," cell growth, accompanied by the upregulation of certain metastatic markers, particularly those upregulated in suspension cells. AB These metastatic markers that are upregulated after culture in NME7 include, but are not limited to, CXCR4, CHD1 (also known as E-cadherin), MUC1, ALDH1, CD44, and pluripotent stem cell markers such as OCT4, SOX2, NANOG, KLF2 / 4, FOXa2, TBX3, ZEB2, and c-Myc (Figures 2, 3, 20, 49, 51). AB Cancer cells cultured in NME7 have an extremely high engraftment rate of over 90% when xenografted into test animals. Furthermore, a very small number of transplanted cancer cells formed tumors in the test animals, which is consistent with the results of NME7. AB This is evidence that NME7 transforms them into cancer stem cells, known as metastatic cancer cells. AB Cancer cells cultured in estrogen-releasing pellets and injected into NOD / SCID / GAMMA mice metastasized in the animals at lower cell counts than parental cells grown in normal medium (Figures 33-38). AB The methods described herein generate alternative isoforms essentially equivalent to NME7. AB The use of recombinant NME7 is not limited to this, and other NME7 species may function similarly. For example, we have found that another NME7 isoform, NME7-X1, is expressed by cancer cells. It is similar to our recombinant NME7 except that the X1 isoform lacks 33 amino acids from the N-terminus. AB The NME7-X1 is the same as the NME7 AB It is predicted to work like this. AB A "like" protein was also detected in cancer cells as an approximately 33 Da species.

[0090] In our previous work, we AB We have shown that NME7 alone can revert human stem cells to a previous, naive state. We have found that culturing cancer cells in the presence of other agents that revert stem cells to a more naive state transforms the cancer cells to a more metastatic state. AB We demonstrated that culturing cancer cells in NME1 (Figure 2), NME1 dimers (Figure 3), or "2i" inhibitors (Figure 4) can transform normal cancer cells into metastatic cancer cells, termed cancer stem cells "CSCs" or tumor-initiating cells "TICs," respectively. However, NME7 AB induced cancer cells to enter a more metastatic state than NME1, also known as NM23-H1, which was better than 2i.

[0091] 2i is the name given to two biochemical inhibitors that researchers found reverted human stem cells to a more naive state. 2i are MEK and GSK3 beta inhibitors, PD0325901 and CHIR99021, which are added to the culture medium at final concentrations of approximately 1 mM and 3 mM, respectively. NME7 AB NME7-X1 and NME7-X2 function well at lower and higher concentrations, ranging from approximately 1 nM to 16 nM, but are used at a final concentration of approximately 4 nM when added to separate batches of minimal medium to transform cancer cells into metastatic cells. Human or bacterial NME1 dimers are used at final concentrations ranging from 4 nM to 32 nM, typically at 16 nM in these experiments, with human NME carrying the S120G mutation. Lower concentrations are necessary when using wild-type NME1. These exact concentrations are not intended to be critical. While certain mutations allow NME1 to exist as a dimer at higher concentrations, it is important that the NME1 protein is dimeric, and the concentration range at which this occurs is in the low nanomolar range. Similarly, the concentration of NME7 protein can be varied. NME7 AB and NME7-X1 are monomers, and the concentrations used to transform cancer cells into metastatic cells must allow the proteins to remain as monomers.

[0092] NME7, NME7 AB In addition to NME7-X1 and 2i inhibitors, MEKi, and GSK3i, other agents and inhibitors have been shown to revert stem cells to a more naive state. These inhibitors include JNKi, p38i, PKCi, ROCKi, BMPi, BRAFi, SRCi, and the growth factors activin and LIF (Gafni et al., 2013, Chan et al., 2013, Valamehr et al., 2014, Ware et al., 2014, Theunissen et al., 2014). These agents can also be used to advance cancer cells to a more metastatic state. Cells induced to transform into a more metastatic state using inhibitors or growth factors that revert stem cells to a more naive state, alone or in combination, can then be used as discovery tools to identify or test drugs to treat or prevent cancer metastasis.

[0093] Various molecular markers have been proposed as indicators of metastatic cancer cells. Different cancer types may have different upregulated molecules. For example, the receptor CXCR4 is upregulated in metastatic breast cancer, while E-cadherin, also known as CHD1, is more upregulated in metastatic prostate cancer. In addition to these specific metastatic markers, typical markers of pluripotency, such as OCT4, SOX2, NANOG, and KLF4, are upregulated as cancers become metastatic. Starting cancer cells and later metastatic cancer cells are assayed by PCR to measure the expression levels of these genes. We have found that NME7, which transforms cancer cells to a more metastatic state, as evidenced by increased expression of metastatic and pluripotent stem cell markers. AB demonstrated that these cancer cells cultured in agents such as EGFR-1, EGFR-2, and EGFR-3 function as metastatic cancer cells.

[0094] A functional test for whether a cancer cell population is metastatic is to inject a very small number of cells, e.g., 200, into immunodeficient mice and see whether they develop into tumors. Typically, 5-6 million cancer cells are required to form tumors in immunodeficient mice. We have shown that as few as 50 NME-induced metastatic cancer cells formed tumors in mice. Furthermore, human NME7 AB Mice injected with NME1 or NME7-X1 during the study period developed distant metastases.

[0095] In one particular experiment, T47D human breast cancer cells were cultured in standard RPMI medium for 14 days, with medium changes every 48 hours, and subjected to trypsinization at approximately 75% confluence. Cells were then seeded into 6-well plates and treated with 4 nM NME7. AB The cells were cultured in minimal stem cell medium (see Example 1) supplemented with CXCR4. The medium was changed every 48 hours. By about day 4, some cells detach from the surface and float. The medium was carefully changed to retain the "floaters," as these are the cells with the highest metastatic potential as revealed by RT-PCR measurement of metastatic markers. On day 7 or 8, the floaters were collected and counted. Samples were retained for RT-PCR measurement. The key markers measured were CXCR4, which is a marker associated with NME7. AB It is upregulated 40-200 fold after short incubation in

[0096] Freshly collected floating metastatic cells were xenografted into the flanks of female nu / nu athymic mice implanted with a 90-day sustained-release estrogen pellet. The floating cells were xenografted at 10,000, 1,000, 100, or 50 cells, respectively. Half of the mice in each group of six were also implanted with 32 nM NME7 near the original implantation site. AB was injected daily. ABParental T47D cells cultured in RPMI medium without NME7 were also implanted into mice at 6 million, 10,000, or 100 cells as controls. Mice implanted with NME7-induced floating cells developed tumors even when only 50 cells were implanted. AB Mice receiving daily injections of NME7 also developed distant tumors or distant metastases in various organs. AB Human NME7 after transplantation of cultured cancer cells AB Eleven of the 12 mice injected with human NME7, or 92%, developed tumors at the injection site. AB Only 7 of the 12 mice, or 58%, that were not injected with human NME7 developed tumors. AB Nine of 11 mice (82%) injected with NME7 developed multiple tumors distant from the injection site. AB None of the mice that were not injected developed multiple visible tumors.

[0097] After sacrifice, RT-PCR and Western blot analysis revealed that NME7 AB The distant tumors in the mice injected with 6 million cells indicated that they were indeed human breast tumors. Similar analysis of their organs showed that in addition to the distant tumors, the mice also had random metastases to the liver and lungs, with the human breast cancer characteristics of the implanted human breast cancer cells. As expected, only the mice injected with 6 million cells grew tumors.

[0098] We used human recombinant NME7 AB We showed that the size and sequence of NME7-X1 are comparable to those of NME7-X1 and the 30-33 kDa NME7 cleavage products. ABWe have shown that NME7-X1 promotes cancerous growth and accelerates cancer cells to a highly metastatic cancer stem cell (CSC) state, also called tumor-initiating cells (TICs). Therefore, we conclude that NME7-X1 and NME7 truncation products lacking the DM10 domain also promote cancerous growth and accelerate cancer cells to a highly metastatic cancer stem cell (CSC) state, also called tumor-initiating cells (TICs). In one example, NME7 AB was added to cancer cells in serum-free medium and in the absence of any other growth factors or cytokines. Within 7-10 days, the cancer cells reverted to highly metastatic CSCs / TICs, as evidenced by a greater than 100-fold increase in the expression of molecular markers such as CXCR4, an indicator of metastatic cancer cells. In one example, T47D breast cancer cells were cultured in standard RPMI medium or in the presence of recombinant NME7 at a final concentration of 16 nM. AB The cells were cultured in minimal stem cell medium (Example 1) supplemented with NME7. After 10 days, the cells were collected and analyzed by RT-PCR for expression of molecular markers of CSCs, which was increased by 10-200 fold (Figure 2). This is a specific, detailed example of how we transformed one cancer cell type to a more metastatic state. The invention is not intended to be limited by these details, as there is a range of cancer cells that can be transformed in this manner, a range of agents that revert stem cells to a more naive state and further promote cancer cells to a more metastatic state, and a range of concentrations at which the added agents transform cancer cells. Other types of cancer cells have been shown to be more sensitive to NME7 due to the dramatic increase in expression of metastatic markers and the ability to form tumors from very low numbers of transplanted cancer cells. AB For example, NME7 required a longer incubation period. AB Prostate cancer cells cultured in 2i, human NME1, or bacterial NME1 with high homology to human NME1 or human NME7 showed a dramatic increase in metastatic markers after 2-3 passages.

[0099] The metastasis marker CXCR4 was specifically elevated in metastatic breast cancer cells, while CHD1 was specifically elevated in metastatic prostate cancer. Here, we show that pluripotent stem cell markers such as OCT4, SOX2, NANOG, KLF2 / 4, and TBX3 are also upregulated when cancer cells transform into more metastatic cells.

[0100] DU145 prostate cancer cells were similarly cultured and treated with NME7 AB These cells cultured in NME7 also showed a dramatic increase in CSC marker expression (Figure 3). In prostate cancer cells, CHD1 (also known as E-cadherin) and CXCR4 were upregulated by NME7. AB Figures 20A-20C show that the ovarian cancer cell lines SK-OV3, OV-90, and breast cancer cell line MDA-MB all transitioned from adherent to non-adherent suspension cells, and NME7 was upregulated along with other pluripotent stem cell markers compared to control cancer cells not growing in suspension. AB We show that ovarian cancer cells, prostate cancer cells, pancreatic cancer cells, and melanoma cells also increased the expression of metastatic markers after 72 or 144 hours of culture with NME7. AB and transformed into a more metastatic state after only 3 days of culture. Figure 21 shows that breast, ovarian, prostate, pancreatic cancer cells, and melanoma cells express MUC1 and MUC1 * It is shown that the gene expresses

[0101] Here we are, NME7 AB We have also shown that cancer cells expressing recombinant NME7 transform a wide range of cancer cells to a more metastatic state. AB It has a molecular weight of approximately 33 kDa (Figures 17, 18, 19, and 22) and is NME7 AB and NME7, which lacks the DM10 domain, except for the N-terminal 33 amino acids. ABWe demonstrated that MUC1 also expresses an alternative isoform, NME7-X1, 30 kDa, which has the same sequence as MUC1. Co-immunoprecipitation experiments were performed in T47D breast cancer cells, where cell extracts were incubated with an antibody against the cytoplasmic tail of MUC1, Ab-5, or a control antibody, IgG, and co-immunoprecipitated. The immunoprecipitated species were separated by gel electrophoresis. The gels were blotted with two different commercially available anti-NME7 antibodies. Both gels show distinctive NME7 bands at approximately 33 kDa and 30 kDa (Figures 22A and 22B). The gels were stripped and MUC1 was detected. * This was reprobed with anti-PSMGFR, an antibody against the extracellular domain of NME7 and MUC1 (Fig. 22C and Fig. 22D). * The recombinant NME7 we created AB and recombinant NME7-X1 were mixed together, run on a gel, and then detected with an anti-NME7 antibody. * Two unique NME7 species interact with NME7 AB The results show that the co-immunoprecipitation of NME7-X1 is consistent with that of the cytoplasmic tail of MUC1 (Figure 22E). Similar experiments were performed with human stem cells. Figures 23A-23C show Western blot photographs of co-immunoprecipitation experiments. Extracts from human induced pluripotent stem cells (iPS7 cells) or embryonic stem cells (HES3 cells) were incubated with Ab-5, an antibody against the cytoplasmic tail of MUC1, or a control antibody, IgG, and co-immunoprecipitated. The gel was blotted with the commercially available anti-NME7 antibody B9 (Figure 23A). Both cell types show distinctive NME7 bands at approximately 33 kDa and 30 kDa. The gel was stripped and analyzed for MUC1. * This was reprobed with anti-PSMGFR (Figure 23B), an antibody against the extracellular domain of NME7 and MUC1. * The recombinant NME7 we created AB and recombinant NME7-X1 were mixed together, run on a gel, and then detected with an anti-NME7 antibody. * Two unique NME7 species interact with NME7 AB NME7-X1 and NME7-X1 (Figure 23C).AB Since NME7 is a recombinant protein, it is possible that the naturally occurring species contains an extra 1-15 additional amino acids or that the recombinant NME7 AB It is unclear whether it lacks 1 to 15 additional amino acids, but it evolves at the same apparent molecular weight. AB "Like" recombinant NME7 in that it can stimulate cancer cell proliferation, induce the transition of cancer cells to a more metastatic state, and fully support the expansion of pluripotency in human stem cells. AB The present invention contemplates NME7 species that have an apparent molecular weight of approximately 33 kDa and can function as such.

[0102] We conclude that cancer cell lines and populations expressing NME7 and lower molecular weight NME7 species contain some cancer cells that are CSCs or metastatic cancer cells. AB Culturing cells in NME7-X1, NME7-X2, or the lower molecular weight NME7 species can increase the population of cells that become or are metastatic. Figure 19 shows the results of culturing cells in NME7-X1, NME7-X2, or the lower molecular weight NME7 species. AB Figure 21 shows Western blots of a series of cancer cell lines that express NME7-X1 at 30 kDa and 30 kDa. The T47D breast cancer cell line, PC3 and DU145 prostate cancer cell lines, BT-474 breast cancer cell line, CHL-1 and A2058 melanoma cell lines, and CAPAN-2 and PANC-1 pancreatic cancer cell lines all express MUC1, MUC1 * In Figure 21A, BT474 cells express MUC1 or MUC1 * However, we previously showed that when these HER2-positive breast cancer cells become resistant to chemotherapy, i.e., metastatic, they express MUC1. * It has been shown that increasing the expression of anti-MUC1 contributes to metastatic potential (Fessler et al., 2009) (Figure 21D). * MUC1 by Fab *Blockade of the receptor reversed their resistance to Herceptin (Figure 21E), Taxol (Figure 21F), and other chemotherapeutic agents. These and other cancer types expressing NME7 and lower molecular weight NME7 species, such as 33 kDa and 30 kDa, are NME7-dependent. AB Culturing cells in NME7-X1 or lower molecular weight NME7 species can make them more metastatic or can enrich the population of cells that are metastatic.

[0103] Conversely, the metastatic potential of these and other cancer types that express NME7 and lower molecular weight NME7 species, such as 33 kDa or 30 kDa, can be reversed by treating the cells with anti-NME7 antibodies. AB Alternatively, antibodies that bind to NME7-X1 are administered to patients for the treatment or prevention of cancer, including breast, prostate, ovarian, pancreatic and liver cancer. AB However, MUC1 * Since anti-NME7 antibodies have been shown to exert their tumorigenic effects by binding to and activating the growth factor receptor, they are useful for targeting any MUC1 tumors, including, but not limited to, breast, lung, liver, pancreas, stomach, colorectal, prostate, brain, melanoma, and kidney. * It appears to be effective against NME7-positive cancers. AB , or anti-NME7-X1 antibody, AB , NME7 AB -like, or NME7-X1 positive, or MUC1 * It is administered to patients for the treatment or prevention of positive cancer.

[0104] Testing patient cancer cells for effective treatment NME7 ABNME7-X1 and 2i, as well as other reagents that revert stem cells to a more naive state, also induce cancer cells to transform into a more metastatic state. After treatment with any one or a combination of these reagents, cancer cells have a higher engraftment rate, requiring up to 100,000-fold fewer cells to produce tumors in test animals. Thus, the methods described in this disclosure can be used to enable xenotransplantation of a patient's primary tumor cells into test animals.

[0105] Candidate therapeutic agents can then be tested in the recipient animal. Effective therapeutic agents thus identified can be used to treat the donor patient or other patients with similar cancers. In one embodiment, a method for identifying effective therapeutic agents for a particular patient or type of cancer comprises the following steps: 1) cancer cells are obtained from a cell line, a patient, or a patient to whom the test therapeutic agent will be administered; 2) the cancer cells are isolated from a cell line, a patient, or a patient to whom the test therapeutic agent will be administered; AB 2) culturing the resulting cancer cells in NME7-X1, human NME1, bacterial NME1 with high homology to human NME1 or NME7, 2i, or other reagents shown to revert stem cells to a more naive state; 3) culturing the resulting cancer cells in human NME7 AB NME7-X1, human NME1, bacterial NME1 with high homology to human NME1 or NME7, 2i, or other reagents shown to revert stem cells to a more naive state may also be administered to the test animals, or the animals may be implanted with human NME7. AB or NME7-X1; 4) the candidate anti-cancer therapeutic is administered to the animal; 5) the efficacy of the therapeutic is evaluated; and 6) the effective therapeutic is administered to the donor patient or another patient with a similar cancer.

[0106] Anti-NME7 antibody Anti-NME7 antibodies are potent anti-cancer agents. NME7 is a growth factor that promotes the proliferation of cancer cells and their progression to more metastatic or more aggressive states. NME7 and its truncated forms of approximately 33 kDa or 30 kDa were shown to fully support cancer growth, even in serum-free medium lacking any other growth factors or cytokines. In pull-down assays, ELISA, and nanoparticle binding experiments, we demonstrated that NME7 binds to the growth factor receptor MUC1. * NME7 and NME7 AB We showed that NME7 is a binding partner of NME7. Enhancement of this interaction by the exclusion of all other growth factors or cytokines increased the expression of cancer stem cell markers. Blockade of this interaction using a polyclonal antibody that specifically binds to NME7 actively killed cancer cells, even in the presence of serum. Therefore, anti-NME7 or anti-NME7 AB Antibodies are potent anti-cancer drugs that can be administered to patients to treat or prevent cancer. Over 75% of all cancers involve MUC1 * It is positive. MUC1 * is a transmembrane cleavage product of MUC1 in which most of the extracellular domain is deleted, leaving a portion of the extracellular domain that contains most of the PSMGFR sequence and may contain an additional 9–20 amino acids N-terminal to the border of the PSMGFR sequence.

[0107] One aspect of the invention is a method of treating or preventing cancer in a subject, comprising administering to the subject an effective amount of an anti-NME7 antibody. AB In another example, the anti-NME7 antibody can bind to NME7-X1. In yet another example, the anti-NME7 antibody administered to a patient inhibits or prevents binding to its target in promoting cancer. In one example, the target is the extracellular domain of cleaved MUC1. More specifically, the cancer-promoting NME7 target is MUC1 * In one embodiment, an effective therapeutic agent is a therapeutic agent that primarily targets MUC1 * NME7 species consisting of the PSMGFR portion or PSMGFR peptide of MUC1 *The agent for treating or preventing cancer is an agent that disrupts or prevents the interaction between the extracellular domain of NME7, NME7 AB In one example, an effective anti-cancer therapeutic agent is one that binds to or neutralizes the oncogenic activity of an NME7 species. Effective therapeutic agents for the treatment or prevention of cancer include NME7, NME7-like cleavage products, or alternative isoforms, including NME7-X1. AB In one embodiment, the therapeutic agent that binds to or disables NME7-X1-like cleavage products, or alternative isoforms, or NME7-X1. In one embodiment, the therapeutic agent that binds to the NME7 species is an antibody. The antibody can be polyclonal, monoclonal, bispecific, bivalent, monovalent, single chain, scFv, or antibody mimic that can be of animal, human, chimeric, humanized, or human antibody. The antibody can be generated by inoculation or immunization with an NME7 species or a fragment thereof, or can be isolated from, for example, an antibody library or antibody pool, to identify NME7, NME7, NME7-X1. AB The antibodies can be selected for their ability to bind to NME7 species, including NME7-like cleavage products or alternative isoforms, including NME7-X1.

[0108] Generation of anti-NME7 antibodies Anti-NME7 antibodies can be generated outside of a patient, such as in a host animal, or in a patient. Antibodies can be generated by immunization with NME7 or an NME7 fragment, or by immunization with NME7. AB Alternatively, antibodies can be selected from a library or pool of antibodies, which can be natural, synthetic, full-length antibodies, or antibody fragments, based on their ability to bind to a desired NME7 species, such as NME7-X1. In one embodiment, the antibodies are generated from immunization with, or selected for their ability to bind to, a peptide selected from those listed in Figures 6-9. In another embodiment, the antibodies are generated from a peptide whose sequence is not identical to that of human NME1, or the antibodies are selected for their ability to bind to an NME7 species and their inability to bind to human NME1.

[0109] One method used to identify NME7- or NME7-X1-derived peptides that generate antibodies that inhibit cancer growth and metastasis progression, or peptides that are themselves inhibitory, is as follows: 1) the protein sequences of human NME1, human NME7, human NME7-X1, and several bacterial or fungal NME proteins with high sequence homology to human NME1 or human NME7 are aligned; 2) regions of high sequence homology among all NMEs are identified; and 3) peptide sequences that are unique to NME7 or NME7-X1 but flank the regions of high sequence homology are identified. The peptides are then synthesized and used to generate antibodies in humans or host animals. The resulting antibodies are selected for therapeutic use if: 1) they are highly homologous to NME7 or NME7-X1. AB or binds to NME7-X1 but not NME1; 2) has the ability to inhibit cancer growth; 3) has the ability to inhibit the transition of cancer cells to a more metastatic state; or 4) inhibits metastasis in vivo. In some cases, antibodies for therapeutic use may bind to NME7-X1 but not NME1; AB or MUC1 in NME7-X1 * They are selected for their ability to disrupt binding to the extracellular domain, binding to the PSMGFR peptide, or binding to the N-10 peptide.

[0110] Use of anti-NME7 antibodies for the treatment of cancer Antibodies that suppress cancer growth or the transition to a more metastatic state can be selected for use as anticancer therapeutics and administered to patients for the treatment or prevention of cancer. Selected antibodies can be further optimized, for example, by designing or generating human chimeric or fully human antibodies. To demonstrate the effectiveness of this approach, we selected NME7 peptides from regions of NME7 believed to be essential for its anticancer function. We then used these peptides to generate antibodies and tested both the resulting antibodies and the immunizing peptides for their ability to a) suppress cancer growth and b) inhibit the induction of cancer cell to metastatic cancer cell transition. NME7 peptides were selected as immunizing agents for antibody production and as inhibitors themselves (Figure 9 and Example 7). Peptides A1 (SEQ ID NO: 141), A2 (SEQ ID NO: 142), B1 (SEQ ID NO: 143), B2 (SEQ ID NO: 144), and B3 (SEQ ID NO: 145), where A refers to the domain from which the peptide is derived, i.e., the NDPK A domain, and B refers to the peptide derived from the NDPK B domain (Figure 5). Each peptide was used as an immunogen and injected into two rabbits for the production of polyclonal antibodies. Antibodies collected from the immunized rabbit blood were purified on a column derivatized with the immunizing peptide. The purified antibodies were then tested for their ability to bind to human NME7. All of the resulting antibodies bound to human NME7, as desired, but not to human NME1 (Figures 10A-10B, Example 8). These results demonstrate that by selecting a peptide whose sequence is found in NME7 but is not exactly identical to NME1, antibodies can be generated that specifically bind to NME7 but not to NME1. Since NME1 has healthy function, it is often desirable to generate antibodies that do not interfere with NME1. Antibodies also bind to MUC1. * The antibodies were tested for their ability to inhibit NME7 binding to extracellular domain peptides. ELISA experiments shown in Figure 11 demonstrate that the antibodies inhibited MUC1 binding much more than they inhibited NME1 binding. * NME7 to extracellular domain peptides ABIt is to be recalled that each of the NME7 A and B domains can bind to the PSMGFR peptide. AB Complete inhibition of NME7 binding to PSMGFR peptides cannot be achieved with a single antibody or peptides derived from only one domain. These antibodies and their respective immunizing peptides also inhibited cancer cell proliferation (Figures 12-13). These antibodies also inhibited NME7 binding to PSMGFR peptides. AB The polyclonal antibody purified by immunization with the B3 peptide inhibited the formation of non-adherent "floating" cells arising from cancer cells growing within the tumor (Figure 14). As can be seen, the polyclonal antibody purified by immunization with the B3 peptide reduced the number of metastatic floating cells by 95%, indicating that anti-NME7 antibodies that bind to the B3 peptide are the most effective at inhibiting cancer metastasis. Similarly, the antibody inhibited the expression of the metastasis marker CXCR4 (Figure 15A). Again, the B3 antibody was most efficient at inhibiting CXCR4 expression; the bar labeled NME7 FL (NME7 floating cells) shows a 70-fold increase in CXCR4, which was reduced by 20-fold by the B3 antibody 61 (bar labeled NME7+61FL). In addition, the immunization peptide itself inhibited the formation of non-adherent "floating" cells arising from cancer cells growing within the tumor (Figure 15B). AB or 2i suppressed the upregulation of CXCR4 and other metastatic markers.

[0111] This is just one example of a selected peptide that generates antibodies that inhibit the oncogenic functions of NME7 and NME7 species. Sequence alignment between human NME1, human NME7, human NME7-X1, and bacterial NME proteins with high sequence homology to human NME1 or NME7 identified five homologous domains. The fact that peptides A1, A2, B1, B2, and B3 all generated antibodies that inhibited cancer growth or their transition to a metastatic state suggests that the five regions from which these peptides are derived are regions of NME7 that are important for its function in promoting cancer. Other peptides from these regions may also inhibit cancer growth and metastasis, thus generating anti-NME7 antibodies that are potential anti-cancer therapeutics. Antibodies generated from peptides A1, A2, B1, B2, and B3 were shown to inhibit cancer growth and the transition to a more metastatic state. Monoclonal antibodies generated by immunization with the same or similar peptides and subsequent testing of the monoclonal antibodies will identify antibodies that, after humanization or other manipulations known to those skilled in the art, can be administered to patients for the treatment or prevention of cancer.

[0112] In certain experiments, antibodies generated by immunization with peptides A1, A2, B1, B2, and B3, as well as the immunizing peptides themselves, were added to cultured cancer cells to determine whether the addition of the antibodies or immunizing peptides inhibited cancer cell growth. At low concentrations and added separately, the antibodies and immunizing peptides suppressed cancer cell growth (Figure 12 for one example). However, when added at higher concentrations or in combination, the antibodies and immunizing peptides strongly inhibited cancer cell growth (Figure 13). The corresponding human NME7 amino acid numbers of immunizing peptides A1, A2, B1, B2, and B3 are 127-142, 181-191, 263-282, 287-301, and 343-371, respectively, from human full-length NME7 having SEQ ID NO: 82 or 147.

[0113] For clarity, when residue numbers of NME7 are discussed, they refer to the residue numbers of NME7 set forth in SEQ ID NO:82 or 147.

[0114] The antibody used in the cancer growth inhibition experiments and one of the antibodies shown in Figure 12 was generated by immunization with an NME7 peptide corresponding to amino acids 100-376 of NME7 (SEQ ID NO: 82 or 147). To generate anti-NME7 antibodies of higher affinity and specificity, the following steps were followed: immunize animals with a peptide containing human NME7 amino acids 100-376, and then: 1) remove antibodies that bind to human NME1; 2) remove antibodies that bind to NME7. AB 2i or other NMEs that induce the transition of cancer cells to a more metastatic state; 3) select antibodies that inhibit cancer cell proliferation; and 4) select antibodies that inhibit MUC1. * 5) Select antibodies that suppress the growth of MUC1-positive cancer cells * NME7 to the extracellular domain AB or 6) selecting an antibody that inhibits binding of NME7-X1 or substantially inhibits binding to the PSMGFR peptide; and / or 7) selecting an antibody that binds to one or more of the peptides A1, A2, B1, B2, or B3 listed in Figure 9.

[0115] Higher affinity monoclonal antibodies or monoclonal antibodies generated from longer peptides may be more effective antibody therapeutics. AB , or a combination of anti-NME7-X1 antibodies, is administered to the patient to enhance efficacy.

[0116] Anti-NME7 antibody inhibits cancer cell migration to metastatic cancer cells Anti-NME7 antibody inhibits the transition of cancer cells into metastatic cancer cells called cancer stem cells (CSCs) or tumor-initiating cells (TICs). AB Recall that culturing a variety of cancer cells in the presence of NME7 has been demonstrated to transition those cancer cells from normal cancer cells to metastatic CSCs or TICs. AB Alternatively, antibodies that bind to NME7-X1 will inhibit the progression of cancer cells to a more metastatic state.

[0117] Cancer cells transform into a more metastatic state when cultured in the presence of agents that revert stem cells to a more naive state. AB demonstrated that culturing cancer cells in human NME1 dimers, bacterial NME1 dimers, or MEK and GSK3beta inhibitors (termed "2i") causes the cells to become more metastatic. As cells transition to a more metastatic state, they become non- or poorly adherent, floating away from the culture dish. These floating cells, "floaters," were collected separately from the adherent ones and shown to: a) express much higher levels of metastatic genes; and b) generate tumors when xenografted into mice at very low copy numbers. RT-PCR measurements of specific metastatic markers, such as CXCR4 for breast cancer and CHD1 for prostate cancer, and other pluripotent stem cell markers, such as OCT4, SOX2, NANOG, and KLF4, demonstrated the efficacy of NME7. AB It was dramatically overexpressed in cancer cells cultured in PBS, and was most overexpressed in cells that had become non-adherent, referred to here and in the figures as "floaters."

[0118] In one example, NME7 produced by immunization with NME7-derived peptides A1, A2, B1, B2, and B3. AB The specific antibodies, as well as the immunizing peptide itself, were tested against NME7 to determine whether they inhibit the transformation of normal cancer cells into metastatic cancer stem cells. AB The antibody and peptide were added to the culture medium together with either 2i or 2i. The antibody and peptide were separately administered to the cells of interest to induce metastatic transformation, in this case NME7. AB or 2i inhibitors PD0325901 and CHIR99021. AB and 2i were used separately to induce cancer cells to transform into a more aggressive metastatic state. 2i was used because the antibody added to the culture medium simply blocked all NME7 AB was used to block the stool, so that it could not be claimed that the causative agent was effectively absent (Example 10).

[0119] Visual observations were recorded independently by two scientists as the experiment progressed (Figure 14). The most striking observation was that the antibody and peptide dramatically reduced the number of non-adherent cells, which was the first indication that the antibody and peptide inhibited the transformation of metastatic cancer cells. Notably, cells against which the antibody was generated from immunization with the B3 peptide produced very few non-adherent cells. mRNA was extracted from both the non-adherent and adherent cells, as well as control cancer cells. The amount of mRNA, which indicates cell viability and proliferation, was measured. Cells treated with the antibody had much less mRNA, indicating fewer viable dividing cells (Figure 16), which is consistent with anti-NME7. AB This confirms that the antibody inhibits cancer cell proliferation and their transition to a more metastatic state. RT-PCR was used to measure the expression levels of metastatic markers, including CXCR4. Treatment with anti-NME7 antibody significantly reduced the amount of metastatic markers such as CXCR4, indicating that anti-NME7 antibody or peptide inhibited the transition to metastatic cancer (Figures 15A-15C). These results support the conclusion that NME7 inhibits cancer cell proliferation and their transition to a more metastatic state. AB can be administered to patients for the treatment or prevention of metastatic cancer.

[0120] NME7 AB , or NME7-X1 derived peptides, intact NME7 AB and competitively inhibits the binding of NME7-X1 and is an anti-cancer agent. In another embodiment of the present invention, a therapeutic agent for the treatment or prevention of cancer is a peptide derived from the NME7 sequence, which is administered to a patient for the treatment or prevention of cancer. In one embodiment, an NME7-derived peptide is administered to a patient, whereby the peptide, which is shorter than the entire NME7 and which should be unable to confer the oncogenic activity of NME7, binds to the target of NME7 and competitively inhibits the cancer-promoting interaction between the target and intact NME7. NME7 AB can fully confer oncogenic activity, NME7 ABThe sequence of NME7 is preferred as a source of shorter peptides, in which case it must be ensured that the peptide is not itself capable of promoting cancerous growth or other tumor or oncogenic activity. In a preferred embodiment, NME7 AB One or more peptides having a portion of the sequence of NME7, preferably about 12-56 amino acids in length, are administered to the patient. To increase half-life, the peptide may be a peptidomimetic, such as a peptide with a non-natural backbone or D-amino acids instead of L-amino acids. In another example, the anti-cancer therapeutic is a peptide or peptidomimetic, and the peptide may be NME7, NME7, or NME7. AB or NME7-X1, or its target, MUC1, including the PSMGFR peptide, also referred to in some instances herein as "FLR" * It has a sequence highly homologous to at least a portion of the extracellular domain of

[0121] Figures 6-9 provide a list of preferred amino acid sequences predicted to inhibit binding of NME7 to its cognate target. In even more preferred embodiments, peptides selected for administration to patients suffering from or at risk of developing cancer are selected because they bind to an NME7 binding partner and because the peptides themselves do not confer anti-tumor activity. In even more preferred embodiments, the NME7 binding partner is MUC1. * In an even more preferred embodiment, the NME7 binding partner is a PSMGFR peptide.

[0122] The term "conferring tumor or oncogenic activity" means that the peptide itself cannot support or promote cancer growth. Another way to test whether a peptide or peptides derived from NME7 can promote tumor formation is to test whether the peptide can support the pluripotent proliferation of human stem cells. NME proteins and peptides that support pluripotent human stem cell growth also support cancer growth. Alternatively, peptides are deselected if they can revert somatic cells to a less mature state.

[0123] NME7 AB Fragments of NME7 inhibit cancer cell proliferation and the transition of cancer cells to a more metastatic state. Illustratively, NME7 peptides A1, A2, B1, B2, and B3, added separately (FIG. 12) or in combination (FIG. 13), inhibit cancer cell proliferation. Furthermore, NME7 peptides A1, A2, B1, B2, and B3 inhibited the transition of cancer cells to a more metastatic state (FIG. 15).

[0124] Therefore, NME7-specific and NME7 AB Alternatively, antibodies generated by immunization with peptides specific for NME7-X1 may block the oncogenic effects of NME7 species and may be potent anti-cancer agents. Similarly, these results suggest that antibodies specific for NME7 and NME7-X1 may be effective anti-cancer agents. AB or peptides specific to NME7-X1 block the oncogenic effects of NME7 species. In one embodiment of the invention, the peptides are selected from the list shown in Figure 6. In one embodiment of the invention, the peptides are selected from the list shown in Figure 7. In one embodiment of the invention, the peptides are selected from the list shown in Figure 8. In yet another embodiment of the invention, the peptides are selected from the list shown in Figure 9. These antibodies may be generated by immunization, or generated or selected by other means, and then used to detect NME7, NME7-X1-specific peptides, including, but not limited to, NME7-derived peptides A1 (SEQ ID NO: 141), A2 (SEQ ID NO: 142), B1 (SEQ ID NO: 143), B2 (SEQ ID NO: 144), or B3 (SEQ ID NO: 145). AB , NME7-X1, or an NME7-derived peptide. Such antibodies may be polyclonal, monoclonal, bispecific, bivalent, monovalent, single-chain, scFv, human, or humanized, or may be antibody mimetics, such as protein scaffolds, that provide recognition regions that bind to specific targets.

[0125] Anti-NME7 antibodies for use in the treatment or prevention of cancer can be prepared by standard methods known to those skilled in the art, and these methods include the isolation of NME7, NME7 ABor NME7, which lacks the additional 10–25 amino acids from the N-terminus AB These antibodies are used to generate antibodies or antibody-like molecules that recognize shorter versions of the antibody. Such antibodies can be human or humanized. Such antibodies can be polyclonal, monoclonal, bispecific, bivalent, monovalent, single-chain, scFv, human, or humanized, or can be antibody mimics, such as protein scaffolds, that provide recognition regions that bind to specific targets.

[0126] Anti-NME7 antibodies generated by immunization with NME7-derived peptides A1 (SEQ ID NO: 141), A2 (SEQ ID NO: 142), B1 (SEQ ID NO: 143), B2 (SEQ ID NO: 144) or B3 (SEQ ID NO: 145), or antibodies that bind to the A1, A2, B1, B2 or B3 peptides, are NME7 antibodies. AB and NME7-X1, but resist binding to NME1, which may be necessary for the function of some healthy cells. AB or their target receptor, MUC1, of NME7-X1 * Antibodies that bind to the A1, A2, B1, B2, or B3 peptides can be administered to patients diagnosed with or at risk of developing cancer or metastasis. Such antibodies can be human or humanized. Such antibodies can be polyclonal, monoclonal, bispecific, bivalent, monovalent, single-chain, scFv, or antibody mimics, such as protein scaffolds, that provide recognition regions that bind to specific targets.

[0127] Anti-NME7 antibodies generated by immunization with B3 peptide, or antibodies that bind to B3 peptide, are NME7 antibodies. AB and NME7-X1. Such antibodies are particularly specific for recognizing NME7. AB or their target receptor, MUC1, of NME7-X1 *Antibodies that bind to B3 peptides are also highly effective in preventing, inhibiting, and ameliorating cancer or cancer metastasis. Such antibodies can be human or humanized. Such antibodies can be polyclonal, monoclonal, bispecific, bivalent, monovalent, single-chain, scFv, or antibody mimics, such as protein scaffolds, that provide recognition regions that bind to specific targets.

[0128] Note that polyclonal antibody #61, generated in rabbits by immunization with the B3 peptide, inhibited the transformation of cancer cells into cancer stem cells, as evidenced by antibody #61 blocking the upregulation of the metastatic marker CXCR4 (Figure 15).

[0129] The B3 peptide derived from NME7 (SEQ ID NO: 145) has a cysteine ​​at position 14, which complicates the generation of anti-NME7 antibodies. We mutated cysteine ​​14 to serine to create AIFGKTKIQNAVHSTDLPEDGLLEVQYFF (SEQ ID NO: 169), immunized animals, and generated anti-NME7 monoclonal antibodies. The resulting antibodies bind to the native B3 sequence as well as the B3Cys14Ser peptide. We generated seven high-affinity and specific monoclonal antibodies: 8F9A5A1, 8F9A4A3, 5F3A5D4, 5D9E2B11, 5D9E10E4, 5D9G2C4, and 8H5H5G4. However, sequence alignment revealed the presence of only three unique sequence antibodies: 8F9A5A1, 8F9A4A3, and 5F3A5D4, as shown below.

[0130] Heavy Chain Alignment

[0131] [ka]

[0132] Light Chain Alignment

[0133] [ka]

[0134] Monoclonal antibodies 5D9E2B11, 5D9E10E4, 5D9G2C4, and 8H5H5G4 all have the same sequence as 5F3A5D4, also known as 5D4. Herein, when we refer to antibody 5F3A5D4, also known as 5D4, it is understood that this also applies to 5D9E2B11, 5D9E10E4, 5D9G2C4, and 8H5H5G4. As can be seen in Figures 24 and 25, anti-NME7 antibodies 8F9A5A1, 8F9A4A3, and 5F3A5D4 all have the same sequence as NME7. AB This is important because the A domain of NME7 is highly homologous to NME1, which is necessary for normal cell function. AB It may be essential to suppress NME1 and not NME1.

[0135] Figures 26, 27 and 28 show that these anti-NME7 antibodies also AB MUC1 * The figure shows that the binding of MUC1 to the PSMGFR peptide and the N-10 PSMGFR peptide can be disrupted. * NME7 from peptides AB However, there is no complete substitution of NME7. AB consists of A and B domains, each of which is * Recall that these antibodies can bind to the B domain of MUC1. * designed to disrupt binding to NME7 AB The A domain of MUC1 is still attached to the plate surface. * To be useful therapeutic agents, antibodies may bind to a single domain of MUC1. * It is sufficient to disrupt binding to MUC1, thereby preventing ligand-induced dimerization and MUC1 *Activation of growth factor receptors can be blocked. Antibodies or antibody mimetics that bind to the NME7 B3 peptide or B3Cys14Ser peptide (SEQ ID NO: 169) are antibodies that can be administered to patients diagnosed with cancer or metastasis or at risk of developing cancer or metastasis.

[0136] It is well known in the field that it is difficult to generate metastatic cancer cells in animal models. It is estimated that in human tumors, only 1 in 100,000 or 1 in 1,000,000 cancer cells break away from the tumor, implant elsewhere, and initiate metastasis [Al-Hajj et al., 2003]. Some researchers have reported that T47D breast cancer cells injected into immune-compromised mice metastasize after approximately 12 weeks [Harrell et al., 2006]. Other researchers have reported that AsPC-1 pancreatic cancer cells metastasize after approximately 4 weeks [Suzuki et al., 2013].

[0137] Here, we demonstrate that T47D breast cancer cells express recombinant NME7 AB The results show that the cells were grown for 10 days in serum-free medium containing NME7 as the only growth factor. AB When grown in HCl, approximately 25% of the cancer cells began to float, stopped dividing, but remained viable. PCR measurements showed that these "floating" cells significantly upregulated the expression of the breast cancer metastatic factor CXCR4.

[0138] In some of the figures provided herein, these floating cells are referred to as cancer stem cells (CSCs). Immunocompromised female nu / nu mice were implanted with estrogen-releasing pellets for 90 days. 500,000 T47D-wt cells or 10,000 T47D-CSCs (cancer stem cells) were injected subcutaneously (sc) into the tail vein (i.v.) or intraperitoneally (ip) into the nu / nu mice. These cancer cells were engineered to express luciferase. To visualize tumors or cancer cells, animals were injected with luciferin and then visualized 10 minutes later using an IVIS instrument. As can be seen from the IVIS measurements in Figures 33A-33B, by day 6, 500,000 T47D-wt cells injected via the tail vein showed no signs of viable cancer cells or cell engraftment.

[0139] In stark contrast, 10,000 T47D-CSCs injected into the tail vein metastasized. Prior to IVIS measurements on day 6, T47D-CSC mice were treated with 32 nM recombinant NME7. AB The following day, one of the two CSC mice was injected with a cocktail of anti-NME7 monoclonal antibodies 8F9A5A1, 8F9A4A3, and 5F3A5D4 at a concentration equivalent to 15 mg / kg in a volume of 200 uL. AB The near simultaneous injection of IFN-γ and anti-NME7 antibodies likely negated the effect of the antibodies. Figure 34 shows that by day 10, the treated mice were nearly completely metastatic. As can be seen, the mice selected for treatment were more metastatic than comparable T47D-CSC mice.

[0140] The animals were again injected with anti-NME7 antibody on day 10. IVIS measurements on day 12 (Figure 35) show that the antibody-treated mice began to clear metastases. By day 14 (Figure 36), untreated mice died from severe metastases, while treated mice had eliminated metastases. Figure 37 shows the time course of IVIS measurements for mice injected with 500,000 T47D-wt cells and T47D-CSCs that received anti-NME7 treatment until day 17, when antibody treatment was discontinued. As can be seen from the figure, at day 17, small clusters of cancer cells remained, which grew larger by day 19. By day 21, metastases had spread, and antibody treatment was resumed. As shown in the figure, after resumption of anti-NME7 antibody treatment, the animals cleared all metastases and showed no signs of ill health.

[0141] Figure 38 shows the IVIS time course for animals injected subcutaneously or intraperitoneally. Antibody infusions for animals injected subcutaneously or intraperitoneally with CSCs also received anti-NME7 antibodies by sc or ip. In these animals, antibody infusions were stopped on day 17 and not resumed. Figures 39 and 40 show that polyclonal anti-NME7 antibodies generated by immunization with the B3 peptide strain promoted advanced and metastatic cancers, but not normal tissue or low-grade cancers, and that 1 in 100,000 or 1 in 1,000,000 cancer cells would be metastatic. Taken together, these data suggest that anti-NME7 antibodies 8F9A5A1, 8F9A4A3, and 5F3A5D4 or 8F9A5A1, or 8F9A4A3, or 5F3A5D4 administered to patients diagnosed with cancer or at risk of developing cancer will prevent, inhibit, or reverse the formation of cancer metastases.

[0142] In addition to treating metastatic animals with the anti-NME7AB antibody cocktail, we also administered monoclonal anti-NME7AB antibodies individually and showed that they could prevent and even reverse cancer metastasis. In one experimental study, 8-10 week-old female nu / nu mice, each weighing approximately 20 g, were implanted with estrogen-releasing pellets for 90 days. Cancer cells secreted the growth factor NME7. AB The cells were made metastatic by culturing them for 10-15 days in serum-free medium supplemented with IgG. Both adherent and floating cells showed upregulation of metastatic markers and were capable of metastasizing within 4-7 days in animals. In this case, floating cells were harvested on day 11 of in vitro culture and injected into the tail vein of the test animals. To test the preventive model, one group of animals was injected with anti-NME7 antibody into the tail vein. AB Antibody 8F9A4A3 was injected at 15 mg / kg 24 hours before injection of metastatic cancer cells, and the same dose was injected approximately every 48 hours thereafter. Figures 42A-42F show a mouse model of a mouse model injected with 10,000 luciferase-positive T47D metastatic breast cancer stem cells via the tail vein and treated with anti-NME7. AB Photographs of a female nu / nu mouse treated with antibody 4A3 (also known as 8F9A4A3) are shown. To image cancer cells, the luciferase substrate luciferin is injected intraperitoneally 10 minutes before imaging with the IVIS instrument. Figures 42A-42C show IVIS photographs of the animal facing downwards. Figures 42D-42F show IVIS photographs of the animal facing upwards. Figures 42A and 42D show control animals injected with phosphate-buffered saline. Figures 42B and 42E show animals treated with anti-NME7 antibody 24 hours before injection of metastatic cancer cells. AB Figures 42C and 42F show a prophylactic model in which animals were injected with antibody 4A3, followed by approximately every other day for a total of 12 antibody injections over a 22-day period. AB The reversal model shows that the mice were injected with antibody 4A3, followed by 11 injections of the antibody approximately every other day for 20 days. AB The antibody 8F9A4A3 is able to prevent as well as reverse established metastases.

[0143] Anti-NME7AB antibodies 5A1 and 5D4 were also tested in a metastasis prevention model and were shown to significantly inhibit cancer metastasis. Figures 43A-43F show that 10,000 luciferase-positive T47D metastatic breast cancer stem cells were injected into the tail vein, and anti-NME7AB antibodies 5A1 and 5D4 were shown to significantly inhibit cancer metastasis. AB Photographs of female nu / nu mice, each weighing approximately 20 g, treated with antibodies 5A1 (also known as 8F9A5A1) and 5D4 (also known as 5F3A5D4). To image cancer cells, the luciferase substrate luciferin is injected intraperitoneally 10 minutes before imaging with an IVIS instrument. Figures 43A-43C show IVIS photographs of animals facing downward. Figures 43D-43F show IVIS photographs of animals facing upward. Figures 43A and 43D show control animals injected with phosphate-buffered saline. Figures 43B, 43E, 43C, and 43F show animals treated with 15 mg / kg of anti-NME7 antibody 24 hours before injection of metastatic cancer cells. AB This shows a prophylactic model in which mice were injected with antibody 5A1 approximately every other day for a total of 12 injections over a 22-day period. Photographs were taken on days 24 and 27. Specifically, mouse #1 in the group treated with antibody 5A1 was photographed on day 27, while mice #2 and #3 were photographed on day 24 because the animal died on day 26.

[0144] anti-NME7 AB Antibodies 5A1 and 5D4 were also tested in a metastasis reversal model and were shown to significantly inhibit established cancer metastasis. In this experiment, animals were treated with NME7 at a final concentration of 32 nM on day 0. AB In addition, animals were injected twice, on days 3 and 4, with 10,000 T47D metastatic cancer cells mixed with NME7, which in our experiments showed to form metastases that were more difficult to reverse. AB The first antibody injection was on day 7. Because the extent of metastasis in each test animal was somewhat variable, we determined that the apparent elimination of metastatic cancer cells was due to the anti-NME7 AB We needed to confirm that the effects were due to the treatment. Therefore, we treated animals with alternating high and low doses. As can be easily seen in Figure 44, high dose anti-NME7 ABThis resulted in the elimination of metastases, and if complete elimination was not achieved, the patient returned to a lower dose and increased it further. This experiment demonstrated the efficacy of all three anti-NME7 antibodies tested. AB Antibodies 5A1, 4A3, and 5D4 are able to bind to NME7-B3 and inhibit cancer metastasis in a concentration-dependent manner. Figures 44A-44D show that NME7 at a final concentration of 32 nM inhibits cancer metastasis. AB This is a photograph of a female nu / nu mouse that was injected into the tail vein with 10,000 luciferase-positive T47D metastatic breast cancer stem cells mixed with 32 nM NME7. AB After injection into the tail vein, individual anti-NME7 AB Figure 44A shows a control animal injected with phosphate buffered saline. Figure 44B shows an anti-NME7 antibody. AB Figure 44C shows animals treated with monoclonal antibody 8F9A5A1. AB Figure 44D shows animals treated with monoclonal antibody 8F9A4A3. AB An animal treated with monoclonal antibody 5F3A5D4 is shown. The green arrow indicates the low dose (5-7 mg / kg) over the indicated time period, while the red arrow indicates the high dose (15 mg / kg). As can be seen, metastases disappear significantly when the antibody is administered at 15 mg / kg.

[0145] Anti-NME7 of the present invention AB In addition to demonstrating that the antibodies can inhibit metastasis, we tested their effect on metastasis from the primary tumor, which would more closely mimic the physiology of cancer metastasis. We transformed T47D metastatic breast cancer cells, also known as cancer stem cells (CSCs), with NME7. ABT47D CSCs were generated by culturing cancer cells in serum-free minimal medium containing NME7 for 10-15 days. These T47D CSCs were then implanted subcutaneously into the right flank of NSG mice implanted with estrogen-releasing pellets for 90 days. Because the implanted cancer cells are luciferase-positive, after injection of the luciferase substrate, luciferin, the cancer cells emit photons, which can be imaged with an IVIS instrument to measure and locate the implanted cancer cells. Figures 45A and 45B show that on day 0, NME7 was added to a final concentration of 32 nM. AB Photographs of female nu / nu mice are shown that were injected subcutaneously into the right flank with 10,000 luciferase-positive T47D metastatic breast cancer stem cells mixed with 1:1 vol:vol of Matrigel and then mixed 1:1 vol:vol with Matrigel. Tumor engraftment was allowed to progress from day 0 to day 6. Animals were then treated with anti-NME7 AB Antibodies were administered intravenously via tail vein injection. Control animals were injected with PBS. Figure 45A shows an IVIS photograph of a control animal. Figure 45B shows an IVIS photograph of an anti-NME7 AB This figure shows IVIS images of animals injected via the tail vein with a cocktail of antibodies 5A1, 4A3, and 5D4 at a total concentration of 15 mg / kg. Antibodies or PBS were administered four times between days 7 and 18. As can be seen from the figure, anti-NME7 AB Antibody-treated animals show fewer metastases (blue dots throughout the body) than the control group. In the treatment group, two of the five animals have primary tumors that are larger than those in the control group. This is likely due to the anti-NME7 AB This is likely because the antibody prevented the cancer cells from spreading and therefore remained concentrated in the primary tumor. In this experiment, PCR analysis performed before injection of the cancer cells showed that NME7 AB After 11 days in culture with , T47D breast cancer cells showed upregulation of CXCR4 109-fold, OCT4 2-fold, NANOG 3.5-fold, and MUC1 2.7-fold.

[0146] In another experiment, we demonstrated the effect of the anti-NME7 antibody of the present invention on metastasis from the primary tumor to organs to which breast cancer commonly metastasizes. AB We tested the effect of the antibody. Breast cancer usually metastasizes to the liver, lung, bone, and brain, in that order. We transformed T47D metastatic breast cancer cells with NME7. ABT47D CSCs were generated by culturing cancer cells in serum-free minimal medium containing NME7 for 11 days. These T47D CSCs were then implanted subcutaneously into the right flank of NSG mice implanted with estrogen-releasing pellets for 90 days. Figures 46A-46P show that on day 0, NME7 was added to a final concentration of 32 nM. AB Photographs of female nu / nu mice are shown that were injected subcutaneously into the right flank with 10,000 luciferase-positive T47D metastatic breast cancer stem cells mixed with 1:1 vol:vol of Matrigel and then mixed 1:1 vol:vol with Matrigel. Tumor engraftment was allowed to progress from day 0 to day 6. Animals were then treated with anti-NME7 AB The animals were treated intravenously with the antibody via tail vein injection. Control animals were injected with PBS. On day 38, the animals were sacrificed, and the livers were harvested and analyzed by IVIS to detect cancer cells that had metastasized to the liver. Figures 46A and 46B show whole-body IVIS photographs of a control animal injected with PBS only. Figures 46C and 46D show whole-body IVIS photographs of an anti-NME7 control animal injected with PBS only. AB Figures 46E and 46F show whole body IVIS photographs of a control animal injected with antibody 5A1. AB Figures 46G and 46H show whole body IVIS photographs of a control animal injected with antibody 4A3. AB Figures 46A, 46C, 46E, and 46G show whole-body IVIS photographs of a control animal injected with antibody 5D4. Figures 46B, 46D, 46F, and 46H show IVIS photographs taken on day 7 before any treatment. AB IVIS photographs taken 31 days after antibody or mock treatment. As can be seen, animals in the PBS control group showed metastases in whole-body IVIS photographs (blue dots), while animals in the anti-NME7 group showed no metastases (blue dots) in whole-body IVIS photographs. AB Antibody-treated animals do not show metastasis. Figures 46I-46P show photographs and IVIS images of the liver and lungs taken from the animals after sacrifice. Figures 46I, 46K, 46M, and 46O are normal photographs. Figures 46J, 46L, 46N, and 46P are IVIS images revealing cancer cells that have metastasized thereto. As can be seen from the figures, anti-NME7 ABThe antibody significantly inhibited metastasis to the liver, which is the primary site for breast cancer metastasis. Figure 46Q is a bar graph of the measured photons emitted and counted by the IVIS instrument for livers taken from control versus treated animals. As can be seen from the inset of the IVIS measurements, inhibition of liver metastasis follows the order of metastasis when cells are injected into the tail vein, which is consistent with the NME7 antibody. AB -MUC1 * It also matches the order of potency in disrupting interactions.

[0147] We performed immunofluorescence imaging of many cancer cell lines and confirmed that cultured cancer cell lines express NME7 AB As Figures 47A-47F and 48A-48I clearly show, each of the MUC1-positive cancer cell lines we tested was positive for NME7AB, and the binding was membranous, consistent with NME7AB being secreted by the cancer cells, which in turn was associated with MUC1. * Figures 47A-47F show photographs of immunofluorescence experiments in which various cancer cell lines were stained for the presence of NME7AB. Figure 47A shows T47D breast cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47B shows ZR-75-1 breast cancer cells (also known as 1500s) stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47C shows H1975 non-small cell lung cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. Figure 47D shows H1975 non-small cell lung cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. AB Figure 47E shows H292 non-small cell lung cancer cells stained with antibody 5D4. AB Figure 47F shows HPAFII pancreatic cancer cells stained with antibody 5D4. Figure 47G shows DU145 prostate cancer cells stained with various concentrations of anti-NME7AB antibody 5D4. As can be seen, all cancer cell lines we tested show strong membranous staining for NME7AB. The monoclonal antibody used in these experiments was 5D4. In parallel, we stained the same cell lines with NME7AB antibodies 5A1 and 4A3, with identical results.

[0148] Figures 48A-48I show that various human lung cancer cell lines express NME7. AB Figures 48A-48C show photographs of immunofluorescence experiments staining for the presence of various concentrations of anti-NME7. AB Figure 48A shows H1975 non-small cell lung cancer cells, an adenocarcinoma, stained with antibody 5D4. Figure 48B shows DAPI and anti-NME7 AB Figure 48B is an overlay of staining. AB Figure 48C shows single staining. AB Figures 48D-48F show magnified staining overlays. AB Figure 48D shows H292 non-small cell lung cancer cells, a pulmonary mucoepidermoid carcinoma, stained with antibody 5D4. AB Figure 48E shows an overlay of staining with anti-NME7. AB Figure 48F shows staining alone. AB Figures 48G-48I show magnified views of staining overlays. AB Figure 48G shows metastatic bronchoalveolar carcinoma, H358 non-small cell lung cancer cells stained with antibody 5D4. AB Figure 48H is an overlay of staining. AB Figure 48I shows staining alone. AB A close-up of the staining overlay.

[0149] Plus, NME7 AB Culturing these cell lines in serum-free medium containing NME7 even further enhanced their expression of stem cell and metastatic markers. In particular, the cells that became non-adherent, referred to here as suspension, have even higher expression of stem cell and metastatic markers than their adherent counterparts. Figures 49A-49I show that NME7 ABFigure 49 shows PCR graphs of cancer cell lines: breast T47D, lung H1975, lung H358, and pancreatic HPAFII before and after culture in 49A. Figure 49B. The stem cell marker OCT4 was measured. Figure 49C. The metastasis marker ALDH1 was measured. Figure 49D. The stem cell marker SOX2 was measured. Figure 49E. The stem cell marker NANOG was measured. Figure 49F. The metastasis marker CDH1 (also known as E-cadherin) was measured. Figure 49G. The metastasis marker CD133 was measured. Figure 49H. The stem cell marker ZEB2 was measured. Figure 49I. The stem, cancer, and metastasis marker MUC1 was measured. Floating cells (also known as tumorspheres) are able to grow independently of the scaffold and show increased metastasis markers compared to adherent cells. Animals injected with cancer stem cells showed increased expression of NME7. AB As can be seen, no metastasis markers, stem cell markers, or epithelial-mesenchymal transition (EMT) markers were detected in NME7 mice injected with proliferating suspension cells. AB Figure 50 shows that NME7 cells expanded after culture in 100% HO, indicating a transformation to a more metastatic state. AB This figure shows IVIS photographs on day 6 of NSG mice injected with 10,000 H358 parental lung cancer cells or H358 cells via the tail vein after 10–12 days in culture with NME7. As can be seen from the figure, AB NCI-H358 lung cancer cells grown in NME7 medium showed significantly enhanced metastatic potential compared to the parental cells, which are themselves reportedly metastatic. AB The functional enhancement of metastasis from metastatic cancer stem cells at 6 days is consistent with Figure 49, which shows that H358 inhibits NME7 AB The results show that the expression of metastatic markers is significantly enhanced after culture in .

[0150] Figure 51 shows the dimer NM23-H1 (also known as NME1), or NME7 AB1 shows PCR graphs of the MUC1-negative prostate cancer line PC3 before and after 2 or 3 passages in culture in . The graphs show the fold difference in stem cell, cancer cell markers, and metastasis markers. As can be seen, NME1 or NME7 AB Repeated culture in induced upregulation of stem, cancer, and metastasis markers, but also resulted in a 5- to 8-fold upregulation of MUC1 expression.

[0151] Taken together, these data suggest that NME7 lacking the DM10 domain is selected by cancer cells and binds to the extracellular domain of MUC1 lacking the tandem repeat domain, resulting in NME7 binding to MUC1. * They demonstrated that the extracellular domains of NME7 dimerize, leading to increased cancer cell proliferation and an increased metastatic potential of cancer cells. AB and MUC1 * It stands to reason that antibodies that disrupt the interaction between the extracellular domains may inhibit cancer cell proliferation and suppress cancer metastasis. AB and MUC1 * Anti-NME7 inhibits interactions between extracellular domains AB The antibody indeed inhibited cancer cell proliferation and metastasis. AB The antibodies could be administered to patients diagnosed with cancer or metastasis, or at risk of developing cancer or metastasis, to treat or prevent cancer.

[0152] Because NME1 is expressed in the cytoplasm of all cells and is lethal when knocked out, and importantly, the NME1 A domain shares high sequence homology with the NME7 A domain, anti-NME7 antibodies for therapeutic use are being developed. AB The antibody is NME7 AB Importantly, the antibody binds to NME7-X1 but not to NME1. In one aspect of the invention, the antibody that is likely to be optimal for therapeutic use binds to NME7. AB or NME7-X1, and were selected for their ability to bind to peptides that are unique to NME7-X1 and not present in the NME1 sequence. AB The peptides specific to the

[0153] In a preferred embodiment, an antibody suitable for administration to a patient for the treatment or prevention of cancer or cancer metastasis is selected from the group of antibodies that bind to the NME7 B3 peptide. In a further preferred embodiment, an antibody suitable for administration to a patient for the treatment or prevention of cancer or cancer metastasis is selected from the group of antibodies that bind to the NME7 B3 peptide, NME7 AB The antibody is selected from a group of antibodies that bind to NME1 but not NME1. Examples of antibodies that have demonstrated such anti-cancer and anti-metastatic activity in vitro and in vivo and are suitable for therapeutic use in treating or preventing cancer or cancer metastasis include the anti-NME7 antibodies 5A1, 4A3, and 5D4. These are merely examples; antibodies generated as described herein and selected as described herein will have the same anti-cancer and anti-metastatic activity. Such antibodies may be complete antibodies or fragments thereof, including scFvs or antibody mimetics, in which the variable domains of an antibody are incorporated into a protein scaffold that mimics an antibody. Antibodies may be human or non-human, including murine, camelid, llama, human, or humanized, and may be monoclonal, polyclonal, scFv, or fragments thereof.

[0154] Anti-NME7 antibodies for the treatment or prevention of cancer or metastasis can be used in many different therapeutic formats. For example, any antibody described herein, or a fragment thereof, can be administered to a patient as a standalone antibody or antibody fragment, or conjugated to a toxin such as an antibody-drug conjugate (ADC), or incorporated into a bispecific antibody, or incorporated into a BiTE (bispecific T cell engager), or incorporated into a chimeric antigen receptor (CAR) or engineered to be expressed by a cell that also expresses a CAR. The cells can be immune cells, T cells, NK cells, or stem or progenitor cells, which can then be differentiated into T cells or NK cells.

[0155] Any of the antibodies described herein, or fragments thereof, may be used to treat NME7, which may be an indicator of cancer or susceptibility to cancer.AB or as a diagnostic agent to detect body fluids, cells, tissues, or body specimens for the presence of NME7-X1. Diagnostic antibodies can be linked to imaging agents, nucleic acid tags, and can be of any species, including camelids, and can be used systemically or in body fluids such as blood, cells, or tissues, or in vitro, in vivo, or intraoperatively.

[0156] The criteria for selecting a therapeutically or diagnostically useful anti-NME7 antibody depend on the therapeutic or diagnostic format or form in which the antibody is incorporated. When an antibody or antibody fragment is administered to a patient as a stand-alone agent for the treatment or prevention of cancer or cancer metastasis, the antibody should: i) inhibit NME7 AB or NME7-X1 but not NME1; ii) binds to the PSMGFR peptide; iii) binds to the N-10 peptide; and iv) NME7 AB Or NME7-X1 and MUC1 * Interactions between the extracellular domains, or NME7 AB Alternatively, the antibody may be selected for its ability to disrupt the interaction between NME7-X1 and N-10. The antibody may also be selected for its ability to bind to the NME7 B3 peptide. This therapeutic format also encompasses cells engineered to express a CAR and a selected anti-NME7 antibody.

[0157] Other formats require other selection criteria for the anti-NME7 antibody. If the anti-NME7 antibody is incorporated into an ADC, the ADC must be internalized by the target cell to induce its death. AB or NME7-X1 is MUC1 * Recall that the antibody binds to the extracellular domain of MUC1. * If the binding of NME to the extracellular domain is disrupted, the toxin-conjugated antibody will be internalized and will not kill the cell. Similarly, if an anti-NME7 antibody is incorporated into a CAR or BiTE, NME7 ABAlternatively, the NME7-X1 interaction may not be disrupted, or immune cells may no longer be able to target killing agents to cancer cells. AB Alternatively, the interaction between NME7 and X1 may not be disrupted, or the antibody and associated label may be washed away. Thus, for ADC, CAR T, or CAR-NK, BiTE, or diagnostic applications, anti-NME7 antibodies may be used to: i) inhibit NME7; AB or NME7-X1 but not NME1; ii) binds to PSMGFR peptide; iii) binds to N-10 peptide; and iv) MUC1 * Interaction with the extracellular domain, or NME7 AB or NME7 without disrupting the interaction between NME7-X1 and the N-10 peptide. AB or NME7-X1. Antibodies may also be selected for their ability to bind to the NME7 B3 peptide.

[0158] In one aspect of the invention, the cells are AB The cells are engineered to express an antibody or fragment thereof. The cells may be immune cells such as T cells, NK cells, or stem or progenitor cells that can be differentiated into more mature immune cells such as T cells or NK cells. In a preferred embodiment, the cells are engineered to express an anti-NME7 antibody or fragment thereof. AB The cells engineered to express the antibody are also engineered to express a chimeric antigen receptor (CAR). In a preferred embodiment, the CAR recognizes a tumor-associated antigen. In a preferred embodiment, the CAR recognizes MUC1. * In a more preferred embodiment, the CAR targets anti-MUC1 * The CAR is directed to the tumor by the antibody MNC2. In another embodiment of the invention, the cells engineered to express the CAR are also engineered to inducibly express an anti-NME7 antibody. In one example, the anti-NME7 AB The nucleic acid encoding the antibody is inserted within a Foxp3 enhancer or promoter. ABThe antibody is present in a NFAT induction system. In one embodiment, the NFAT induction system is an anti-NME7 AB Incorporates an NFATc1 response element inserted upstream of the antibody sequence, which may be inserted into the IL2 promoter, Foxp3 enhancer or promoter, or other suitable promoter or enhancer.

[0159] In another aspect of the invention, NME7 AB Alternatively, peptides specific to NME7-X1 are incorporated into materials used to immunize or vaccinate humans against cancer or cancer metastasis. In a preferred embodiment, the peptide comprises all or a portion of the NME7 B3 peptide, which may be an NME7 B3 peptide with a Cys-14-Ser mutation.

[0160] Another aspect of the present invention is the use of anti-NME7 antibodies in a host animal. AB The method for generating antibodies includes immunizing an animal with an NME7 B3 peptide. In a preferred embodiment, the NME7 B3 peptide has cysteine ​​14 mutated to serine (SEQ ID NO: 169) to avoid disulfide bond formation, which inhibits NME7-specific antibody generation.

[0161] Another aspect of the invention is to inactivate cells with NME7 AB or a method for generating cells with enhanced metastatic potential, comprising culturing the cells with NME7-X1. These cells can then be used in many aspects of drug discovery.

[0162] Another aspect of the present invention is an anti-NME7 AB or cells engineered to also express NME7-X1. AB Alternatively, NME7-X1 is a human sequence. Their expression can be inducible. In one embodiment, the cell is an egg, which contains human NME7. AB or developed into an animal which may be a transgenic animal capable of expressing NME7-X1.

[0163] NME7 is MUC1 *It binds to the extracellular domain of growth factor receptors and dimerizes them. Tissue studies have shown that MUC1 * Here, we show that NME7 expression increases with increasing tumor grade and metastasis (Fig. 39-Fig. 41). Here, we show that NME7 and MUC1 expression increase with increasing tumor grade and metastasis (Fig. 39-Fig. 41). * Antibodies that inhibit this interaction have been shown to inhibit tumor growth and metastasis.

[0164] Other NME family members include MUC1 * They can bind to and dimerize the extracellular domain of growth factor receptors. For example, we have shown that NME1, NME2, and NME6 exist as dimers and that they interact with MUC1. * It was shown to bind to the extracellular domains and dimerize them. AB and NME7-X1, MUC1 * They have two domains that can bind to the extracellular domain, so they form MUC1 as monomers. * NME7 dimerizes and activates growth factor receptors. We have now shown that anti-NME7 antibodies inhibit cancer and cancer metastasis. Similarly, antibodies or antibody mimics that bind to these other NME proteins can be anti-cancer or anti-metastasis therapeutics that can be administered to patients diagnosed with cancer or metastasis, or patients at risk of developing cancer or metastasis. In one embodiment of the invention, antibodies that can be used therapeutically for the treatment of cancer or metastasis are antibodies that bind to NME1, NME2, NME3, NME4, NME5, NME6, NME7, NME8, NME9, or NME10. In one embodiment of the invention, the therapeutic antibody or antibody mimic inhibits binding of the NME protein to its cognate growth factor receptor. In one embodiment of the invention, the therapeutic antibody or antibody mimic inhibits binding of the NME protein to MUC1 * In another aspect of the invention, the therapeutic antibody or antibody mimetic binds to a peptide derived from NME1, NME2, NME3, NME4, NME5, NME6, NME7, NME8, NME9 or NME10, wherein the peptide is homologous to the NME7 A1, A2, B1, B2 or B3 peptide.

[0165] Below is a sequence alignment showing the homology and identity alignment between NME7 and other NME family members. Underlined or underlined and bolded sequences correspond to NME7 peptides A1 (SEQ ID NO: 141), A2 (SEQ ID NO: 142), B1 (SEQ ID NO: 143), B2 (SEQ ID NO: 144) or B3 (SEQ ID NO: 145).

[0166] Nucleoside diphosphate kinase 7 isoform a [Homo sapiens] (Hu_7)

[0167] [ka]

[0168] >NME2 Theoretical pI / Mw:8.52 / 17298.04

[0169] [ka]

[0170] Global / Global (NW) score: 171; 26.5% identity (56.8% similarity) in 155 aa overlap (1-131:1-152)

[0171] [ka]

[0172] Global / Global (NW) score: 104; 24.4% identity (51.3% similarity) in 156 aa overlap (1-134:1-152)

[0173] [ka]

[0174] >NME3 Theoretical pI / Mw:5.96 / 19088.97

[0175] [ka]

[0176] >NME4 Theoretical pI / Mw:10.30 / 20658.59

[0177] [ka]

[0178] 29.3% identity (68.4% similarity) among 133aa overlap (1-131:56-185)

[0179] [ka]

[0180] 28.8% identity (56.8% similarity) among 132aa overlap (3-134:40-167)

[0181] [ka]

[0182] >NME5 Theoretical pI / Mw:6.08 / 29296.23

[0183] [ka]

[0184] 44.3% identity (74.8% similarity) among 131aa overlap (1-131:13-143)

[0185] [ka]

[0186] 28.0% identity (58.3% similarity) among 132aa overlap (3-134:15-143)

[0187] [ka]

[0188] >NME6 Theoretical pI / Mw:7.81 / 22003.16

[0189] [ka]

[0190] 37.6% identity (68.4% similarity) among 133aa overlap (3-131:22-153)

[0191] [ka]

[0192] 29.3% identity (57.9% similarity) among 133aa overlap (3-134:22-153)

[0193] [ka]

[0194] >NME8 Theoretical pI / Mw:4.90 / 67269.94

[0195] [ka]

[0196] 36.1% identity (69.2% similarity) among 133aa overlap (1-131:316-448)

[0197] [ka]

[0198] Waterman-Eggert score: 269; 85.9 bits; E(1)<1.1e-21 33.6% identity (72.7% similarity) among 128aa overlap (1-127:451-577)

[0199] [ka]

[0200] Waterman-Eggert score: 119; 40.4 bits; E(1)<5.3e-08 33.8% identity (73.8% similarity) within 65aa overlap (3-65:156-220)

[0201] [ka]

[0202] 33.6% identity (65.5% similarity) among 116aa overlap (3-118:453-566)

[0203] [ka]

[0204] Waterman-Eggert score: 128; 41.3 bits; E(1)<2.9e-08 22.3% identity (60.3% similarity) in 116aa overlap (20-134:334-448)

[0205] [ka]

[0206] >--Waterman-Eggert score: 76; 26.4 bits; (E(1)<0.00088 23.4% identity (46.8% similarity) among 111aa overlap (6-105:159-268)

[0207] [ka]

[0208] >NME9

[0209] [ka]

[0210] 46aa overlap (3-46: 100-145) 41.3% identity (67.4% similarity)

[0211] [ka]

[0212] >--Waterman-Eggert score: 30; 13.5 bits; (E(1)<0.85 28.6% identity (71.4% similarity) among 14aa overlap (69-82:100-113)

[0213] [ka]

[0214] >--Waterman-Eggert score: 29; 13.2 bits; (E(1)<0.91 25.8% identity (74.2% similarity) among 31aa overlap (12-42: 121-149)

[0215] [ka]

[0216] 39.6% identity (69.8% similarity) among 53aa overlap (1-53:98-150)

[0217] [ka]

[0218] >NME10 NP_008846.2 Protein XRP2 [Homo sapiens]

[0219] [ka]

[0220] 68aa overlap (11-78:246-308) 23.5% identity (66.2% similarity)

[0221] [ka]

[0222] >Waterman-Eggert score: 35; 15.1 bits; (E(1)<0.73 45aa overlap (66-108:200-244) 28.9% identity (57.8% similarity)

[0223] [ka]

[0224] >--Waterman-Eggert score: 33; 14.4 bits; (E(1)<0.87 75aa overlap (7-80:35-109) 14.7% identity (52.0% similarity)

[0225] [ka]

[0226] >--Waterman-Eggert score: 45; 17.5 bits; (E(1)<0.22 21.6% identity (58.8% similarity) among 51aa overlap (4-50:130-180)

[0227] [ka]

[0228] As an example, antibodies or antibody mimetics that bind to peptides homologous to NME7 ("homologous peptides"), particularly peptides homologous to the A1, A2, B1, B2 or B3 peptides, may be administered to patients diagnosed with cancer or cancer metastasis, or at risk of developing cancer or cancer metastasis.

[0229] Peptides homologous to the A1, A2, B1, B2 or B3 peptides Peptides homologous to the A1, A2, B1, B2, or B3 peptides may include, but are not limited to: NME2A1 (amino acid) RASEEHLKQHYIDLKD (SEQ ID NO: 247)

[0230] NME2A2 (amino acid) PADSKPGT (SEQ ID NO: 248)

[0231] NME2B1 (amino acid) QKGFRLVAMKFLRASEEHLK (SEQ ID NO: 249)

[0232] NME2B2 (amino acid) IDLKDRPFPGLVKY (SEQ ID NO: 250)

[0233] NME2B3 (amino acid) GDFCIQVGRNIIHGSDSVKSAEKEISLWF (SEQ ID NO: 251)

[0234] NME3A1 (amino acid) QASEELLREHYVELRE (SEQ ID NO: 252)

[0235] NME3A1 (amino acid) PGDATPGT (SEQ ID NO: 253)

[0236] NME3B1 (amino acid) RKGFKLVALKLVQASEELLR (SEQ ID NO: 254)

[0237] NME3B2 (amino acid) VELRERPFYSRLVKY (SEQ ID NO: 255)

[0238] NME3B3 (amino acid) GDFCVEVGKNVIHGSDSVESAQREIALWF (SEQ ID NO: 256)

[0239] NME4A1 (amino acid) QAPESVLAEHYQDLRR (SEQ ID NO: 257)

[0240] NME4A2 (amino acid) SAEAAPGT (SEQ ID NO: 258)

[0241] NME4B1 (amino acid) RRGFTLVGMKMLQAPESVLA (SEQ ID NO: 259)

[0242] NME4B2 (amino acid) QDLRRKPFYPALIRY (SEQ ID NO: 260)

[0243] NME4B3 (amino acid) GDFSVHISRNVIHASDSVEGAQREIQLWF (SEQ ID NO: 261)

[0244] NME5A1 (amino acid) RLSPEQCSNFYVEKYG (SEQ ID NO: 262)

[0245] NME5A2 (amino acid) SLVAKETHPDS (SEQ ID NO: 263)

[0246] NME5B1 (amino acid) RSGFTIVQRRKLRLSPEQCS (SEQ ID NO: 264)

[0247] NME5B2 (amino acid) VEKYGKMFFPNLTAY (SEQ ID NO: 265)

[0248] NME5B3 (amino acid) AIYGTDDLRNALHGSNDFAAAEREIRFMF (SEQ ID NO: 266)

[0249] NME6A1 (amino acid) LWRKEDCQRFYREHEG (SEQ ID NO: 267)

[0250] NME6A2 (amino acid) VFRARHVAPDS (SEQ ID NO: 268)

[0251] NME6B1 (amino acid) SNKFLIVRMRELLWRKEDCQ (SEQ ID NO: 269)

[0252] NME6B2 (amino acid) REHEGRFFYQRLVEF (SEQ ID NO: 270)

[0253] NME6B3 (amino acid) GSFGLTDTRNTTHGSDSVVSASREIAAFF (SEQ ID NO: 271)

[0254] NME8A1 (amino acid) VLSEKEAQALCKEYEN (SEQ ID NO: 272)

[0255] NME8A2 (amino acid) VEEAIEYFPES (SEQ ID NO: 273)

[0256] NME8A3 (amino acid) FLTPEQIEKIYPKVTG (SEQ ID NO: 274)

[0257] NME8A4 (amino acid) PEEAKLLSPDS (SEQ ID NO: 275)

[0258] NME8A5 (amino acid) VLTEEQVVNFYSRIAD (SEQ ID NO: 276)

[0259] NME8B1 (amino acid) EAGFDLTQVKKMFLTPEQIE (SEQ ID NO: 277)

[0260] NME8B2 (amino acid) PKVTGKDFYKDLLEM (SEQ ID NO: 278)

[0261] NME8B3 (amino acid) AQFGISKLKNIVH (SEQ ID NO: 279)

[0262] NME8B4 (amino acid) DEDFKILEQRQVVLSEKEAQ (SEQ ID NO: 280)

[0263] NME8B5 (amino acid) KEYENEDYFNKLIEN (SEQ ID NO: 281)

[0264] NME8B6 (amino acid) AQFAMDSLPVNQLYGSDSLETAEREIQHFF (SEQ ID NO: 282)

[0265] NME8B7 (amino acid) KAGFIIEAEHKTVLTEEQVV (SEQ ID NO: 283)

[0266] NME8B8 (amino acid) SRIADQCDFEEFVSF (SEQ ID NO: 284)

[0267] NME9A1 (amino acid) TMTEAEVRLFY (SEQ ID NO: 285)

[0268] NME9B1 (amino acid) EAGFEILTNEERTMTEAEVR (SEQ ID NO: 286)

[0269] NME10A1 (amino acid) SMKAEDAQRVFREK (SEQ ID NO: 287)

[0270] NME10A2 (amino acid) GQRQKSSDES (SEQ ID NO: 288)

[0271] NME10A3 (amino acid) IQDCENCNIYIFDHSA (SEQ ID NO: 289)

[0272] NME10B1 ELAFQFKDAGLSIFNNTWSNIH (SEQ ID NO: 290)

[0273] In some cases, peptides derived from other NME proteins can be made more homologous to the NME7 A1, A2, B1, B2, or B3 peptides by frame shifting or by extending the NME7 peptide such that the extended peptide is more homologous to the NME7 peptide that gives rise to antibodies that inhibit cancer or cancer metastasis. As another example, antibodies or antibody mimetics that bind to extended peptides ("extended peptides") homologous to NME7 can be administered to patients diagnosed with cancer or cancer metastasis, or to patients at risk of developing cancer or cancer metastasis.

[0274] an extended peptide homologous to the A1, A2, B1, B2, or B3 peptide

[0275] Peptides homologous to the extending peptides A1, A2, B1, B2 or B3 peptides may include, but are not limited to:

[0276] NME2A1 (amino acid) RASEEHLKQHYIDLKDRPFFPGL (SEQ ID NO: 291)

[0277] NME2A2 (amino acid) LGETNPADSKPGTIRGDF (SEQ ID NO: 292)

[0278] NME2B1 (amino acid) GLVGEIIKRFEQKGFRLVAMKFLRASEEHLKQHY (SEQ ID NO: 293)

[0279] NME2B2 (amino acid) YIDLKDRPFFPGLVKYMNSGPVVAM (SEQ ID NO: 294)

[0280] NME2B3 (amino acid) PGTIRGDFCIQVGRNIIHGSDSVKSAEKEISLWF (SEQ ID NO: 295)

[0281] NME3A1 (amino acid) LKLVQASEELLREHYVELRERPFYSRL (SEQ ID NO: 296)

[0282] NME3A1 (amino acid) LIGATDPGDATPGTIRGDF (SEQ ID NO: 297)

[0283] NME3B1 (amino acid) LVGEIVRRFERKGFKLVALKLVQASEELLRE (SEQ ID NO: 298)

[0284] NME3B2 (amino acid) EHY-VELRERPFYSRLVKYMGSGPVVAM (SEQ ID NO: 299)

[0285] NME3B3 (amino acid) PGTIRGDFCVEVGKNVIHGSDSVESAQREIALWF (SEQ ID NO: 300)

[0286] NME4A1 (amino acid) GFTLVGMKMLQAPESVLAEHYQDLRRKPF (SEQ ID NO: 301)

[0287] NME4A2 (amino acid) GHTDSAEAAPGTIRGDF (SEQ ID NO: 302)

[0288] NME4B1 (amino acid) LVGDVIQRFERRGFTLVGMKMLQAPESVLAEHY (SEQ ID NO: 303)

[0289] NME4B2 (amino acid) EHYQDLRRKPFYPALIRYMSSGPVVAM (SEQ ID NO: 304)

[0290] NME4B3 (amino acid) PGTIRGDFSVHISRNVIHASDSVEGAQREIQLWF (SEQ ID NO: 305)

[0291] NME5A1 (amino acid) GFTIVQRRKLRLSPEQCSNFYVEKYGKMFF (SEQ ID NO: 306)

[0292] NME5A2 (amino acid) LLGPNNSLVAKETHPDSLRAIYGTD (SEQ ID NO: 307)

[0293] NME5B1 (amino acid) IQDIILRSGFTIVQRRKLRLSPEQCSNFY (SEQ ID NO: 308)

[0294] NME5B2 (amino acid) FYVEKYGKMFFPNLTAYMSSGPLVAM (SEQ ID NO: 309)

[0295] NME5B3 (amino acid) PDSLRAIYGTDDLRNALHGSNDFAAAEREIRFMF (SEQ ID NO: 310)

[0296] NME6A1 (amino acid) FLIVRMRELLWRKEDCQRFYREHEGRFFYQRL (SEQ ID NO: 311)

[0297] NME6A2 (amino acid) LMGPTRVFRARHVAPDSIRGSFG (SEQ ID NO: 312)

[0298] NME6B1 (amino acid) ILSNKFLIVRMRELLWRKEDCQRFY (SEQ ID NO: 313)

[0299] NME6B2 (amino acid) FYREHEGRFFYQRLVEFMASGPIRA (SEQ ID NO: 314)

[0300] NME6B3 (amino acid) ARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFF (SEQ ID NO: 315)

[0301] NME8A1 (amino acid) FKILEQRQVVLSEKEAQALCKEYENEDYFNKLI (SEQ ID NO: 316)

[0302] NME8A2 (amino acid) WKQLLGPRTVEEAIEYFPESLCAQFAMD (SEQ ID NO: 317)

[0303] NME8A3 (amino acid) AGFDLTQVKKMFLTPEQIEKIYPKVTGKDFYKDL (SEQ ID NO: 318)

[0304] NME8A4 (amino acid) EWRRLMGPTDPEEAKLLSPDSIRAQFG (SEQ ID NO: 319)

[0305] NME8A5 (amino acid) KAGFIIEAEHKTVLTEEQVVNFYSRIADQCDFEE (SEQ ID NO: 320)

[0306] NME8B1 (amino acid) ILKIVKEAGFDLTQVKKMFLTPEQIEKIY (SEQ ID NO: 321)

[0307] NME8B2 (amino acid) YPKVTGKDFYKDLLEMLSVGP (SEQ ID NO: 322)

[0308] NME8B3 (amino acid) DPEEAKLLSPDSIRAQFGISKLKNIVH (SEQ ID NO: 323)

[0309] NME8B4 (amino acid) LRIIKDEDFKILEQRQVVLSEKEAQ (SEQ ID NO: 324)

[0310] NME8B5 (amino acid) KEYENEDYFNKLIENMTSGPSLA (SEQ ID NO: 325)

[0311] NME8B6 (amino acid) PESLCAQFAMDSLPVNQLYGSDSLETAEREIQHFF (SEQ ID NO: 326)

[0312] NME8B7 (amino acid) IKRKITKAGFIIEAEHKTVLTEEQVVNFY (SEQ ID NO: 327)

[0313] NME8B8 (amino acid) FYSRIADQCDFEEFVSFMTSG (SEQ ID NO: 328)

[0314] NME9A1 (amino acid) AGFEILTNEERTMTEAEVRLFY (SEQ ID NO: 329)

[0315] NME9B1 (amino acid) IIMKIQEAGFEILTNEERTMTEAEVRLFY (SEQ ID NO: 330)

[0316] NME10A1 (amino acid) GFFLVQTKEVSMKAEDAQRVFREKAP (SEQ ID NO: 331)

[0317] NME10A2 (amino acid) EANRSIVPISRGQRQKSSDESCLVVLFAGD (SEQ ID NO: 332)

[0318] NME10A3 (amino acid) IQDCENCNIYIFDHSA (SEQ ID NO: 333)

[0319] NME10B1 ELAFQFKDAGLSIFNNTWSNIHDFTPVDCT (SEQ ID NO: 334)

[0320] Some NME proteins perform functions that are necessary for normal cell growth or development. For example, NME1 is thought to be required for normal cell function. Other NME proteins have catalytic domains whose functions are required in normal cells or tissues. In these cases, therapeutic antibodies can be selected based on their ability to bind to targeted cancer-associated NMEs but not non-targeted NMEs. For example, the anti-NME7 antibodies presented herein, 8F9A5A1, 8F9A4A3, and 5F3A5D4, bind to NME7. AB are selected for their ability to bind to NME1 but not NME1; they are further selected based on their ability to inhibit cancer and cancer metastasis.

[0321] In another embodiment of the invention, an anti-NME7 antibody, antibody fragment, e.g., scFv, or antibody mimetic fragment, is incorporated into a chimeric antigen receptor (CAR) that is engineered for expression in immune cells. The immune cells are engineered to express an anti-NME7 CAR, an anti-MUC1 CAR, or an anti-NME7 CAR. *The immune cells can be engineered to express either the MUC1 or CAR, or both. One of the CARs can be expressed from an inducible promoter. Alternatively, the immune cells can be engineered to express either the MUC1 or CAR. * The CAR can be engineered to express a CAR and an inducible anti-NME7 antibody or antibody fragment. In some cases, the inducible promoter can include an NFAT response element. In one embodiment, these engineered species are expressed in T cells, NK cells, or dendritic cells. Immune cells can be obtained from patients or donors. In some cases, immune molecules such as MHC, checkpoint inhibitors, or receptors for checkpoint inhibitors are mutated or deleted, for example, using CrisPR or CrisPR-like technology. In another embodiment, ITAM molecules, Fos, or Jun are genetically deleted in patient- or donor-derived immune cells by mutagenesis or Talen, Sleeping Beauty, CrisPR, or CrisPR-like technology.

[0322] In one embodiment of the invention, an anti-NME7 antibody or antibody mimetic for use in a CAR T format is specific for NME7 but MUC1 * The antibody or antibody mimic is selected from a group of antibodies or antibody mimics that do not disrupt the binding of NME7 to the extracellular domain of NME1 or NME2. Thus, an anti-NME7 antibody or antibody mimic that targets CAR T to tumors will not simply deprive the receptor of the ligand, thereby preventing the T cell from injecting granzyme B into the target cancer cells. Such antibodies or antibody mimics are generated by immunizing animals with NME7 peptides, such as NME7 peptides A1, A2, B1, B2, or B3, or selected for their ability to bind to NME7 peptides A1, A2, B1, B2, or B3. The antibody or antibody mimic is selected for its ability to specifically bind to NME7, but not to NME1 or NME2, and for its ability to specifically bind to NME7 and MUC1. * The bond between the extracellular domains can be screened for as they cannot be disrupted. For example, in an ELISA device, the PSMGFR peptide is immobilized on a surface. AB Surface-immobilized MUC1 *It binds to the extracellular domain and binds to NME7 AB Detection of the label is measured in the presence or absence of the test antibody or antibody mimetic. AB and surface-immobilized MUC1 * Antibodies that do not reduce binding between the extracellular domain peptide are selected as antibodies to be incorporated into CARs, engineered to be expressed in immune cells, and then administered to patients for the treatment or prevention of cancer or cancer metastasis.

[0323] In one embodiment of the invention, an anti-NME7 antibody or fragment thereof is administered to a patient diagnosed with, or at risk of developing, cancer or cancer metastasis. In one embodiment, the anti-NME7 antibody or antibody fragment binds to an NME peptide, particularly those discussed above in the sections "Peptides homologous to the A1, A2, B1, B2 or B3 peptides" and "Extended peptides homologous to the A1, A2, B1, B2 or B3 peptides."

[0324] In another embodiment, the antibody, antibody fragment, or antibody mimetic binds to an NME7-derived peptide selected from A1, A2, B1, B2, or B3 (SEQ ID NOs: 141-145). In yet another embodiment, the antibody, antibody fragment, or antibody mimetic binds to an NME7 peptide that includes most or all of the B3 peptide. In one embodiment of the invention, the anti-NME7 antibody, antibody fragment, or antibody mimetic comprises sequences from the variable domains of the anti-NME7 antibodies 8F9A4A3 (SEQ ID NOs: 1001-1015), 8F9A5A1 (SEQ ID NOs: 1016-1030), or 8H5H5G4 (SEQ ID NOs: 1031-1045) shown below.

[0325] Anti-NME7 B3 peptide monoclonal antibody 8F9A4A3 heavy chain variable region sequence mouse (DNA) Gtccagctgcaacagtctggacctgaactggtgaagcctggggcttcagtgaagatatcctgcaagacttctggaaacacattcactgaatacaccatgcactgggtgaagcagagccatggaaagagccttgagtggattggaggttttaatcctaacaatggtgttactaactacaaccagaag ttcaagggcaaggccacattgactgtagacaagtcctccagcacagcctacatggagctccgcagcctgacatctgaggattctgcagtctattactgtgcaagacggtactaccatagtctctacgtgttttactttgactactggggccaaggcaccactctcacagtctcctca (SEQ ID NO: 335) (amino acid) VQLQQSGPELVKPGASVKISCKTSGNTFTEYTMHWVKQSHGKSLEWIGGFNPNNGVTNYNQKFKGKATLTVDKSSSTAYMELRSLTSEDSAVYYCARRYYHSLYVFYFDYWGQGTTLTVSS (SEQ ID NO: 1001)

[0326] IGHV1-24 * 01 V-region sequence human (closest match hu antibody sequence) (DNA) Caggtccagctggtacagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaaggtttccggatacaccctcactgaattatccatgcactgggtgcgacaggctcctggaaaagggcttgagtggatgggaggtt ttgatcctgaagatggtgaaacaatctacgcacagaagttccagggcagagtcaccatgaccgaggacacatctacagacacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtattactgtgcaaca (SEQ ID NO: 336) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGYTLTELSMHWVRQAPGKGLEWMGGFDPEDGETIYAQKFQGRVTMTEDTSTDTAYMELSSLRSEDTAVYYCAT (SEQ ID NO: 1002)

[0327] Human (closest match hu antibody sequence) IGHJ4*01 J region sequence (DNA) tactttgactactggggccaaggaaccctggtcaccgtctcctca (SEQ ID NO: 337) (amino acid) YFDYWGQGTLVTVSS (SEQ ID NO: 1003)

[0328] Humanized heavy chain variable sequence (SEQ ID NO:1001 + SEQ ID NO:1002 + SEQ ID NO:1003) Humanized 8F9A4A3 heavy chain variable region sequence (DNA) caggtccagctggtacagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctctcctgcaaggtttcggaaacacattcactgaatacaccatgcactgggtgcgacaggctcctggaaaagggcttgagtggatgggaggttttaatcctaacaatggtgttactaactacaaccaga agttcaagggcagagtcaccatgaccgagacacatctacagacacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtattactgtgcaagacggtactaccatagtctctacgtgttttactttgactactggggccaaggaaccctggtcaccgtctcctca (SEQ ID NO: 338) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWMGGFNPNNGVTNYNQKFKGRVTMTEDTSTDTAYMELSSLRSEDTAVYYCARRYYHSLYVFYFDYWGQGTLVTVSS (SEQ ID NO: 1004)

[0329] Humanized heavy chain variable sequence (codon-optimized version of 1004) Humanized 8F9A4A3 heavy chain variable region sequence (codon optimized) (DNA) caggttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggtgtccggaaataccttcaccgagtacaccatgcactgggtccgacaggcccctggcaaaggacttgaatggatgggcggcttcaaccccaacaacggcgtgaccaactacaaccaga aattcaagggccgcgtgaccatgaccgaggacacaagcacagacaccgcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagaaggtactaccacagcctgtacgtgttctacttcgactactggggccagggcaccctggtcacagtttcttct (SEQ ID NO: 339) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWMGGFNPNNGVTNYNQKFKGRVTMTEDTSTDTAYMELSSLRSEDTAVYYCARRYYHSLYVFYFDYWGQGTLVTVSS (SEQ ID NO: 1005)

[0330] Humanized heavy chain variable sequence ("modified" SEQ ID NO: 1005 sequence, modified means that certain amino acids thought to be important for binding or structure were added back to the murine sequence) Modified humanized 8F9A4A3 heavy chain variable region sequence (DNA) caggtccagctggtacagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctctcctgcaaggtttccggaaacacattcactgaatacaccatgcactgggtgcgacaggctcctggaaaagggcttgagtggattggaggttttaatcctaacaatggtgttactaactacaaccaga agttcaagggcaaagtcaccctgaccgtggacacatctagcagcacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtattactgtgcaagacggtactaccatagtctctacgtgttttactttgactactggggccaaggaaccctggtcaccgtctcctca (SEQ ID NO: 340) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWIGGFNPNNGVTNYNQKFKGKVTLTVDTSSSTAYMELSSLRSEDTAVYYCARRYYHSLYVFYFDYWGQGTLVTVSS (SEQ ID NO: 1006)

[0331] Humanized heavy chain variable sequence (codon optimized SEQ ID NO: 1006) Modified humanized 8F9A4A3 heavy chain variable region sequence (codon optimized) (DNA) caggttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggtgtccggaaataccttcaccgagtacaccatgcactgggtccgacaggcccctggcaaaggactggaatggatcggcggcttcaaccccaacaacggcgtgaccaactacaaccaga aattcaagggcaaagtgaccctgaccgtggacaccagcagcagcacagcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagaaggtactaccacagcctgtacgtgttctacttcgactactggggccagggcaccctggtcacagtttcttct (SEQ ID NO: 341) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWIGGFNPNNGVTNYNQKFKGKVTLTVDTSSSTAYMELSSLRSEDTAVYYCARRYYHSLYVFYFDYWGQGTLVTVSS (SEQ ID NO: 1007)

[0332] 8F9A4A3 light chain variable region sequence mouse (DNA) gaaacaactgtgacccagtctccagcatccctgtccatggctataggagaaaaagtcaccatcagatgcataaccagcactgatattgatgatgatatgaactggtaccagcagaagccaggggaacctcctaagctccttatttcagaaggcaatactcttcgtc ctggagtcccatcccgattctccagcagtggctatggtacagattttgtttttacaattgaaaacatgctctcagaagatgttgcagattactactgtttgcaaagtgataacttgcctctcacgttcggctcggggacaaagttggaaataaaacgg (SEQ ID NO: 342) (amino acid) ETTVTQSPASLSMAIGEKVTIRCITSTDIDDDMNWYQQKPGEPPKLLISEGNTLRPGVPSRFSSSGYGTDFVFTIENMLSEDVADYYCLQSDNLPLTFGSGTKLEIKR (SEQ ID NO: 1008)

[0333] Human (closest match hu antibody sequence) IGKV5-2 * 01 V region array (DNA) gaaacgacactcacgcagtctccagcattcatgtcagcgactccaggagacaaagtcaacatctcctgcaaagccagccaagacattgatgatgatatgaactggtaccaacagaaaccaggagaagctgctattt tcattattcaagaagctactactctcgttcctggaatcccacctcgattcagtggcagcgggtatggaacagattttaccctcacaattaataacatagaatctgaggatgctgcatattacttctgt (SEQ ID NO: 343) (amino acid) ETTLTQSPAFMSATPGDKVNISCKASQDIDDDMNWYQQKPGEAAIFIIQEATTLVPGIPPRFSGSGYGTDFTLTINNIESEDAAYYFC (SEQ ID NO: 1009)

[0334] Human (closest match hu antibody sequence) IGKJ4 * 02 J region array (DNA) ctcacgttcggcggagggaccaaggtggagatcaaa (SEQ ID NO: 344) (amino acid) LTFGGGTKVEIK (SEQ ID NO: 1010)

[0335] Humanized light chain variable sequence (SEQ ID NO:1008 + SEQ ID NO:1009 + SEQ ID NO:1) Humanized 8F9A4A3 light chain variable region sequence (DNA) gaaacgacactcacgcagtctccagcattcatgtcagcgactccaggagacaaagtcaacatctcctgcataaccagcactgatattgatgatgatatgaactggtaccaacagaaaccaggagaagctgctattttcattattcaagaaggcaatactcttcgtc ctggaatcccacctcgattcagtggcagcgggtatggaacagattttaccctcacaattaataacatagaatctgaggatgctgcatattacttctgtttgcaaagtgataacttgcctctcacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 345) (amino acid) ETTLTQSPAFMSATPGDKVNISCITSTDIDDDMNWYQQKPGEAAIFIIQEGNTLRPGIPPRFSGSGYGTDFTLTINNIESEDAAYYFCLQSDNLPLTFGGGTKVEIKR (SEQ ID NO: 1011)

[0336] Humanized light chain variable sequence (codon optimized version of SEQ ID NO: 1011) Modified humanized 8F9A4A3 light chain variable region sequence (codon optimized) (DNA) Gagacaaccctgacacagagccctgccttcatgtctgccacacctggcgacaaagtgaacatcagctgcatcaccagcaccgacatcgacgacgacatgaactggtatcagcagaagcctggcgaggccgccatcttcatcatccaagagggcaacacactgcggc ctggcatccctcctagattttctggcagcggctacggcaccgacttcaccctgaccatcaacaacatcgagagcgaggacgccgcctactacttctgcctgcaaagcgacaacctgcctctgacctttggcggaggcaccaaggtggaaatcaagcgg (SEQ ID NO: 346) (amino acid) ETTLTQSPAFMSATPGDKVNISCITSTDIDDDMNWYQQKPGEAAIFIIQEGNTLRPGIPPRFSGSGYGTDFTLTINNIESEDAAYYFCLQSDNLPLTFGGGTKVEIKR (SEQ ID NO: 1012)

[0337] Humanized light chain variable sequence ("modified" SEQ ID NO: 1012 sequence, modified means that certain amino acids thought to be important for binding or structure were added back to the murine sequence) Modified humanized 8F9A4A3 light chain variable region sequence (DNA) gaaacgacagtgacgcagtctccagcattcatgtcagcgactccaggagacaaagtcaccatctcctgcataaccagcactgatattgatgatgatatgaactggtaccaacagaaaccaggagaagctgctattctgctgattagcgaaggcaatactcttcgtc ctggaatcccacctcgattcagtagcagcgggtatggaacagattttaccctcacaattaataacatagaatctgaggatgctgcatattacttctgtttgcaaagtgataacttgcctctcacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 347) (amino acid) ETTVTQSPAFMSATPGDKVTISCITSTDIDDDMNWYQQKPGEAAILLISEGNTLRPGIPPRFSSSGYGTDFTLTINNIESEDAAYYFCLQSDNLPLTFGGGTKVEIKR (SEQ ID NO: 1013)

[0338] Humanized light chain variable sequence (codon optimized SEQ ID NO: 1013) Modified humanized 8F9A4A3 light chain variable region sequence (codon optimized) (DNA) gagacaaccgtgacacagagccctgccttcatgtctgccacacctggcgacaaagtgaccatcagctgcatcaccagcaccgacatcgacgacgacatgaactggtatcagcagaagcctggcgaggccgccatcctgcttatctctgagggaaacacactgcggc ctggcatccctcctagattttccagcagcggctacggcaccgacttcaccctgaccatcaacaacatcgagagcgaggacgccgcctactacttctgcctgcaaagcgacaacctgcctctgacctttggcggaggcaccaaggtggaaatcaagcgg (SEQ ID NO: 348) (amino acid) ETTVTQSPAFMSATPGDKVTISCITSTDIDDDMNWYQQKPGEAAILLISEGNTLRPGIPPRFSSSGYGTDFTLTINNIESEDAAYYFCLQSDNLPLTFGGGTKVEIKR (SEQ ID NO: 1014)

[0339] Humanized heavy and light chains linked via a flexible linker. Modified humanized 8F9A4A3 sequence (codon optimized) (DNA) Caggttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggtgtccggaaataccttcaccgagtacaccatgcactgggtccgacaggcccctggcaaaggactggaatggatcggcggcttcaaccccaacaacggcgtgaccaactacaaccagaaattcaagggcaaagtgaccctgaccgtggacaccagcagcagcacagcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagaaggtactaccacagcctgtacgtgttctacttcgactactggggccagggcaccctggtcacagtttcttctggcggtggcggaagcggaggcggtggctccggtggcggaggcagcgaaacgacagtgacgcagtctccagcattcatgtcagcgactccaggagacaaagtcaccatctcctgcataaccagcactgatattgatgatgatatgaactggtaccaacagaaaccaggagaagctgctattctgctgattagcgaaggcaatactcttcgtcctggaatcccacctcgattcagtagcagcgggtatggaacagattttaccctcacaattaataacatagaatctgaggatgctgcatattacttctgtttgcaaagtgataacttgcctctcacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 349) (Amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWIGGFNPNNGVTNYNQKFKGKVTLTVDTSSSTAYMELSSLRSEDTAVYYCARRYYHSLYVFYFDYWGQGTLVTVSSGGGGSGGGGSGGGGSETTVTQSPAFMSATPGDKVTISCITSTDIDDDMNWYQQKPGEAAILLISEGNTLRPGIPPRFSSSGYGTDFTLTINNIESEDAAYYFCLQSDNLPLTFGGGTKVEIKR (SEQ ID NO: 1015)

[0340] 8F9A5A1 heavy chain variable region sequence (DNA) atccagttggtgcagtctggacctgagctgaagaagcctggagagacagtcaagatctcctgcaaggcttctgggtataccttcacaaactatggaatgaactgggtgaagcaggctccaggaaagggtttaaagtggatgggctggataaacacctacactggagagccaacatatgttgat gacttcaagggacggtttgccttctctttggaaacctctgccaccactgcctatttgcagatcaacaacctcaaaaatgaggacacgtctacatatttctgtgcaagattgagggggatacgaccgggtcccttggcttactggggccaagggactctggtcactgtctctgca (SEQ ID NO: 350) (amino acid) IQLVQSGPELKKPGETVKISCKASGYTFTNYGMNWVKQAPGKGLKWMGWINTYTGEPTYVDDFKGRFAFSLETSATTAYLQINNLKNEDTSTYFCARLRGIRPGPLAYWGQGTLVTVSA (SEQ ID NO: 1016)

[0341] IGHV7-81 * 01 V region array (DNA) caggtgcagctggtgcagtctggccatgaggtgaagcagcctggggcctcagtgaaggtctcctgcaaggcttctggttacagtttcaccacctatggtatgaattgggtgccacaggcccctggacaagggcttgagtggatgggatggt tcaacacctacactgggaacccaacatatgcccagggcttcacaggacggtttgtcttctccatggacacctctgccagcacagcatacctgcagatcagcagcctaaaggctgaggacatggccatgtattactgtgcgaga (SEQ ID NO: 351) (amino acid) QVQLVQSGHEVKQPGASVKVSCKASGYSFTTYGMNWVPQAPGQGLEWMGWFNTYTGNPTYAQGFTGRFVFSMDTSASTAYLQISSLKAEDMAMYYCAR (SEQ ID NO: 1017)

[0342] IGHJ4 * 03 J region array (DNA) tactttgactactggggccaagggaccctggtcaccgtctcctca (SEQ ID NO: 352) (amino acid) YFDYWGQGTLVTVSS (SEQ ID NO: 1018) SEQ ID NO: 1019

[0343] Humanized 8F9A5A1 heavy chain variable region sequence (DNA) Caggtgcagctggtgcagtctggccatgaggtgaagcagcctggggcctcagtgaaggtctcctgcaaggcttctgggtataccttcacaaactatggaatgaactgggtgccacaggcccctggacaagggcttgagtggatgggatggataaacacctacactggagagccaacatatgttg atgacttcaagggacggtttgtcttctccatggacacctctgccagcacagcatacctgcagatcagcagcctaaaggctgaggacatggccatgtattactgtgcaagattgagggggatacgaccgggtcccttggcttactggggccaagggaccctggtcaccgtctcctca (SEQ ID NO: 353) (amino acid) QVQLVQSGHEVKQPGASVKVSCKASGYTFTNYGMNWVPQAPGQGLEWMGWINTYTGEPTYVDDFKGRFVFSMDTSASTAYLQISSLKAEDMAMYYCARLRGIRPGPLAYWGQGTLVTVSS (SEQ ID NO: 1019)

[0344] Humanized 8F9A5A1 heavy chain variable region sequence (codon optimized) (DNA) caggttcagctggtgcagtctggccacgaagtgaaacagcctggcgcctctgtgaaggtgtcctgtaaagccagcggctacacctttaccaactacggcatgaactgggtgccccaggctcctggacaaggcttggaatggatgggctggatcaacacctacacccggcgagcctacctacgtgg acgacttcaagggcagattcgtgttcagcatggacaccagcgccagcacagcctacctgcagatcagctctctgaaggccgaggatatggccatgtactactgcgccagactgagaggcatcagacctggacctctggcctattggggacagggcacactggtcacagtgtcctct (SEQ ID NO: 354) (amino acid) QVQLVQSGHEVKQPGASVKVSCKASGYTFTNYGMNWVPQAPGQGLEWMGWINTYTGEPTYVDDFKGRFVFSMDTSASTAYLQISSLKAEDMAMYYCARLRGIRPGPLAYWGQGTLVTVSS (SEQ ID NO: 1020)

[0345] Modified humanized 8F9A5A1 heavy chain variable region sequence (DNA) cagatccagctggtgcagtctggccccgaggtgaagcagcctggggcctcagtgaaggtctcctgcaaggcttctgggtataccttcacaaactatggaatgaactgggtgaagcaggcccctggacaagggcttgagtggatgggatggataaacacctacactggagagccaacatatgttg atgacttcaagggacggtttgccttctccatggacacctctgccagcacagcatacctgcagatcagcagcctaaaggctgaggacaccgccacctattactgtgcaagattgagggggatacgaccgggtcccttggcttactggggccaagggaccctggtcaccgtctcctca (SEQ ID NO: 355) (amino acid) QIQLVQSGPEVKQPGASVKVSCKASGYTFTNYGMNWVKQAPGQGLEWMGWINTYTGEPTYVDDFKGRFAFSMDTSASTAYLQISSLKAEDTATYYCARLRGIRPGPLAYWGQGTLVTVSS (SEQ ID NO: 1021)

[0346] Modified humanized 8F9A5A1 heavy chain variable region sequence (codon optimized) (DNA) cagattcagctggtgcagtctggccccgaagtgaaacaacctggcgcctctgtgaaggtgtcctgcaaggccagcggctacacctttaccaactacggcatgaactgggtcaagcaggcccctggacaaggcctggaatggatgggctggatcaacacctacacggcgagcctacctacgtgg acgacttcaagggcagattcgccttcagcatggacaccagcgccagcacagcctacctgcagatcagctctctgaaggccgaggacaccgccacctactactgtgccagactgagaggcatcagacccggacctctggcctattggggacagggaacactggtcaccgtgtcctct (SEQ ID NO: 356) (amino acid) QIQLVQSGPEVKQPGASVKVSCKASGYTFTNYGMNWVKQAPGQGLEWMGWINTYTGEPTYVDDFKGRFAFSMDTSASTAYLQISSLKAEDTATYYCARLRGIRPGPLAYWGQGTLVTVSS (SEQ ID NO: 1022)

[0347] 8F9A5A1 light chain variable region sequence (DNA) gaaattttgctcacccagtctccagcaatcatagctgcatctcctggggagaaggtcaccatcacctgcagtgccagctcaagtgtaagttacatgaactggtaccagcagaaaccaggatcctcccccaaaatatggatttatggtatatccaacctggcttct ggagttcctgctcgcttcagtggcagtgggtctgggacatctttctctttcacaatcaacagcatggaggctgaagatgttgccacttattactgtcagcaaaggagtagttacccacccacgttcggaggggggaccaagctggaaataaaacgg (SEQ ID NO: 357) (amino acid) EILLTQSPAIIAASPGEKVTITCSASSSVSYMNWYQQKPGSSPKIWIYGISNLASGVPARFSGSGSGTSFSFTINSMEAEDVATYYCQQRSSYPPTFGGGTKLEIKR (SEQ ID NO: 1023)

[0348] IGKV3D-15 * 02 V region array (DNA) gaaatagtgatgatgcagtctccagccaccctgtctgtgtctccaggggaaagagccaccctctcctgcagggccagtcagagtgttagcagcaacttagcctggtaccagcagaaacctggccaggctcccaggctcctcatc tatggtgcatccaccagggccactggcatcccagccaggttcagtggcagtgggtctgggacagagttcactctcaccatcagcagcctgcagtctgaagattttgcagtttattactgtcagcagtataataac (SEQ ID NO: 358) (amino acid) EIVMMQSPATLSVSPGERATLSCRASQSVSSNLAWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTEFTLTISSLQSEDFAVYYCQQYNN (SEQ ID NO: 1024)

[0349] IGKJ4 * 02 J region array (DNA) ctcacgttcggcggagggaccaaggtggagatcaaa (SEQ ID NO: 359) (amino acid) LTFGGGTKVEIK (SEQ ID NO: 1025)

[0350] Humanized 8F9A5A1 light chain variable region sequence (DNA) gaaatagtgatgatgcagtctccagccaccctgtctgtgtctccaggggaaagagccaccctctcctgcagtgccagctcaagtgtaagttacatgaactggtaccagcagaaacctggccaggctcccaggctcctcatctatggtatatccaacctggcttct ggcatcccagccaggttcagtggcagtgggtctgggacagagttcactctcaccatcagcagcctgcagtctgaagattttgcagtttattactgtcagcaaaggagtagttacccacccacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 360) (amino acid) EIVMMQSPATLSVSPGERATLSCSASSSVSYMNWYQQKPGQAPRLLIYGISNLASGIPARFSGSGSGTEFTLTISSLQSEDFAVYYCQQRSSYPPTFGGGTKVEIKR (SEQ ID NO: 1026)

[0351] Humanized 8F9A5A1 light chain variable region sequence (codon optimized) (DNA) gagatcgtgatgatgcagagccccgccacactgagtgtgtctccaggcgaaagagccacactgtcctgtagcgccagcagcagcgtgtcctacatgaactggtatcagcagaagcccggacaggcccctagactgctgatctacggcatcagcaatctggccagc ggcatccctgccagattttctggctctggctccggcaccgagttcaccctgacaatctctagcctgcagagcgaggacttcgccgtgtactactgccagcagagaagcagctaccctcctacctttggcggaggcaccaaggtggaaatcaagcgg (SEQ ID NO: 361) (amino acid) EIVMMQSPATLSVSPGERATLSCSASSSVSYMNWYQQKPGQAPRLLIYGISNLASGIPARFSGSGSGTEFTLTISSLQSEDFAVYYCQQRSSYPPTFGGGTKVEIKR (SEQ ID NO: 1027)

[0352] Modified humanized 8F9A5A1 light chain variable region sequence (DNA) gaaatagtgctgacccagtctccagccaccctgtctgtgtctccaggggaaagagccaccctctcctgcagtgccagctcaagtgtaagttacatgaactggtaccagcagaaacctggccaggctcccaggctctggatctatggtatatccaacctggcttct ggcatcccagccaggttcagtggcagtgggtctgggacaagcttcagcctcaccatcagcagcctgcagtctgaagattttgcagtttattactgtcagcaaaggagtagttacccacccacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 362) (amino acid) EIVLTQSPATLSVSPGERATLSCSASSSVSYMNWYQQKPGQAPRLWIYGISNLASGIPARFSGSGSGTSFSLTISSLQSEDFAVYYCQQRSSYPPTFGGGTKVEIKR (SEQ ID NO: 1028)

[0353] Modified humanized 8F9A5A1 light chain variable region sequence (codon optimized) (DNA) gagatcgtgctgacacagtctcccgccacactgagtgtgtctccaggcgaaagagccacactgtcctgtagcgccagcagcagcgtgtcctacatgaactggtatcagcagaagcccggacaggcccctagactgtggatctacggcatcagcaatctggccagc ggcatccctgccagattttctggctctggctccggcaccagcttcagcctgacaatcagcagcctgcagagcgaggacttcgccgtgtactactgccagcagagaagcagctaccctcctacctttggcggaggcaccaaggtggaaatcaagcgg (SEQ ID NO: 363) (amino acid) EIVLTQSPATLSVSPGERATLSCSASSSVSYMNWYQQKPGQAPRLWIYGISNLASGIPARFSGSGSGTSFSLTISSLQSEDFAVYYCQQRSSYPPTFGGGTKVEIKR (SEQ ID NO: 1029)

[0354] Modified humanized 8F9A5A1 scFv sequence (codon optimized) (DNA) Cagattcagctggtgcagtctggccccgaagtgaaacaacctggcgcctctgtgaaggtgtcctgcaaggccagcggctacacctttaccaactacggcatgaactgggtcaagcaggcccctggacaaggcctggaatggatgggctggatcaacacctacaccggcgagcctacctacgtggacgacttcaagggcagattcgccttcagcatggacaccagcgccagcacagcctacctgcagatcagctctctgaaggccgaggacaccgccacctactactgtgccagactgagaggcatcagacccggacctctggcctattggggacagggaacactggtcaccgtgtcctctggcggtggcggaagcggaggcggtggctccggtggcggaggcagcgagatcgtgctgacacagtctcccgccacactgagtgtgtctccaggcgaaagagccacactgtcctgtagcgccagcagcagcgtgtcctacatgaactggtatcagcagaagcccggacaggcccctagactgtggatctacggcatcagcaatctggccagcggcatccctgccagattttctggctctggctccggcaccagcttcagcctgacaatcagcagcctgcagagcgaggacttcgccgtgtactactgccagcagagaagcagctaccctcctacctttggcggaggcaccaaggtggaaatcaagcgg(SEQ ID NO: 364) (Amino acid) QIQLVQSGPEVKQPGASVKVSCKASGYTFTNYGMNWVKQAPGQGLEWMGWINTYTGEPTYVDDFKGRFAFSMDTSASTAYLQISSLKAEDTATYYCARLRGIRPGPLAYWGQGTLVTVSSGGGGSGGGGSGGGGSEIVLTQSPATLSVSPGERATLSCSASSSVSYMNWYQQKPGQAPRLWIYGISNLASGIPARFSGSGSGTSFSLTISSLQSEDFAVYYCQQRSSYPPTFGGGTKVEIKR (SEQ ID NO: 1030)

[0355] 8H5H5G4 heavy chain variable region sequence (DNA) gtccagctgcaacagtctggacctgatctggtgaagcctggagacttcagtgaagatatcctgtaagacttctggaaacacattcactgaatacaccatgcactgggtgaagcagagccatggaaagagccttgagtggattggaggttttaatcctaacaatggtgttactaactacaaccagaag ttcaagggcaaggccacattgactgtagacaagtcctccagcacagcctacatggagctccgcagcctgacatctgaggattctgcagtctattactgtgcaagacgttactaccatagtacctacgtgttctactttgactcctggggccaaggcaccactctcacagtctcctca (SEQ ID NO: 365) (amino acid) VQLQQSGPDLVKPGTSVKISCKTSGNTFTEYTMHWVKQSHGKSLEWIGGFNPNNGVTNYNQKFKGKATLTVDKSSSTAYMELRSLTSEDSAVYYCARRYYHSTYVFYFDSWGQGTTLTVSS (SEQ ID NO: 1031)

[0356] IGHV1-24 * 01 V region array (DNA) caggtccagctggtacagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaaggtttccggatacaccctcactgaattatccatgcactgggtgcgacaggctcctggaaaagggcttgagtggatgggaggtt ttgatcctgaagatggtgaaacaatctacgcacagaagttccagggcagagtcaccatgaccgaggacacatctacagacacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtattactgtgcaaca (SEQ ID NO: 366) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGYTLTELSMHWVRQAPGKGLEWMGGFDPEDGETIYAQKFQGRVTMTEDTSTDTAYMELSSLRSEDTAVYYCAT (SEQ ID NO: 1032)

[0357] IGHJ4 * 03 J region array (DNA) tactttgactactggggccaagggaccctggtcaccgtctcctca (SEQ ID NO: 367) (amino acid) YFDYWGQGTLVTVSS (SEQ ID NO: 1033)

[0358] Humanized 8H5H5G4 heavy chain variable region sequence (DNA) caggtccagctggtacagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctctcctgcaaggtttccggaaacacattcactgaatacaccatgcacTgggtgcgacaggctcctggaaaagggcttgagtggatgggaggttttaatcctaacaatggtgttactaactacaaccaga agttcaagggcAgagtcaccatgaccgagacacatctacagacacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtattactgtGcaagacgttactaccatagtacctacgtgttctactttgactcctggggccaagggaccctggtcaccgtctcctca (SEQ ID NO: 368) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWMGGFNPNNGVTNYNQKFKGRVTMTEDTSTDTAYMELSSLRSEDTAVYYCARRYYHSTYVFYFDSWGQGTLVTVSS (SEQ ID NO: 1034)

[0359] Humanized 8H5H5G4 heavy chain variable region sequence (codon optimized) (DNA) caggttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggtgtccggaaataccttcaccgagtacaccatgcactgggtccgacaggcccctggcaaaggacttgaatggatgggcggcttcaaccccaacaacggcgtgaccaactacaaccaga aattcaagggccgcgtgaccatgaccgaggacacaagcacagacaccgcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagaaggtactaccacagcacctacgtgttctacttcgacagctggggccagggcacactggtcacagtttcttct (SEQ ID NO: 369) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWMGGFNPNNGVTNYNQKFKGRVTMTEDTSTDTAYMELSSLRSEDTAVYYCARRYYHSTYVFYFDSWGQGTLVTVSS (SEQ ID NO: 1035)

[0360] Modified humanized 8H5H5G4 heavy chain variable region sequence (DNA) caggtccagctggtacagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctctcctgcaaggtttcggaaacacattcactgaatacaccatgcactgggtgcgacaggctcctggaaaagggcttgagtggatcggaggttttaatcctaacaatggtgttactaactacaaccaga agttcaagggcaaggtcaccctgaccgtggacacatctagcagcacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtattactgtgcaagacgttactaccatagtacctacgtgttctactttgactcctggggccaagggaccctggtcaccgtctcctca (SEQ ID NO: 370) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWIGGFNPNNGVTNYNQKFKGKVTLTVDTSSSTAYMELSSLRSEDTAVYYCARRYYHSTYVFYFDSWGQGTLVTVSS (SEQ ID NO: 1036)

[0361] Modified humanized 8H5H5G4 heavy chain variable region sequence (codon optimized) (DNA) caggttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggtgtccggaaataccttcaccgagtacaccatgcactgggtccgacaggcccctggcaaaggactggaatggatcggcggcttcaaccccaacaacggcgtgaccaactacaaccaga aattcaagggcaaagtgaccctgaccgtggacaccagcagcagcacagcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagaaggtactaccacagcacctacgtgttctacttcgacagctggggccagggcacactggtcacagtttcttct (SEQ ID NO: 371) (amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWIGGFNPNNGVTNYNQKFKGKVTLTVDTSSSTAYMELSSLRSEDTAVYYCARRYYHSTYVFYFDSWGQGTLVTVSS (SEQ ID NO: 1037)

[0362] 8H5H5G4 light chain variable region sequence (DNA) gatatccagatgacacagactacatcctccctgtctgcctctctgggagacagagtcaccatcagttgcagtgcaagtcagggcattagcaattatttaaactggtttcagcagaaaccagatggaactattaagctcctgatctattacacatcaagtttacatt caggagtcccatcaaggttcagtggcagtgggtctgggacagattattctctcaccatcagtaatgtggaacctgaagatattgccacttactattgtcagcagtatagtaagcttccttacacgttcggaggggggaccaagctggagataaaacgg (SEQ ID NO: 372) (amino acid) DIQMTQTTSSLSASLGDRVTISCSASQGISNYLNWFQQKPDGTIKLLIYYTSSLHSGVPSRFSGSGSGTDYSLTISNVEPEDIATYYCQQYSKLPYTFGGGTKLEIKR (SEQ ID NO: 1038)

[0363] IGKV1-27 * 01 V region array (DNA) gacatccagatgacccagtctccatcctccctgtctgcatctgtaggagacagagtcaccatcacttgccgggcgagtcagggcattagcaattatttagcctggtatcagcagaaaccagggaaagttcctaagctcctgatctat gctgcatccactttgcaatcaggggtcccatctcggttcagtggcagtggatctgggacagatttcactctcaccatcagcagcctgcagcctgaagatgttgcaacttattactgtcaaaagtataacagtgcccct (SEQ ID NO: 373) (amino acid) DIQMTQSPSSLSASVGDRVTITCRASQGISNYLAWYQQKPGKVPKLLIYAASTLQSGVPSRFSGSGSGTDFTLTISSLQPEDVATYYCQKYNSAP (SEQ ID NO: 1039)

[0364] IGKJ4 * 02 J region array (DNA) ctcacgttcggcggagggaccaaggtggagatcaaa (SEQ ID NO: 374) (amino acid) LTFGGGTKVEIK (SEQ ID NO: 1040)

[0365] Humanized 8H5H5G4 light chain variable region sequence (DNA) gacatccagatgacccagtctccatcctccctgtctgcatctgtaggagacagagtcaccatcacttgcagtgcaagtcagggcattagcaattatttaaacTggtatcagcagaaaccagggaaagttcctaagctcctgatctattacacatcaagtttacatt caggggtcccatctcggttcagtggcagtggatctgggacagatttcactctcaccatcagcagcctgcagcctgaagatgttgcaacttattactgtcagcagtatagtaagcttccttacacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 375) (amino acid) DIQMTQSPSSLSASVGDRVTITCSASQGISNYLNWYQQKPGKVPKLLIYYTSSLHSGVPSRFSGSGSGTDFTLTISSLQPEDVATYYCQQYSKLPYTFGGGTKVEIKR (SEQ ID NO: 1041)

[0366] Modified humanized 8H5H5G4 light chain variable region sequence (codon optimized) (DNA) gacatccagatgacacagagccctagcagcctgtctgccagcgtgggagacagtgaccatcacatgtagcgccagccagggcatcagcaactacctgaactggtatcagcagaaacccggcaaggtgcccaagctgctgatctactacaccagcagcctgcaca gcggcgtgccaagcagattttctggcagcggctctggcaccgacttcaccctgaccatatctagcctgcagcctgaggacgtggccacctactactgtcagcagtacagcaagctgccctacacctttggcggaggcaccaaggtggaaatcaagcgg (SEQ ID NO: 376) (amino acid) DIQMTQSPSSLSASVGDRVTITCSASQGISNYLNWYQQKPGKVPKLLIYYTSSLHSGVPSRFSGSGSGTDFTLTISSLQPEDVATYYCQQYSKLPYTFGGGTKVEIKR (SEQ ID NO: 1042)

[0367] Modified humanized 8H5H5G4 light chain variable region sequence (DNA) gacatccagatgacccagtctccatcctccctgtctgcatctgtaggagacagagtcaccatcacttgcagtgcaagtcagggcattagcaattatttaaactggtatcagcagaaaccagggaaagttcctaagctcctgatctattacacatcaagtttacatt caggggtcccatctcggttcagtggcagtggatctgggacagattacactctcaccatcagcagcctgcagcctgaagatgttgcaacttattactgtcagcagtatagtaagcttccttacacgttcggcggagggaccaaggtggagatcaaacgg (SEQ ID NO: 377) (amino acid) DIQMTQSPSSLSASVGDRVTITCSASQGISNYLNWYQQKPGKVPKLLIYYTSSLHSGVPSRFSGSGSGTDYTLTISSLQPEDVATYYCQQYSKLPYTFGGGTKVEIKR (SEQ ID NO: 1043)

[0368] Modified humanized 8H5H5G4 light chain variable region sequence (codon optimized) (DNA) gacatccagatgacacagagccctagcagcctgtctgccagcgtgggagacagtgaccatcacatgtagcgccagccagggcatcagcaactacctgaactggtatcagcagaaacccggcaaggtgcccaagctgctgatctactacaccagcagcctgcaca gcggcgtgccaagcagattttctggcagcggctctggcaccgactacaccctgaccatatctagcctgcagcctgaggacgtggccacctactactgtcagcagtacagcaagctgccctacacctttggcggaggcaccaaggtggaaatcaagcgg (SEQ ID NO: 378) (amino acid) DIQMTQSPSSLSASVGDRVTITCSASQGISNYLNWYQQKPGKVPKLLIYYTSSLHSGVPSRFSGSGSGTDYTLTISSLQPEDVATYYCQQYSKLPYTFGGGTKVEIKR (SEQ ID NO: 1044)

[0369] Modified humanized 8H5H5G4 scFv sequence (codon optimized) (DNA) Caggttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggtgtccggaaataccttcaccgagtacaccatgcactgggtccgacaggcccctggcaaaggactggaatggatcggcggcttcaaccccaacaacggcgtgaccaactacaaccagaaattcaagggcaaagtgaccctgaccgtggacaccagcagcagcacagcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagaaggtactaccacagcacctacgtgttctacttcgacagctggggccagggcacactggtcacagtttcttctggcggtggcggaagcggaggcggtggctccggtggcggaggcagcgacatccagatgacacagagccctagcagcctgtctgccagcgtgggagacagagtgaccatcacatgtagcgccagccagggcatcagcaactacctgaactggtatcagcagaaacccggcaaggtgcccaagctgctgatctactacaccagcagcctgcacagcggcgtgccaagcagattttctggcagcggctctggcaccgactacaccctgaccatatctagcctgcagcctgaggacgtggccacctactactgtcagcagtacagcaagctgccctacacctttggcggaggcaccaaggtggaaatcaagcgg(SEQ ID NO: 379) (Amino acid) QVQLVQSGAEVKKPGASVKVSCKVSGNTFTEYTMHWVRQAPGKGLEWIGGFNPNNGVTNYNQKFKGKVTLTVDTSSSTAYMELSSLRSEDTAVYYCARRYYHSTYVFYFDSWGQGTLVTVSSGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCSASQGISNYLNWYQQKPGKVPKLLIYYTSSLHSGVPSRFSGSGSGTDYTLTISSLQPEDVATYYCQQYSKLPYTFGGGTKVEIKR (SEQ ID NO: 1045)

[0370] Human IgG1 heavy chain constant region sequence: (for generation of complete antibody - paired with kappa or lambda constant region; 2 plasmids, expressed together) (DNA) (amino acid) ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 1046)

[0371] Human IgG2 heavy chain constant region sequence: (for generation of complete antibody - paired with kappa or lambda constant region; 2 plasmids, expressed together) (DNA) (query number 381) (amino acid) ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 1047)

[0372] Human kappa light chain constant region sequence: (DNA) aggacggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgttgtgtgcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcgggtaactcccaggagagtg tcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctgacgctgagcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagctcgcccgtcacaaagagcttcaacaggggagagtgttag (SEQ ID NO: 382) (amino acid) RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 1048)

[0373] Human lambda light chain constant region sequence: (DNA) ggtcagcccaaggctgccccctcggtcactctgttcccgccctcctctgaggagcttcaagccaacaaggccacactggtgtgtctcataagtgacttctacccgggagccgtgacagtggcctggaaggcagatagcagccccgtcaaggcgggagtggagacc accacaccctccaaacaaagcaacaacaagtacgcggccagcagctatctgagcctgacgcctgagcagtggaagtcccacagaagctacagctgccaggtcacgcatgaagggagcaccgtggagaagacagtggcccctacagaatgttcatag (SEQ ID NO: 383) (amino acid) GQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS (SEQ ID NO: 1049)

[0374] Human IgG1 Fc region sequence: (fused to scFv for homodimerization) (DNA) (query number 384) (amino acid) EPKSCDKTHTCPPCPAPELLGGPSVFLFPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLPGK*(request number 1050)

[0375] Human IgG2 Fc region sequence: (DNA) gagcgcaaatgttgtgtcgagtgcccaccgtgcccagcaccacctgtggcaggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacgtgcgtggtggtggacgtgagccacgaagaccccgaggtccagttcaactggtacgtggacggcgtggaggtgcataatgccaagacaaagccacgggaggagcagttcaacagcacgttccgtgtggtcagcgtcctcaccgttgtgcaccaggactggctgaacggcaaggagtacaagtgcaaggtctccaacaaaggcctcccagcccccatcgagaaaaccatctccaaaaccaaagggcagccccgagaaccacaggtgtacaccctgcccccatcccgggaggagatgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctaccccagcgacatcgccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacacctcccatgctggactccgacggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcatgctccgtgatgcatgaggctctgcacaaccactacacgcagaagagcctctccctgtctccgggtaaatag (SEQ ID NO: 385) (Amino acid) ERKCCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* (SEQ ID NO: 1051)

[0376] In another embodiment of the invention, immune cells engineered to express a CAR are administered to a patient diagnosed with or at risk of developing cancer or cancer metastasis, and the immune cells are also engineered to express an anti-NME7 antibody or antibody fragment, which may be expressed from an inducible promoter. * The CAR is derived from an antibody fragment. In one example, the CAR is huMNC2-CAR44. In one embodiment, the anti-NME7 antibody or antibody fragment binds to an NME7 peptide listed above in the sections "Peptides Homologous to A1, A2, B1, B2, or B3 Peptides" and "Extended Peptides Homologous to A1, A2, B1, B2, or B3 Peptides." In another embodiment, the antibody or antibody fragment binds to an NME7-derived peptide selected from A1, A2, B1, B2, or B3 (SEQ ID NOs: 141-145). In yet another embodiment, the antibody, antibody fragment, or antibody mimetic binds to an NME7 peptide, including the B3 peptide. In one embodiment of the invention, the anti-NME7 antibody, antibody fragment or antibody mimetic comprises a sequence derived from the variable domain of anti-NME7 antibody 8F9A4A3 (SEQ ID NOs: 1001-1015), 8F9A5A1 (SEQ ID NOs: 1016-1030), or 8H5H5G4 (SEQ ID NOs: 1031-1045).

[0377] Such antibodies can be human or humanized. They can be polyclonal, monoclonal, bispecific, bivalent, monovalent, single-chain, scFv, or antibody mimics, such as protein scaffolds, that provide recognition regions for binding to specific targets. As will be appreciated by those skilled in the art, antibodies can be of non-human origin, human, or humanized. Methods for humanizing antibodies include fusing all or part of the murine variable region to the V and J regions of the closest-matched human antibody sequence, such as those shown in SEQ ID NOS: 1001-1045. Full antibodies, rather than single-chain constructs, can also be generated. For example, a heavy chain variable murine sequence is fused to human V and J regions, which are then fused to a human heavy chain constant region for IgG1, IgG2, or IgG3. Similarly, a light chain variable murine sequence is fused to human V and J regions, which are then fused to a human kappa or lambda constant region for IgG1, IgG2, or IgG3. The plasmids are expressed together and combined to form a full antibody (SEQ ID NOS: 1047-1051).

[0378] In another aspect of the invention, the small molecule is NME7, NME7 AB or anti-cancer agents selected for their ability to inhibit the tumorigenic effects of NME7-X1. For example, high-throughput screening can identify small molecules that treat cancer. In multi-well plates, small molecules are screened to identify small molecules in which cancer cells express NME7-X1. AB The small molecules are then added separately to wells where the cells are cultured in a medium containing NME7. If the small molecule reduces the amount of cells that float and / or reduces the expression of metastasis markers or pluripotent stem cell markers, such as CXCR4 and CHD1, the small molecule is a potential anti-cancer drug. Another method for identifying small molecules that are anti-cancer drugs is to use NME7. AB Alternatively, small molecules that bind to NME7-X1 or inhibit the expression of NME7 species can be selected. * NME7 to PSMGFR peptide in the extracellular domain AB The goal is to select small molecules that inhibit the binding of these small molecules, which are anti-cancer drugs.

[0379] NME7 AB The sequence of NME7-X1 is AB The only difference is that it lacks some of the N-terminal sequence found in NME7. AB We have shown that there are seven naturally occurring NMEs that are nearly identical to NMEs. AB Antibodies that bind to NME7-X1 are called NME7 variants unless there is a conformational difference that the antibody can distinguish. AB It can also bind to naturally occurring species that mimic NME7. AB If it is desired to inhibit NME7-X1 but not other species, or vice versa, siRNA, antisense nucleic acid, or gene editing techniques can be used to inhibit expression in one species but not the other.

[0380] In one example, the anti-cancer therapeutic agent is NME7, NME7-X1 or NME7 AB The therapeutic agent is a nucleic acid that directly or indirectly suppresses the specific expression of various species. Such nucleic acids can be siRNA, RNAi, antisense nucleic acids, etc. that directly suppress NME7 species. In another embodiment of the present invention, the nucleic acid can indirectly suppress NME7 species, for example, by altering the expression of a molecule that regulates it. For example, the super enhancer BRD4 suppresses NME7 expression. Therefore, an effective therapeutic agent for treating or preventing cancer is an agent that increases BRD4 expression. An effective therapeutic agent can also be an agent that increases the expression of JMJD6, a cofactor of BRD4.

[0381] NME7 AB Alternatively, NME7-X1 derived peptides or the entire protein can be used to generate anti-NME7 and anti-NME7-X1 antibodies in animals, which we have demonstrated inhibit cancer growth and inhibit the migration of cancer cells to metastatic cancer cells. Similarly, NME7 derived peptides can be administered to humans to generate antibodies that treat or prevent cancer or inhibit the migration of cancer cells to metastatic cancer cells. The NME7 peptide or protein can be administered to recipients to generate anti-NME7 and anti-NME7-X1 antibodies in recipients. ABThe results shown in Figures 12 and 13 demonstrate that the NME7-X1 or NME7-X1 antibodies derived from NME7 are specifically expressed in the NME7-X1 or NME7-X1-derived antibodies. AB It has been shown that immunizing humans with a collection of peptides in a sequence may be a more effective vaccine than immunization with a single peptide. The peptide or protein may be further conjugated to a carrier protein or other adjuvant known to those skilled in the art to help stimulate an immune response.

[0382] NME7 peptides located outside the DM10 domain are preferred for generating antibodies for treating or preventing cancer. Peptides that can be administered to patients for the prevention of cancer or metastasis include the peptide sequences listed in Figures 6-9. A1, A2, B1, B2, and B3 are NME7 peptides. AB and NME7-X1, and are examples of peptides that bind to NME7-X1 and generate antibodies that can be administered to patients for the treatment or prevention of cancer. The present invention is not limited to peptides of the exact sequence, such as the naturally occurring sequence of NME7 or NME7-X1. As known to those skilled in the art, substitution of some amino acids in the peptide sequence can still generate antibodies that specifically recognize the native protein sequence. It is not intended that the present invention be limited to the peptides demonstrated herein that inhibit cancer growth or inhibit the transition of normal cancer cells to metastatic cancer cells. The methods used herein to identify peptides A1, A2, B1, B2, and B3 can also be used to identify other peptide sequences that may be equally or more effective than the peptides demonstrated herein.

[0383] Human NME7 AB or includes part of the NME7-X1 or NME7 AB Alternatively, chimeric antigen receptor molecules comprising antibody fragments that bind to NME7-X1 are anti-cancer therapeutic agents and are administered to patients to treat or prevent cancer or cancer metastasis.

[0384] In one example, the recognition unit or variable region of an anti-NME7 antibody is fused to a molecule on a T cell using a technique known as CAR (chimeric antigen receptor) or CAR T technology. A hallmark of antibodies or fragments thereof that can be used therapeutically to treat or prevent cancer is the identification of an antibody-like variable region that recognizes NME7 and prevents its interaction with cancer-promoting targets. In one example, the target is MUC1 * This is the PSMGFR region.

[0385] Antibodies, antibody fragments, or single-chain antibodies can be engineered into chimeric molecules containing chimeric antigen receptors, also known as CARs, which are then transduced or transduced into immune system cells such as T cells and administered to patients. A humanized antibody or antibody fragment, typically an scFv, contains much of the extracellular domain of the CAR. The antibody fragment is biochemically fused to immune system signaling molecules, such as CD8 as a transmembrane domain, a cytoplasmic signaling motif, also known as an activation domain, of T cell receptor signaling molecules, or costimulatory domains, including but not limited to CD3 zeta, CD28, 41bb, and OX40. The CAR can be transduced into T cells or other cells, preferably immune system cells, and administered to patients. Here, we describe a chimeric antibody fragment in which the extracellular portion is an anti-NME7 or anti-NME7 antibody. AB Alternatively, a CAR is described that comprises an anti-NME7-X1 antibody, antibody fragment, or single-chain, scFv antibody fragment. In a preferred embodiment, the antibody or antibody fragment is a human or humanized antibody.

[0386] Effective anti-NME7 or anti-NME7-X1 antibodies, or fragments, include those derived from native NME7, NME7 AB or NME7-X1. Indeed, the parent antibody from which the extracellular domain of the CAR is engineered is NME7, NME7 ABor by immunizing an animal with an NME7-X1 derived peptide. In one embodiment of the invention, the immunizing peptide consists of NME7 amino acids 1-376. In one embodiment of the invention, the immunizing peptide consists of NME7 amino acids 92-376. In another embodiment of the invention, the immunizing peptide consists of NME7 amino acids 125-376. In yet another embodiment of the invention, the immunizing peptide consists of the sequence listed in Figures 6-8. In another embodiment of the invention, the immunizing peptide consists of the sequence listed in Figure 9. Alternatively, parent antibodies or antibody fragments are selected from an antibody library or pool, which may be natural, synthetic, or fragments of either, and which may be NME7, NME7 AB or NME7-X1, a peptide listed in FIGS. 6-8 or a peptide listed in FIG.

[0387] The targeting portion of a CAR does not have to be an antibody or antibody fragment. Here we describe a CAR whose extracellular domain comprises an NME7 fragment. NME7-derived peptides have been engineered into different types of CARs in which the targeting portion of the extracellular domain is a protein fragment or peptide, rather than an antibody or antibody fragment. Peptide CARs are transduced or transduced into immune system cells, typically T cells. NME7 fragments or NME7-derived peptides bind to their cognate binding partners, but do not bind to intact NME7, NME7. AB NME7 fragments or NME7-derived peptides are selected for their ability to function like NME7-X1 and to confer oncogenic activity. NME7 fragments or NME7-derived peptides are biochemically fused to immune system signaling molecules such as CD8 as a transmembrane domain, cytoplasmic signaling motifs such as T cell receptor signaling molecules, also known as activation domains, or costimulatory domains, including but not limited to CD3 zeta, CD28, 41bb, and OX40.

[0388] In one embodiment of the invention, the NME7 fragment is most or all of the NME7 NDPK B domain. In another embodiment of the invention, the NME7 fragment is an NME7 peptide comprising one or more of the peptide sequences listed in Figures 6-9. Experimental results support the use of NME7 or NME7, NME7 as a targeting moiety in a chimeric antigen receptor (CAR) for engineered immune cell therapy. AB or in the case of a strategy using a fragment of NME7-X1, NME7 AB Alternatively, larger fragments of NME7-X1 may be more effective than shorter peptides, e.g., peptides less than 15 amino acids in length. AB Collectively, a panel of CARs bearing derived peptides can be transduced into immune system cells and administered to patients for cancer treatment or prevention. Experiments shown in Figures 12 and 13 support the validity of this approach.

[0389] A CAR comprising an NME7 fragment in its extracellular domain is transduced or transduced into immune system cells, typically T cells, and administered to a patient for the treatment or prevention of cancer. * In another embodiment, the cancer is a metastatic cancer.

[0390] Drugs that inhibit the enzyme that cleaves NME7 can be used to treat or prevent cancer. Some forms of NME7 are sequestered intracellularly and therefore not secreted from cells, where they can act as growth factors to promote cancer. Full-length NME7 is 42 kDa. However, we have found that the recombinant NME7 we generated lacks the DM10 domain and is a 42 kDa mutant. AB We found that an approximately 33 kDa NME7 species, which appears to be virtually identical to NME7, is secreted from cancer cells and stem cells. This approximately 33 kDa NME7 species and another approximately 25 kDa NME7 species may be cleavage products removed by the use of agents that inhibit NME7 cleavage.

[0391] High levels of NME7, an approximately 33 kDa NME7 species (which we refer to as NME7), were observed in patient samples. AB Detection of NME7-X1, or NME7-X1, is an indicator of the presence of cancer or its progression to a more aggressive or metastatic state. The present inventors have demonstrated that the expression of NME7-X1 in both naive stem cells and cancer cells, particularly MUC1, is essential for the development of cancer-like tumors. * We found that positive cancer cells expressed high levels of approximately 33 kDa NME7 and NME7-X1, which lack the DM10 domain.

[0392] NME7-X1 was recently listed in protein databases as a theoretical alternative isoform of NME7. However, it has not been detected in tissues or cells. We designed primers to distinguish NME7-X1 from NME7 by PCR. Expression levels of human NME7, NME7a, NME7b, and NME7-X1 were measured by PCR in a range of cells, including fibroblasts, human embryonic stem cells, human iPS cells, T47D human breast cancer cells, DU145 human prostate cancer cells, PC3 human prostate cancer cells, HEK295 human fetal liver cells, and other human stem cell lines. NME7 is expressed at higher levels in cancer cells than in stem cells. In particular, NME7-X1 is expressed 10-fold higher in prostate cancer cells and 3-fold higher in breast cancer cells than in fibroblasts or stem cells. NME7-X1 is expressed approximately 5-fold higher in HEK293 fetal liver cells than in fibroblasts or stem cells, and therefore NME7-X1 is expected to be elevated in liver cancer. NME7b is expressed 17-25-fold higher in prostate cancer cells than in stem cells.

[0393] Detection of high levels of NME7 species in a patient sample is an indication that the patient has cancer or is at risk of developing cancer. NME7 species levels can be measured or evaluated by PCR, hybridization schemes, cycling probe technology, FISH, immunocytochemistry, IHC, Western blot, immunoprecipitation, sandwich analysis, ELISA assay, etc. Patient samples can be liquid samples, blood samples, milk, urine, cells, liquid biopsies, biopsies, etc. In patients diagnosed with cancer, high levels of NME7 species are an indicator of increased metastatic potential. High levels of NME7-X1 are an indicator of prostate cancer. The antibodies of the present invention are used to detect and distinguish NME7 species and are used as diagnostic tools.

[0394] Because adult human cells and tissues do not express or secrete significant levels of NME7, an effective method for diagnosing cancer or diagnosing a shift to more aggressive or metastatic forms or a shift to more aggressive forms is to measure the level of NME7 in a sample from a patient, from a cell or tissue collection, or from cultured cells, relative to the level of NME7 in a healthy sample and / or relative to the level of NME7 known to be present in healthy adult cells or tissues. Increased levels of NME7 indicate the presence of cancer, the presence of metastatic cancer, or the onset of metastasis. Increased levels of NME7 also indicate the presence of MUC1. *indicates positive cancer. The sample assayed for the presence of NME7 can be a collection of cells, which can be cells from a patient, a cultured cell line, a body fluid, a blood sample, a tissue specimen, or a biopsy. Thus, a diagnostic assay to detect the presence of cancer or the progression of cancer includes the following steps: 1) obtaining a sample from a cancer patient or from a patient at risk of developing cancer; 2) subjecting the sample to an assay capable of detecting or measuring the level of NME7 or the level of a nucleic acid encoding NME7; 3) comparing the measured level of NME7 protein or NME7-encoding nucleic acid in the test sample to that of a control patient or control cell; 4) identifying that the level of NME7 or the level of a nucleic acid encoding NME7 is elevated compared to the control; and 5) concluding that the cancer is progressing if the donor of the test sample is afflicted with cancer or the control to which the test article is compared is from a donor previously diagnosed with cancer.

[0395] In this assay, the control sample to which the test sample is compared can be noncancerous cells, cultured cells, a sample from a healthy donor, a noncancerous sample from a donor, or a sample from the donor of the test sample that is a control sample taken from the donor at an earlier time point. Such sample sources can be any specimen taken from a patient being tested for the presence or progression of cancer, including bodily fluids, cerebrospinal fluid, bone marrow samples, blood, tissues, cells, biopsy tissue or cells, cultured cells derived from patient cells, etc. The sample source to which the test sample is compared can be bodily fluids, cerebrospinal fluid, bone marrow samples, blood, tissues, cells, biopsy tissue or cells, or cultured cells derived from a healthy donor or a test patient from whom the sample was obtained at an earlier time point. The measured levels to which the test sample is compared can be from previously recorded data and compiled in a list for comparison to the test sample.

[0396] Theranostics Patients diagnosed with elevated levels of NME7 protein or nucleic acid encoding NME7 are then treated with therapeutic agents that reduce NME7 expression, inhibit NME7 cleavage, or inhibit NME7 binding to its targets (such interactions promote cancer). An important target or cleavage product of NME7 is MUC1. * NME7 is a MUC1 * The cleaved form of MUC1 binds to and dimerizes the extracellular domain of NME7. Thus, patients diagnosed with elevated levels of NME7 may benefit from treatment with a therapeutic agent that inhibits NME7 and / or inhibits the dimerization of a truncated form of MUC1 whose extracellular domain consists of some or all of the PSMGFR sequence. Thus, assessing the suitability of cancer therapy and administering an effective amount of a therapeutic agent for treating or preventing cancer comprises the following steps: 1) obtaining a sample from a patient suspected of having cancer, or at risk of developing cancer, or at risk of developing metastatic cancer; 2) measuring the amount of NME7, or a cleavage product thereof, or an NME7-encoding nucleic acid, where the measured level is significantly higher than the amount measured in a control sample; 3) identifying the patient as having cancer or having progressed to a more aggressive or metastatic cancer; 4) administering to the patient an effective amount of a therapeutic agent that inhibits expression of NME7, inhibits cleavage of NME7, or inhibits binding of NME7 to its target, and / or inhibits expression of MUC1. * inhibiting MUC1 cleavage to its target or * administering to the patient an effective amount of a therapeutic agent that inhibits NME7 binding to its target. In a preferred embodiment, the therapeutic agent that inhibits NME7 binding to its target is MUC1 * In a more preferred embodiment, it inhibits its interaction with MUC1, which is substantially composed of a PSMGFR sequence. * In a preferred embodiment, the target is a polypeptide that inhibits its interaction with the extracellular domain of MUC1. * Therapeutic drugs that inhibit binding are MUC1 * In a more preferred embodiment, the interaction between MUC1 and NME7 is inhibited. *Therapeutic agents that inhibit the interaction between NME7 and NME7 AB MUC1 to a portion of NME7 consisting essentially of the sequence * inhibits the binding of

[0397] Chemically modified peptides Polypeptide or antibody therapeutic drugs have problems with short circulation half-life, proteolysis, and low solubility.To improve the pharmacokinetic and pharmacodynamic properties of the biopharmaceuticals of the present invention, methods such as manipulating amino acid sequences can reduce or increase immunogenicity and reduce proteolytic cleavage; peptides can be fused or conjugated to immunoglobulins such as albumin and serum proteins; biopharmaceuticals such as the peptides and antibodies of the present invention can also be incorporated into drug delivery carriers for protection and sustained release; and conjugation to natural or synthetic polymers can also be considered.In particular, for synthetic polymer conjugation, PEGylation or acylation such as N-acylation, S-acylation, etc. can also be considered.

[0398] nucleic acid construct Also provided are expression vectors comprising the nucleic acid molecules of the invention as described herein, wherein the nucleic acid molecules are operably linked to an expression control sequence. Also provided are host-vector systems for producing polypeptides comprising the expression vectors of the invention introduced into a host cell suitable for expression of the polypeptide. Suitable host cells can be bacterial cells such as E. coli, yeast cells such as Pichia pastoris, insect cells such as Spodoptera frugiperda, or mammalian cells such as COS, HEK, or CHO cells.

[0399] The invention also provides methods for producing the polypeptides of the invention by growing cells of the host-vector systems described herein under conditions that allow for the production of the polypeptide, and methods for recovering the polypeptides so produced. Polypeptides useful for practicing the invention can be prepared by expression in prokaryotic or eukaryotic expression systems.

[0400] The recombinant gene is expressed and the polypeptide is purified using a number of methods. The gene can be subcloned into a bacterial expression vector, such as, but not limited to, pZErO.

[0401] The polypeptide can be purified by any technique that allows the subsequent formation of a stable biologically active protein.For example, but not limited to, the factor can be recovered from cells either as a soluble protein or as an inclusion body, from which it is quantitatively extracted with 8M guanidinium hydrochloride and dialyzed.To further purify the factor, numerous purification methods can be used, including but not limited to, conventional ion exchange chromatography, affinity chromatography, different sugar chromatography, hydrophobic interaction chromatography, reversed-phase chromatography, or gel filtration.

[0402] As used herein, a polypeptide includes functionally equivalent molecules in which an amino acid residue is substituted for another within the sequence, resulting in a silent or conservative change. For example, one or more amino acid residues within the sequence can be substituted by another amino acid of a similar polarity, which acts as a functional equivalent, resulting in a silent or conservative change. Substitutes for an amino acid within the sequence can be selected from other members of the class to which the amino acid belongs. For example, nonpolar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, and methionine. Polar-neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine. Positively charged (basic) amino acids include arginine, lysine, and histidine. Negatively charged (acidic) amino acids include aspartic acid and glutamic acid. Potential glycosylated amino acids include serine, threonine, and asparagine. Also included within the scope of the present invention are proteins or fragments or derivatives thereof that exhibit the same or similar biological activity, and derivatives that are differentially modified during or after translation, for example, by glycosylation, proteolytic cleavage, linkage to antibody molecules or other cellular ligands, etc.

[0403] Any method known to those skilled in the art for inserting a DNA fragment into a vector can be used to construct an expression vector encoding the polypeptide of the present invention using appropriate transcriptional / translational control signals and protein-coding sequences. These methods can include in vitro recombinant DNA and synthetic techniques, and in vivo recombination (genetic recombination). The expression of the nucleic acid sequence encoding the polypeptide of the present invention can be regulated by a second nucleic acid sequence, thereby expressing the polypeptide in a host transformed with the recombinant DNA molecule. For example, the expression of the polypeptide described herein can be controlled by any promoter / enhancer sequence known in the art. Promoters that can be used to control the expression of a polypeptide include, but are not limited to, the long terminal repeat described in Squinto et al. (1991, Cell 65:1-20); the SV40 early promoter region (Bernoist and Chambon, 1981, Nature 290:304-310), the CMV promoter, the M-MuLV 5' terminal repeat, the promoter contained in the 3' long terminal repeat of Rous sarcoma virus (Yamamoto, et al., 1980, Cell 22:787-797), the herpes thymidine kinase promoter (Wagner et al., 1981, Proc. Natl. Acad. Sci. USA 78:144-1445), the regulatory sequence of the metallothionein gene (Brinster et al., 1982, Nature 296:39-42); prokaryotic expression vectors such as the β-lactamase promoter (Villa-Kamaroff, et al., 1978, Proc. Natl. Acad. Sci. USA 75:3727-3731), or the tac promoter (DeBoer, et al., 1983, Proc. Natl. Acad. Sci. USA80:21-25; see also "Useful proteins from recombinant bacteria" in Scientific American, 1980, 242:74-94; promoter sequences derived from yeast or other fungi, e.g., the gal4 promoter, the ADH (alcohol dehydrogenase) promoter, the PGK (phosphoglycerate kinase) promoter, the alkaline phosphatase promoter, and the following animal transcriptional control regions that exhibit tissue specificity and have been used in transgenic animals: the elastase I gene control region, which is active in pancreatic acinar cells (Swift et al., 1984, Cell 38:639-646; Ornitz et al., 1986, Cold Spring Harbor Symp. Quant. Biol. 50:399-409; MacDonald, 1987, Hepatology 7:425-515); the insulin gene control region, which is active in pancreatic β-cells (Hanahan, 1985, Nature 315:115-122), immunoglobulin gene control region active in lymphoid cells (Grosschedl et al., 1984, Cell 38:647-658; Adames et al., 1985, Nature 318:533-538; Alexander et al., 1987, Mol. Cell. Biol. 7:1436-1444), mouse mammary tumor virus control region active in testicular, breast, lymphoid, and mast cells (Leder et al., 1986, Cell 45:485-495), albumin gene control region active in Sendai virus, lentivirus, and liver (Pinkert et al., 1987, Genes and Devel. 1:268-276), alpha-fetoprotein gene control region active in liver (Krumlauf et al., 1987, Genes and Devel. 1:268-276), al., 1985, Mol. Cell. Biol. 5:1639-1648; Hammer et al., 1987, Science 235:53-58); the alpha 1-antitrypsin gene control region active in the liver (Kelsey et al., 1987, Genes and Devel.1:161-171), the beta-globin gene control region active in myeloid cells (Mogram et al., 1985, Nature 315:338-340; Kollias et al., 1986, Cell 46:89-94); the myelin basic protein gene control region active in oligodendrocyte cells of the brain (Readhead et al., 1987, Cell 48:703-712); the myosin light chain-2 gene control region active in skeletal muscle (Shani, 1985, Nature 314:283-286), and the gonadotropin-releasing hormone gene control region active in the hypothalamus (Mason et al., 1986, Science 234:1372-1378).

[0404] Thus, according to the invention, expression vectors capable of replicating in a bacterial or eukaryotic host, which contain nucleic acids encoding the polypeptides described herein, are used to transfect the host, thereby inducing expression of such nucleic acids to produce polypeptides which can be recovered in a biologically active form. As used herein, a biologically active form includes a form which is capable of binding to the relevant receptor, effecting a differentiated function, and / or affecting the phenotype of cells expressing the receptor.

[0405] Expression vectors containing nucleic acid inserts can be identified by at least three general techniques: (a) DNA-DNA hybridization, (b) the presence or absence of a "marker" gene function, and (c) expression of the inserted sequence. In the first technique, the presence of a heterologous nucleic acid inserted into an expression vector can be detected by DNA-DNA hybridization using a probe containing a sequence homologous to the inserted nucleic acid sequence. In the second technique, recombinant vector / host systems can be identified and selected based on the presence or absence of a specific "marker" gene function (e.g., thymidine kinase activity, antibiotic resistance, transformed phenotype, occlusion body formation in baculovirus, etc.) caused by the insertion of the heterologous nucleic acid sequence into the vector. For example, if an efl nucleic acid sequence is inserted within the marker gene sequence of a vector, recombinants containing the insert can be identified by the absence of the marker gene function. In the third technique, recombinant expression vectors can be identified by assaying for the heterologous nucleic acid product expressed by the recombinant construct. Such assays can be based, for example, on the physical or functional properties of the nucleic acid product of interest, such as, for example, binding of a ligand to a receptor or portion thereof which can be tagged with a detectable antibody or portion thereof, or by binding to an antibody raised against the protein or portion thereof of interest.

[0406] Polypeptides, particularly modified polypeptides of the present invention, can be expressed transiently, constitutively or permanently in host cells.

[0407] The effective amount useful for treating the diseases or disorders described herein can be determined by methods known to those skilled in the art (e.g., Fingl, et al., The Pharmacological Basis of Therapeutics, Goodman and Gilman, eds. Macmillan Publishing Co., New York, pp. 1-46 (1975)). Pharmaceutical compositions for use according to the present invention comprise the polypeptides described above in a pharmaceutically acceptable liquid, solid, or semi-solid carrier, linked to a carrier or targeting molecule (e.g., an antibody, hormone, growth factor, etc.), and / or incorporated into liposomes, microcapsules, and sustained-release formulations prior to in vivo administration. For example, pharmaceutical compositions can comprise the polypeptide in an aqueous solution such as sterile water, saline, phosphate buffer, or dextrose solution. Alternatively, the active agent can be contained in a solid (e.g., wax) or semi-solid (e.g., gelatin) formulation that can be implanted in a patient in need of such treatment. The route of administration may be any method known in the art, including, but not limited to, intravenous, intrathecal, subcutaneous, intrauterine, injection into diseased tissue, intraarterial, intranasal, oral, or via an implanted device.

[0408] Administration can result in the distribution of the active agent of the present invention throughout the body or in a localized area. For example, in some conditions involving remote areas of the nervous system, intravenous or intrathecal administration of the agent may be desirable. In some cases, an implant containing the active agent can be placed at or near the damaged area. Suitable implants include, but are not limited to, gelfoam, wax, spray, or microparticle-based implants.

[0409] The present invention also provides pharmaceutical compositions comprising the polypeptides described herein in a pharmaceutically acceptable carrier. The compositions can be administered systemically or locally. Any administration method known in the art can be used, including but not limited to, intravenous, intrathecal, intraarterial, intranasal, oral, subcutaneous, intraperitoneal, or local injection or surgical implant. Sustained-release formulations are also provided.

[0410] Gene therapy Gene therapy refers to therapy performed by the administration to a subject of expressed or expressible nucleic acids. In this embodiment of the invention, the nucleic acids produce their encoded proteins that mediate a therapeutic effect.

[0411] Any of the methods for gene therapy available in the art can be used in accordance with the present invention. Exemplary methods are described below.

[0412] For reviews of gene therapy, see Goldspiel et al., Clinical Pharmacy 12:488-505 (1993); Wu and Wu, Biotherapy 3:87-95 (1991); Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993); Mulligan, Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev. Biochem. 62:191-217 (1993); May, TIBTECH 11(5):155-215 (1993). Methods generally known in the field of recombinant DNA technology that can be used are described in Ausubel et al. (eds), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); and Kriegler, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990).

[0413] Delivery of nucleic acids to a patient can be direct, where the patient is directly exposed to the nucleic acid or nucleic acid-carrying vector, or indirect, where cells are first transformed in vitro with the nucleic acid and then transplanted into the patient. These two approaches are known, respectively, as in vivo or ex vivo gene therapy.

[0414] In certain embodiments, nucleic acid sequences are directly administered in vivo, where they are expressed to produce the encoded product. This can be achieved by any of a number of methods known in the art, such as by constructing them as part of a suitable nucleic acid expression vector and administering them so that they become intracellular; by infection with a defective or weakened retrovirus or other viral vector, by direct injection of naked DNA, by coating with lipids or cell surface receptors or gene transfer agents, by encapsulation in liposomes, microparticles, or microcapsules, or by administering them linked to peptides known to enter the nucleus, by administering them linked to a ligand that is subject to receptor-mediated endocytosis (see, e.g., Wu and Wu, J. Biol. Chem. 262:4429-4432 (1987)), which can be used to specifically target cell types that express the receptor, etc. In another embodiment, a nucleic acid-ligand complex is formed in which the ligand comprises a fusogenic viral peptide that disrupts endosomes, allowing the nucleic acid to avoid lysosomal degradation. In another embodiment, nucleic acids can be targeted in vivo for cell-specific uptake and expression by targeting specific receptors. Alternatively, nucleic acids can be introduced inside cells and integrated into host cell DNA for expression by homologous recombination (Koller and Smithies, Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); Zijlstra et al., Nature 342:435-438 (1989)).

[0415] In certain embodiments, viral vectors containing nucleic acid sequences encoding polypeptides are used. The nucleic acid sequences encoding the polypeptides used in gene therapy are cloned into one or more vectors, which facilitates the delivery of the gene into patients. Lentiviral vectors, such as retroviral vectors, and other vectors, such as adenoviral vectors and adeno-associated viruses, are examples of viral vectors that can be used. Retroviral vectors contain the components necessary for the correct packaging of the viral genome and integration into host cell DNA.

[0416] Adenoviruses are particularly attractive carriers for delivering genes to respiratory epithelia because they naturally infect respiratory epithelia, causing mild disease. Other targets for adenovirus-based delivery systems are the liver, central nervous system, endothelial cells, and muscle. Adenoviruses have the advantage of being able to infect non-dividing cells. In addition, adeno-associated viruses (AAVs) have also been proposed for use in gene therapy.

[0417] Another approach to gene therapy involves transferring a gene to cells in tissue culture by methods such as electroporation, lipofectin, calcium phosphate-mediated transfection, or viral infection. Usually, the method of transfer includes the transfer of a selectable marker to the cells. The cells are then subjected to selection to isolate those cells that have taken up and are expressing the transgene. These cells are then delivered to the patient.

[0418] In this embodiment, the nucleic acid is introduced into the cell prior to in vivo administration of the resulting recombinant cell. Such introduction can be carried out by any method known in the art, including, but not limited to, transfection, electroporation, microinjection, infection with a viral or bacteriophage vector containing the nucleic acid sequence, cell fusion, chromosome-mediated gene transfer, microcell-mediated gene transfer, spheroplast fusion, etc. Numerous techniques for introducing foreign genes into cells are known in the art and can be used in accordance with the present invention, so long as the necessary developmental and physiological functions of the recipient cell are not disrupted. The technique should provide for the stable transfer of the nucleic acid into the cell, so that the nucleic acid is expressible by the cell and, preferably, heritable and expressible by its cell progeny.

[0419] Cells into which nucleic acids can be introduced for purposes of gene therapy include any desired available cell type, including, but not limited to, epithelial cells, endothelial cells, keratinocytes, fibroblasts, myocytes, hepatocytes; blood cells such as T lymphocytes, B lymphocytes, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes, granulocytes; various stem or progenitor cells, particularly hematopoietic stem or progenitor cells, obtained, for example, from bone marrow, umbilical cord blood, peripheral blood, fetal liver, etc.

[0420] In a preferred embodiment, the cells used in gene therapy are autologous to the patient.

[0421] In embodiments in which recombinant cells are used in gene therapy, nucleic acid sequences encoding polypeptides are introduced into cells so that they can be expressed by the cells or their progeny, and the recombinant cells are then administered in vivo for therapeutic effect. In certain embodiments, stem or progenitor cells are used. Any stem and / or progenitor cells that can be isolated and maintained in vitro can potentially be used in accordance with this embodiment of the invention.

[0422] In certain embodiments, the nucleic acid introduced for purposes of gene therapy comprises an inducible promoter operably linked to the coding region, whereby expression of the nucleic acid can be controlled by regulating the presence or absence of the appropriate transcription inducer.

[0423] therapeutic composition Formulations of therapeutic compounds are generally known in the art, and reference may be made conveniently to Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Co., Easton, Pa., USA. For example, about 0.05 ng to about 20 mg per kg of body weight per day may be administered. The dosage regimen may be adjusted to obtain the optimal therapeutic response. For example, several divided doses may be administered daily, or the dosage may be proportionally reduced as indicated by the exigencies of the therapeutic situation. The active compound may be administered by any convenient route, such as oral, intravenous (if water-soluble), intramuscular, subcutaneous, intranasal, intraocular, intradermal, or suppository routes, or by implantation (e.g., intraperitoneal route using sustained-release molecules, or by using cells, such as monocytes or dendritic cells, sensitized in vitro and adoptively transferred into the recipient). Depending on the route of administration, the peptide may need to be coated with a material to protect it from the action of enzymes, acids, and other natural conditions that may inactivate the component.

[0424] For example, the low lipophilicity of peptides allows them to be destroyed in the digestive tract by enzymes capable of cleaving peptide bonds or in the stomach by acid hydrolysis. To administer peptides by methods other than parenteral administration, the peptides are coated with or administered with a substance that prevents their inactivation. For example, peptides can be administered in adjuvants, co-administered with enzyme inhibitors, or in liposomes. Adjuvants contemplated herein include resorcinol; non-ionic surfactants such as polyoxyethylene oleyl ether and n-hexadecyl polyethylene ether. Enzyme inhibitors include pancreatic trypsin inhibitor, diisopropyl fluorophosphate (DEP), and trasylol. Liposomes include water-in-oil-in-water CGF emulsions as well as conventional liposomes.

[0425] Active compound can also be administered parenterally or intraperitoneally.Dispersion can also be prepared in glycerol, liquid polyethylene glycol and their mixture, and in oil.Under normal conditions of storage and use, these preparations contain preservatives to prevent the growth of microorganisms.

[0426] Dosage forms suitable for injectable use include sterile aqueous solutions (where soluble) or dispersions and sterile powders for extemporaneous preparation in sterile injectable solutions or dispersions. In all cases, the form must be sterile and fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetable oils. Proper fluidity can be maintained by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. Prevention of the action of microorganisms can be brought about by various antibacterial and antifungal agents, such as chlorobutanol, phenol, sorbic acid, thiomersal, and the like. In many cases, it is preferable to include an isotonic agent, such as sucrose or sodium chloride. Prolonged absorption of the injectable compositions can be brought about by the use in the compositions of agents delaying absorption, for example, aluminum monostearate and gelatin.

[0427] Sterile injectable solutions are prepared by incorporating the active compound in the required amount in a suitable solvent with various other ingredients as listed above, followed by sterile filtration as needed. Generally, dispersions are prepared by incorporating various sterile active ingredients into a sterile carrier containing a basic dispersion medium and the necessary other ingredients selected from those listed above. In the case of sterile powders for preparing sterile injectable solutions, the preferred preparation method is vacuum drying and freeze-drying technology, which produces a powder mixture of the active ingredient and any additional desired ingredient from its previously sterile-filtered solution.

[0428] When the peptide is suitably protected as described above, the active compound may be administered orally, for example, with an inert diluent or an assimilable edible carrier, or enclosed in a hard or soft shell gelatin capsule, or compressed into tablets, or incorporated directly into edible food. For oral therapeutic administration, the active compound may be incorporated with excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. Such compositions and preparations should contain at least 1% by weight of the active compound. The percentage of the compositions and preparations may, of course, be varied and may conveniently be between about 5 and about 80% by weight. The amount of active compound in such therapeutically useful compositions is such that a suitable dosage will be obtained. Preferred compositions or preparations according to the present invention are prepared so that an oral dosage unit contains between about 0.1 μg and 2000 mg of the active ingredient.

[0429] Tablets, pills, capsules, etc. may also contain the following: binders such as gum tragacanth, gum arabic, corn starch, or gelatin; excipients such as dicalcium phosphate; disintegrating agents such as corn starch, potato starch, or alginic acid; lubricants such as magnesium stearate; and sweeteners such as sucrose, lactose, or saccharin, or flavorings such as peppermint, wintergreen oil, or cherry flavoring. When the dosage unit is a capsule, it may contain a liquid carrier in addition to the above types of materials. Various other materials may be present as coatings or to otherwise modify the physical form of the dosage unit. For example, tablets, pills, or capsules may be coated with shellac, sucrose, or both. A syrup or elixir may contain the active compound, sucrose as a sweetener, methyl and propylparabens as preservatives, and a flavoring such as cherry or orange flavor. Of course, any material used in preparing any dosage unit form should be pharmaceutically pure and substantially non-toxic in the amounts employed. In addition, the active compounds may be incorporated into sustained-release preparations and formulations.

[0430] Delivery System Various delivery systems are known and can be used to administer the compounds of the present invention, including encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the compound, receptor-mediated endocytosis, constructs of nucleic acids as part of retroviruses or other vectors, etc. Methods of introduction include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, intraocular, epidural, and oral routes. The compounds or compositions can be administered by any convenient route, for example, by infusion or bolus injection, by absorption through epithelial or mucocutaneous surfaces (e.g., oral, rectal, and intestinal mucosa), and can also be administered in conjunction with other biologically active substances. Administration can be systemic or local. In addition, it may be desirable to introduce the pharmaceutical compounds or compositions of the present invention into the central nervous system by any suitable route, including intraventricular and intrathecal injection. Intraventricular injection can be facilitated, for example, by an intraventricular catheter attached to a reservoir, such as an Ommaya reservoir. Pulmonary administration can also be employed, eg, by use of an inhaler or nebulizer and formulation with an aerosolizing agent.

[0431] In certain embodiments, it may be desirable to administer a pharmaceutical compound or composition of the present invention locally to the area in need of treatment; this may be achieved, for example, but not limited to, by local infusion during surgery, topical application, e.g., in conjunction with a wound dressing after surgery, by injection, by catheter, by suppository, or by implant. Implants can be porous, non-porous, or gelatinous materials, including membranes such as silastic membranes, or fibers. Preferably, when administering proteins, including antibodies or peptides of the present invention, care must be taken to use materials to which the protein does not absorb. In another embodiment, the compound or composition can be delivered in vesicles, particularly liposomes. In yet another embodiment, the compound or composition can be delivered in a controlled-release system. In one embodiment, a pump can be used. In another embodiment, polymeric materials can be used. In yet another embodiment, the controlled-release system can be placed in the vicinity of the therapeutic target, thereby requiring only a fraction of the systemic dose.

[0432] Sequence Listing Free Text For the use of nucleotide abbreviations other than a, g, c, and t, they follow the rules set out in WIPO Standard ST.25, Supplementary Material 2, Table 1, where k represents t or g; n represents a, c, t, or g; m represents a or c; r represents a or g; s represents c or g; w represents a or t, and y represents c or t.

[0433] MTPGTQSPFF LLLLLTVLTV VTGSGHASST PGGEKETSAT QRSSVPSSTE KNAVSMTSSV LSSHSPGSGS STTQGQDVTL APATEPASGS AATWGQDVTS VPVTRPALGS TTPPAHDVTS APDNKPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDTRPAPGSTAPPAHGVTS APDTRPAPGS TAPPAHGVTS APDNRPALGS TAPPVHNVTS ASGSASGSAS TLVHNGTSAR ATTTPASKST PFSIPSHHSD TPTTLASHST KTDASSTHHS SVPPLTSSNH STSPQLSTGV SFFFLSFHIS NLQFNSSLED PSTDYYQELQ RDISEMFLQI YKQGGFLGLS NIKFRPGSVV VQLTLAFREG TINVHDVETQ FNQYKTEAAS RYNLTISDVS VSDVPFPFSA QSGAGVPGWG IALLVLVCVL VALAIVYLIA LAVCQCRRKN ​​YGQLDIFPAR DTYHPMSEYP TYHTHGRYVP PSSTDRSPYE KVSAGNGGSS LSYTNPAVAA ASANL (SEQ ID NO: 1) represents the full-length MUC1 receptor (mucin 1 precursor, GenBank accession number 15941).

[0434] MTPGTQSPFFLLLLLTVLT (SEQ ID NO: 2) MTPGTQSPFFLLLLLTVLT VVTA (SEQ ID NO: 3) MTPGTQSPFFLLLLLTVLT VVTG (SEQ ID NO: 4) SEQ ID NOS: 2, 3, and 4 are N-terminal MUC-1 signaling sequences for targeting the MUC1 receptor and truncated isoforms to the cell membrane surface. Up to three amino acid residues can be deleted at the C-terminus, as shown by the variants in SEQ ID NOS: 2, 3, and 4.

[0435] GTINVHDVETQFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGAGVPGWGIALLVLVCVLVALAIVYLIALAVCQCRRKNYGQLDIFPARDTYHPMSEYPTYHTHGRYVPPSSTDRSPYEKVSAGNGGSSLSYTNPAVAAASANL (SEQ ID NO: 5) represents a truncated MUC1 receptor isoform with the native PSMGFR at its N-terminus and contains the transmembrane and cytoplasmic sequences of the full-length MUC1 receptor.

[0436] GTINVHDVETQFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 6) represents the native primary sequence extracellular domain of the MUC1 receptor (nat-PSMGFR - an example of "PSMGFR").

[0437] TINVHDVETQFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 7) represents the native primary sequence extracellular domain of the MUC1 growth factor receptor (nat-PSMGFR - an example of "PSMGFR") with a single amino acid deletion at the N-terminus of SEQ ID NO: 6.

[0438] GTINVHDVETQFNQYKTEAASPYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 8) represents the extracellular domain of the "SPY" functional variant of the native primary sequence of the MUC1 growth factor receptor with increased stability (var-PSMGFR - an example of "PSMGFR").

[0439] TINVHDVETQFNQYKTEAASPYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 9) represents the extracellular domain of the "SPY" functional variant of the MUC1 growth factor receptor with increased stability (var-PSMGFR - an example of "PSMGFR") having a single amino acid deletion at the C-terminus of SEQ ID NO: 8.

[0440] tgtcagtgccgccgaaagaactacgggcagctggacatctttccagcccgggatacctaccatcctatgagcgagtaccccacctaccacacccatgggcgctatgtgccccctagcagtaccgatcgtagcccctatgagaaggtttctgcaggtaacggtggcagcagcctctcttacacaaacccagcagtggcagccgcttctgccaacttg (SEQ ID NO: 10) represents the MUC1 cytoplasmic domain nucleotide sequence.

[0441] CQCRRKNYGQLDIFPARDTYHPMSEYPTYHTHGRYVPPSSTDRSPYEKVSAGNGGSSLSYTNPAVAAASANL (SEQ ID NO: 11) represents the MUC1 cytoplasmic domain amino acid sequence.

[0442] gagatcctgagacaatgaatcatagtgaaagattcgttttcattgcagagtggtatgatccaaatgcttcacttcttcgacgttatgagcttttattttacccaggggatggatctgttgaaatgcatgatgtaaagaatcatcgcacctttttaaagcggaccaaatatgataacctgcacttggaagatttatttataggcaacaaagtgaatgtcttttctcgacaactggtattaattgactatggggatcaatatacagctcgccagctgggcagtaggaaagaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgttttttccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggtatgttgaatacactatattcagtacattttgttaataggagagcaatgtttattttcttgatgtactttatgtatagaaaataa (SEQ ID NO: 12) represents the NME7 nucleotide sequence (NME7: GenBank accession number AB209049).

[0443] DPETMNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGMLNTLYSVHFVNRRAMFIFLMYFMYRK (SEQ ID NO: 13) represents the NME7 amino acid sequence (NME7: GenBank Accession Number AB209049).

[0444] (SEQ ID NO: 14) represents the NM23-H1 nucleotide sequence (NM23-H1: GenBank accession number AF487339).

[0445] MVLLSTLGIVFQGEGPPISSCDTGTMANCERTFIAIKPDGVQRGLVGEIIKRFEQKGFRLVGLKFMQASEDLLKEHYVDLKDRPFFAGLVKYMHSGPVVAMVWEGLNVVKTGRVMLGETNPADSKPGTIRGDFCIQVGRNIIHGSDSVESAEKEIGLWFHPEELVDYTSCAQNWIYE (SEQ ID NO: 15) NM23-H1 represents the amino acid sequence (NM23-H1: GenBank accession number AF487339).

[0446] (SEQ ID NO: 16) represents the NM23-H1 S120G mutant nucleotide sequence (NM23-H1: GenBank accession number AF487339).

[0447] MVLLSTLGIVFQGEGPPISSCDTGTMANCERTFIAIKPDGVQRGLVGEIIKRFEQKGFRLVGLKFMQASEDLLKEHYVDLKDRPFFAGLVKYMHSGPVVAMVWEGLNVVKTGRVMLGETNPADSKPGTIRGDFCIQVGRNIIHGGDSVESAEKEIGLWFHPEELVDYTSCAQNWIYE (SEQ ID NO: 17) represents the NM23-H1 S120G mutant amino acid sequence (NM23-H1: GenBank Accession Number AF487339).

[0448] atggccaacctggagcgcaccttcatcgcatcaagccggacggcgtgcagcgcggcctggtgggcgagatcatcaagcgcttcgagcagaagggattccgcctcgtggccatgaagttcctccgggcc tctgaagaacacctgaagcagcactacattgacctgaaagaccgaccattcttccctgggctggtgaagtacatgaactcagggccggttgtggccatggtctgggaggggctgaacgtggtgaagaca ggccgagtgatgcttggggagaccaatccagcagattcaaagccaggcaccattcgtggggacttctgcattcaggttggcaggaacatcattcatggcagtgattcagtaaaaagtgctgaaaaagaaatcagcctatggtttaagcctgaagaactggttgactacaagtcttgtgctcatgactgggtctatgaataa (SEQ ID NO: 18) represents the NM23-H2 nucleotide sequence (NM23-H2: GenBank Accession Number AK313448).

[0449] MANLERTFIAIKPDGVQRGLVGEIIKRFEQKGFRLVAMKFLRASEEHLKQHYIDLKDRPFFPGLVKYMNSGPVVAMVWEGLNVVKTGRVMLGETNPADSKPGTIRGDFCIQVGRNIIHGSDSVKSAEKEISLWFKPEELVDYKSCAHDWVYE (SEQ ID NO: 19) represents the NM23-H2 amino acid sequence (NM23-H2: GenBank Accession Number AK313448).

[0450] Human NM23-H7-2 sequence optimized for E. coli expression: (DNA) (amino acid) MHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 21)

[0451] Human NME7-A: (DNA) atggaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctat tattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcat tagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgtttttttga (SEQ ID NO: 22) (amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFF (SEQ ID NO: 23)

[0452] Human NME7-A1: (DNA) atggaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgct ttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagag atgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaat gcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgtttttccttcaagtggaggttgtgggccggcaaacactgctaaatttacttga (SEQ ID NO: 24) (amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFT (SEQ ID NO: 25)

[0453] Human NME7-A2: (DNA) (SEQ ID NO: 26) (amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFF (SEQ ID NO: 27)

[0454] Human NME7-A3: (DNA) atgaatcatagtgaaagattcgttttcattgcagagtggtatgatccaaatgcttcacttcttcgacgttatgagcttttattttacccaggggatggatctgttgaaatgcatgatgtaaagaatcatcgcacctttttaaagcggaccaaatatgataacctgcacttggaagatttatttataggcaacaaagtgaatgtcttttctcgacaactggtattaattgactatggggatcaatatacagctcgccagctgggcagtaggaaagaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgttttttccttcaagtggaggttgtgggccggcaaacactgctaaatttacttga(SEQ ID NO: 28) (Amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFT (SEQ ID NO: 29)

[0455] Human NME7-B: (DNA) atgaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctatccgagatgcaggttttgaaatctcagctat gcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacagaaatgtattctggcc cttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaact ctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttctga (SEQ ID NO: 30) (amino acid) MNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 31)

[0456] Human NME7-B1: (DNA) atgaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctatccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacagaaatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaactctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttcaagatcttggataattagtga(SEQ ID NO: 32) (Amino acid) MNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN(SEQ ID NO: 33)

[0457] Human NME7 - B2: (DNA) atgccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatgg ctatccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacat ggtgacagaaatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcattta cgccctggaactctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttctga (SEQ ID NO: 34) (amino acid) MPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 35)

[0458] Human NME7-B3: (DNA) atgccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctat ccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacaga aatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaactct cagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttcaagatcttggataattagtga (SEQ ID NO: 36) (amino acid) MPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 37)

[0459] Human NME7, also known as NME7-AB AB : (DNA) atggaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgttttttccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctatccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacagaaatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaactctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttcaagatcttggataattagtga(SEQ ID NO: 38) (Amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN -- (SEQ ID NO: 39)

[0460] Human NME7-AB1: (DNA) atggaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgttttttccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctatccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacagaaatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaactctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttctga(SEQ ID NO: 40) (Amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 41)

[0461] Human NME7-A sequence optimized for E. coli expression: (DNA) atggaaaaaacgctggccctgattaaaccggatgcaatctccaaagctggcgaaattatcgaaattatcaacaaagcgggtttcaccatcacgaaactgaa aatgatgatgctgagccgtaaagaagccctggattttcatgtcgaccaccagtctcgcccgtttttcaatgaactgattcaattcatcaccacgggtccgat tatcgcaatggaaattctgcgtgatgacgctatctgcgaatggaaacgcctgctgggcccggcaaactcaggtgttgcgcgtaccgatgccagtgaatccat tcgcgctctgtttggcaccgatggtatccgtaatgcagcacatggtccggactcattcgcatcggcagctcgtgaaatggaactgtttttctga (SEQ ID NO: 42) (amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFF (SEQ ID NO: 43)

[0462] Human NME7-A1 sequence optimized for E. coli expression: (DNA) atggaaaaaacgctggccctgattaaaccggatgcaatctccaaagctggcgaaattatcgaaattatcaacaaagcgggtttcaccatcacgaaactgaaaatgatgatgct gagccgtaaagaagccctggattttcatgtcgaccaccagtctcgcccgtttttcaatgaactgattcaattcatcaccacgggtccgattatcgcaatggaaattctgcgtg atgacgctatctgcgaatggaaacgcctgctgggcccggcaaactcaggtgttgcgcgtaccgatgccagtgaatccattcgcgctctgtttggcaccgatggtatccgtaat gcagcacatggtccggactcattcgcatcggcagctcgtgaaatggaactgtttttcccgagctctggcggttgcggtccggcaaacaccgccaaatttacctga (SEQ ID NO: 44) (amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFT (SEQ ID NO: 45)

[0463] Human NME7-A2 sequence optimized for E. coli expression: (DNA) (SEQ ID NO: 46) (amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFF (SEQ ID NO: 47)

[0464] NME7-A3 sequence optimized for E. coli expression: (DNA) atgaatcactccgaacgctttgtttttatcgccgaatggtatgacccgaatgcttccctgctgcgccgctacgaactgctgttttatccgggcgatggtagcgtggaaatgcatgacgttaaaaatcaccgtacctttctgaaacgcacgaaatatgataatctgcatctggaagacctgtttattggcaacaaagtcaatgtgttctctcgtcagctggtgctgatcgattatggcgaccagtacaccgcgcgtcaactgggtagtcgcaaagaaaaaacgctggccctgattaaaccggatgcaatctccaaagctggcgaaattatcgaaattatcaacaaagcgggtttcaccatcacgaaactgaaaatgatgatgctgagccgtaaagaagccctggattttcatgtcgaccaccagtctcgcccgtttttcaatgaactgattcaattcatcaccacgggtccgattatcgcaatggaaattctgcgtgatgacgctatctgcgaatggaaacgcctgctgggcccggcaaactcaggtgttgcgcgtaccgatgccagtgaatccattcgcgctctgtttggcaccgatggtatccgtaatgcagcacatggtccggactcattcgcatcggcagctcgtgaaatggaactgtttttcccgagctctggcggttgcggtccggcaaacaccgccaaatttacctga(SEQ ID NO: 48) (Amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFT (SEQ ID NO: 49)

[0465] Human NME7-B sequence optimized for E. coli expression: (DNA) atgaattgtacgtgctgtattgtcaaaccgcacgcagtgtcagaaggcctgctgggtaaaattctgatggcaatccgtgatgctggctttgaaatctcggccat gcagatgttcaacatggaccgcgttaacgtcgaagaattctacgaagtttacaaaggcgtggttaccgaatatcacgatatggttacggaaatgtactccggtc cgtgcgtcgcgatggaaattcagcaaaacaatgccaccaaaacgtttcgtgaattctgtggtccggcagatccggaaatcgcacgtcatctgcgtccgggtacc ctgcgcgcaatttttggtaaaacgaaaatccagaacgctgtgcactgtaccgatctgccggaagacggtctgctggaagttcaatactttttctga (SEQ ID NO: 50) (amino acid) MNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 51)

[0466] Human NME7-B1 sequence optimized for E. coli expression: (DNA) atgaattgtacgtgctgtattgtcaaaccgcacgcagtgtcagaaggcctgctgggtaaaattctgatggcaatccgtgatgctggctttgaaatctcggccatgca gatgttcaacatggaccgcgttaacgtcgaagaattctacgaagtttacaaaggcgtggttaccgaatatcacgatatggttacggaaatgtactccggtccgtgcgt cgcgatggaaattcagcaaaacaatgccaccaaaacgtttcgtgaattctgtggtccggcagatccggaaatcgcacgtcatctgcgtccgggtaccctgcgcgcaat ttttggtaaaacgaaaatccagaacgctgtgcactgtaccgatctgccggaagacggtctgctggaagttcaatactttttcaaaattctggataattga (SEQ ID NO: 52) (amino acid) MNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 53)

[0467] Human NME7-B2 sequence optimized for E. coli expression: (DNA) atgccgagctctggcggttgcggtccggcaaacaccgccaaatttaccaattgtacgtgctgtattgtcaaaccgcacgcagtgtcagaaggcctgctgggtaaaattctgatgg caatccgtgatgctggctttgaaatctcggccatgcagatgttcaacatggaccgcgttaacgtcgaagaattctacgaagtttacaaaggcgtggttaccgaatatcacgatat ggttacggaaatgtactccggtccgtgcgtcgcgatggaaattcagcaaaacaatgccaccaaaacgtttcgtgaattctgtggtccggcagatccggaaatcgcacgtcatctg cgtccgggtaccctgcgcgcaattttggtaaaacgaaaatccagaacgctgtgcactgtaccgatctgccggaagacggtctgctggaagttcaatactttttctga (SEQ ID NO: 54) (amino acid) MPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 55)

[0468] Human NME7-B3 sequence optimized for E. coli expression: (DNA) atgccgagctctggcggttgcggtccggcaaacaccgccaaatttaccaattgtacgtgctgtattgtcaaaccgcacgcagtgtcagaaggcctgctgggtaaaattctgatggcaat ccgtgatgctggctttgaaatctcggccatgcagatgttcaacatggaccgcgttaacgtcgaagaattctacgaagtttacaaaggcgtggttaccgaatatcacgatatggttacgg aaatgtactccggtccgtgcgtcgcgatggaaattcagcaaaacaatgccaccaaaacgtttcgtgaattctgtggtccggcagatccggaaatcgcacgtcatctgcgtccgggtacc ctgcgcgcaatttttggtaaaacgaaaatccagaacgctgtgcactgtaccgatctgccggaagacggtctgctggaagttcaatactttttcaaaattctggataattga (SEQ ID NO: 56) (amino acid) MPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 57)

[0469] NME7 optimized for E. coli expression AB Human NME7-AB sequence, also known as: (DNA) atggaaaaaacgctggccctgattaaaccggatgcaatctccaaagctggcgaaattatcgaaattatcaacaaagcgggtttcaccatcacgaaactgaaaatgatgatgctgagccgtaaagaagccctggattttcatgtcgaccaccagtctcgcccgtttttcaatgaactgattcaattcatcaccacgggtccgattatcgcaatggaaattctgcgtgatgacgctatctgcgaatggaaacgcctgctgggcccggcaaactcaggtgttgcgcgtaccgatgccagtgaatccattcgcgctctgtttggcaccgatggtatccgtaatgcagcacatggtccggactcattcgcatcggcagctcgtgaaatggaactgtttttcccgagctctggcggttgcggtccggcaaacaccgccaaatttaccaattgtacgtgctgtattgtcaaaccgcacgcagtgtcagaaggcctgctgggtaaaattctgatggcaatccgtgatgctggctttgaaatctcggccatgcagatgttcaacatggaccgcgttaacgtcgaagaattctacgaagtttacaaaggcgtggttaccgaatatcacgatatggttacggaaatgtactccggtccgtgcgtcgcgatggaaattcagcaaaacaatgccaccaaaacgtttcgtgaattctgtggtccggcagatccggaaatcgcacgtcatctgcgtccgggtaccctgcgcgcaatttttggtaaaacgaaaatccagaacgctgtgcactgtaccgatctgccggaagacggtctgctggaagttcaatactttttcaaaattctggataattga(SEQ ID NO: 58) (Amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 59)

[0470] NME7 optimized for E. coli expression AB Human NME7-AB1 sequence, also known as 1: (DNA) Atggaaaaaacgctggccctgattaaaccggatgcaatctccaaagctggcgaaattatcgaaattatcaacaaagcgggtttcaccatcacgaaactgaaaatgatgatgctgagccgtaaagaagccctggattttcatgtcgaccaccagtctcgcccgtttttcaatgaactgattcaattcatcaccacgggtccgattatcgcaatggaaattctgcgtgatgacgctatctgcgaatggaaacgcctgctgggcccggcaaactcaggtgttgcgcgtaccgatgccagtgaatccattcgcgctctgtttggcaccgatggtatccgtaatgcagcacatggtccggactcattcgcatcggcagctcgtgaaatggaactgtttttcccgagctctggcggttgcggtccggcaaacaccgccaaatttaccaattgtacgtgctgtattgtcaaaccgcacgcagtgtcagaaggcctgctgggtaaaattctgatggcaatccgtgatgctggctttgaaatctcggccatgcagatgttcaacatggaccgcgttaacgtcgaagaattctacgaagtttacaaaggcgtggttaccgaatatcacgatatggttacggaaatgtactccggtccgtgcgtcgcgatggaaattcagcaaaacaatgccaccaaaacgtttcgtgaattctgtggtccggcagatccggaaatcgcacgtcatctgcgtccgggtaccctgcgcgcaatttttggtaaaacgaaaatccagaacgctgtgcactgtaccgatctgccggaagacggtctgctggaagttcaatactttttctga(SEQ ID NO: 60) (Amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 61)

[0471] Mouse NME6 (DNA) (SEQ ID NO: 62) (amino acid) MTSILRSPQALQLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRTRELQWKLEDCRRFYREHEGRFFYQRLVEFMTSGPIRAYILAHKDAIQLWRTLMGPTRVFRARYIAPDSIRGSLGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPVHYSPEEGIHCAAETGGHKQPNKT (SEQ ID NO: 63)

[0472] Human NME6: (DNA) (SEQ ID NO: 64) (amino acid) MTQNLGSEMASILRSPQALQLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPVCYSPEGGVHYVAGTGGLGPA (SEQ ID NO: 65)

[0473] Human NME6 1: (DNA) (SEQ ID NO: 66) (amino acid) MTQNLGSEMASILRSPQALQLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPV (SEQ ID NO: 67)

[0474] Human NME6 2: (DNA) Atgctcactctagccctgatcaagcctgacgcagtcgcccatccactgattctggaggctgttcatcagcagattctaagcaacaagttcctgattgtacgaatgagagaactactgtg gagaaaggaagattgccagaggttttaccgagagcatgaagggcgttttttctatcagaggctggtggagttcatggccagcgggccaatccgagcctacatccttgcccacaaggatg ccatccagctctggaggagctcatgggacccaccagagtgttccgagcacgccatgtggccccagattctatccgtgggagtttcggcctcactgacacccgcaacaccacccatggt tcggactctgtggtttcagccagcagagagattgcagccttcttccctgacttcagtgaacagcgctggtatgaggaggaagagccccagttgcgctgtggccctgtgtga (SEQ ID NO: 68) (amino acid) MLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPV (SEQ ID NO: 69)

[0475] Human NME6 3: (DNA) (SEQ ID NO: 70) (amino acid) MLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPVCYSPEGGVHYVAGTGGLGPA (SEQ ID NO: 71)

[0476] Human NME6 sequence optimized for E. coli expression: (DNA) (SEQ ID NO: 72) (amino acid) MTQNLGSEMASILRSPQALQLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPVCYSPEGGVHYVAGTGGLGPA (SEQ ID NO: 73)

[0477] Human NME6 1 sequence optimized for E. coli expression: (DNA) (SEQ ID NO: 74) (amino acid) MTQNLGSEMASILRSPQALQLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPV (SEQ ID NO: 75)

[0478] Human NME6 2 sequence optimized for E. coli expression: (DNA) Atgctgaccctggctctgatcaaaccggacgctgttgctcatccgctgattctggaagcggtccaccagcaaattctgagcaacaaatttctgatcgtgcgtatgcgcgaactgctgtg gcgtaaagaagattgccagcgtttttatcgcgaacatgaaggccgtttcttttatcaacgcctggttgaattcatggcctctggtccgattcgcgcatatatcctggctcacaaagatg cgattcagctgtggcgtaccctgatgggtccgacgcgcgtctttcgtgcacgtcatgtggcaccggactcaatccgtggctcgttcggtctgaccgatacgcgcaataccacgcacggt agcgactctgttgttagtgcgtcccgtgaaatcgcggcctttttcccggacttctccgaacagcgttggtacgaagaagaagaaccgcaactgcgctgtggcccggtctga (SEQ ID NO: 76) (amino acid) MLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPV (SEQ ID NO: 77)

[0479] Human NME6 3 sequence optimized for E. coli expression: (DNA) Atgctgaccctggctctgatcaaaccggacgctgttgctcatccgctgattctggaagcggtccaccagcaaattctgagcaacaaatttctgatcgtgcgtatgcgcgaactgctgtggcgtaaagaagattgccagcgtttttatcgcgaacatgaaggccgtttcttttatcaacgcctggttgaattcatggcctctggtccgattcgcgcatatatcctggctcacaaagatgcgattcagctgtggcgtaccctgatgggtccgacgcgcgtctttcgtgcacgtcatgtggcaccggactcaatccgtggctcgttcggtctgaccgatacgcgcaataccacgcacggtagcgactctgttgttagtgcgtcccgtgaaatcgcggcctttttcccggacttctccgaacagcgttggtacgaagaagaagaaccgcaactgcgctgtggcccggtctgttattctccggaaggtggtgtccattatgtggcgggcacgggtggtctgggtccggcatga(SEQ ID NO: 78) (Amino acid) MLTLALIKPDAVAHPLILEAVHQQILSNKFLIVRMRELLWRKEDCQRFYREHEGRFFYQRLVEFMASGPIRAYILAHKDAIQLWRTLMGPTRVFRARHVAPDSIRGSFGLTDTRNTTHGSDSVVSASREIAAFFPDFSEQRWYEEEEPQLRCGPVCYSPEGGVHYVAGTGGLGPA(SEQ ID NO: 79)

[0480] OriGene - NME7 - 1 full length (DNA) (amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDNTRTRRLEQKLISEEDLAANDILDYKDDDDKV (SEQ ID NO: 81)

[0481] Abnova NME7-1 full length (amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 82)

[0482] Abnova partial NME7-B (amino acid) DRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKIL (SEQ ID NO: 83)

[0483] Histidine tag (ctcgag)caccaccaccaccaccactga (SEQ ID NO: 84)

[0484] Strep II tag (accggt)tggagccatcctcagttcgaaaagtaatga (SEQ ID NO: 85)

[0485] N-10 peptide: QFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGA (SEQ ID NO: 86)

[0486] C-10 peptides GTINVHDVETQFNQYKTEAASRYNLTISDVSVSDV (SEQ ID NO: 87) LALIKPDA (SEQ ID NO: 88) MMMLSRKEALDFHVDHQS (SEQ ID NO: 89) ALDFHVDHQS (SEQ ID NO: 90) EILRDDAICEWKRL (SEQ ID NO: 91) FNELIQFITTGP (SEQ ID NO: 92) RDDAICEW (SEQ ID NO: 93) SGVARTDASESIRALFGTDGIRNAA (SEQ ID NO: 94) ELFFPSSGG (SEQ ID NO: 95) KFTNCTCCIVKPHAVSEGLLGKILMA (SEQ ID NO: 96) LMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVT (SEQ ID NO: 97) EFYEVYKGVVTEYHD (SEQ ID NO: 98) EIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNA (SEQ ID NO: 99) YSGPCVAM (SEQ ID NO: 100) FREFCGP (SEQ ID NO: 101) VHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 102) IQNAVHCTD (SEQ ID NO: 103) TDLPEDGLLEVQYFFKILDN (SEQ ID NO: 104) PEDGLLEVQYFFK (SEQ ID NO: 105) EIINKAGFTITK (SEQ ID NO: 106) MLSRKEALDFHVDHQS (SEQ ID NO: 107) NELIQFITT (SEQ ID NO: 108) EILRDDAICEWKRL (SEQ ID NO: 109) SGVARTDASESIRALFGTDGI (SEQ ID NO: 110) SGVARTDASES (SEQ ID NO: 111) ALFGTDGI (SEQ ID NO: 112) NCTCCIVKPHAVSE (SEQ ID NO: 113) LGKILMAIRDA (SEQ ID NO: 114) EISAMQMFNMDRVNVE (SEQ ID NO: 115) EVYKGVVT (sequence number 116) EYHDMVTE (SEQ ID NO: 117) EFCGPADPEIARHLR (sequence number 118) AIFGKTKIQNAV (SEQ ID NO: 119) LPEDGLLEVQYFFKILDN (sequence number 120) GPDSFASAAREMELFFP (SEQ ID NO: 121)

[0487] Human NME7-derived immunizing peptide ICEWKRL (SEQ ID NO: 122) LGKILMAIRDA (SEQ ID NO: 123) HAVSEGLLGK (sequence number 124) VTEMYSGP (SEQ ID NO: 125) NATKTFREF (SEQ ID NO: 126) AIRDAGFEI (SEQ ID NO: 127) AICEWKRLLGPAN (SEQ ID NO: 128) DHQSRPFF (SEQ ID NO: 129) AICEWKRLLGPAN (SEQ ID NO: 130) VDHQSRPF (SEQ ID NO: 131) PDSFAS (SEQ ID NO: 132) KAGEIIEIINKAGFTITK (SEQ ID NO: 133)

[0488] Human NME1-derived immunizing peptide MANCERTFIAIKPDGVQRGLVGEIIKRFE (SEQ ID NO: 134) VDLKDRPF (sequence number 135) HGSDSVESAEKEIGLWF (SEQ ID NO: 136) ERTFIAIKPDGVQRGLVGEIIKRFE (SEQ ID NO: 137) VDLKDRPFFAGLVKYMHSGPVVAMVWEGLN (SEQ ID NO: 138) NIIHGSDSVESAEKEIGLWFHPEELV (SEQ ID NO: 139) KPDGVQRGLVGEII (SEQ ID NO: 140)

[0489] Immunizing peptide derived from human NME7 that does not bind to NME1 MLSRKEALDFHVDHQS (SEQ ID NO: 141) Peptide A1 SGVARTDASES (SEQ ID NO: 142) Peptide A2 DAGFEISAMQMFNMDRVNVE (SEQ ID NO: 143) Peptide B1 EVYKGVVTEYHDMVTE (SEQ ID NO: 144) Peptide B2 AIFGKTKIQNAVHCTDLPEDGLLEVQYFF (SEQ ID NO: 145) Peptide B3

[0490] Human NME7a (DNA) (amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 147)

[0491] Human NME7b (DNA) (amino acid) MHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 149)

[0492] NME7 AB Also known as human NME7-AB (DNA) atggaaaaaacgctagccctaattaaaccagatgcaatatcaaaggctggagaaataattgaaataataaacaaagctggatttactataaccaaactcaaaatgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgttttttccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctatccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacagaaatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaactctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttcaagatcttggataattag(SEQ ID NO: 150) (Amino acid) MEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN (SEQ ID NO: 151)

[0493] Human NME7-X1 (DNA) atgatgatgctttcaaggaaagaagcattggattttcatgtagatcaccagtcaagaccctttttcaatgagctgatccagtttattacaactggtcctattattgccatggagattttaagagatgatgctatatgtgaatggaaaagactgctgggacctgcaaactctggagtggcacgcacagatgcttctgaaagcattagagccctctttggaacagatggcataagaaatgcagcgcatggccctgattcttttgcttctgcggccagagaaatggagttgttttttccttcaagtggaggttgtgggccggcaaacactgctaaatttactaattgtacctgttgcattgttaaaccccatgctgtcagtgaaggactgttgggaaagatcctgatggctatccgagatgcaggttttgaaatctcagctatgcagatgttcaatatggatcgggttaatgttgaggaattctatgaagtttataaaggagtagtgaccgaatatcatgacatggtgacagaaatgtattctggcccttgtgtagcaatggagattcaacagaataatgctacaaagacatttcgagaattttgtggacctgctgatcctgaaattgcccggcatttacgccctggaactctcagagcaatctttggtaaaactaagatccagaatgctgttcactgtactgatctgccagaggatggcctattagaggttcaatacttcttcaagatcttggataattag(SEQ ID NO: 152) (Amino acid) MMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVS EGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPADPEIARHLRPGTLRAIFGKTKIQNAVHCTDLPEDGLLEVQYFFKILDN * (SEQ ID NO: 153)

[0494] Human NME7α (optimized for E. coli expression) (DNA) (amino acid) MNHSERFVFIAEWYDPNASLLRRYELLFYPGDGSVEMHDVKNHRTFLKRTKYDNLHLEDLFIGNKVNVFSRQLVLIDYGDQYTARQLGSRKEKTLALIKPDAISKAGEIIEIINKAGFTITKLKMMMLSRKEALDFHVDHQSRPFFNELIQFITTGPIIAMEILRDDAICEWKRLLGPANSGVARTDASESIRALFGTDGIRNAAHGPDSFASAAREMELFFPSSGGCGPANTAKFTNCTCCIVKPHAVSEGLLGKILMAIRDAGFEISAMQMFNMDRVNVEEFYEVYKGVVTEYHDMVTEMYSGPCVAMEIQQNNATKTFREFCGPA...

Claims

1. An NME7 specific antibody or fragment thereof which binds to the NME7 B3 peptide of SEQ ID NO:145 or SEQ ID NO:

169.

2. The antibody of claim 1 which is a monoclonal antibody.

3. The antibody of claim 1 which is a bivalent, monovalent, Fab, or single chain variable fragment antibody (scFv).

4. The antibody of claim 1 , which is linked to an antibody-drug conjugate.

5. The antibody of claim 1 , wherein the drug is linked to a toxin or protoxin.

6. An isolated nucleic acid encoding the monoclonal antibody of claim 2.

7. An isolated hybridoma expressing the monoclonal antibody of claim 2.

8. NME7 ＡＢ Or the antibody of claim 1, which specifically binds to NME7-X1 but does not specifically bind to NME1.

9. NME7 ＡＢ and MUC1 ＊ Between the extracellular domains or between NME7-X1 and MUC1 ＊ The antibody of claim 1 , which disrupts the interaction between the extracellular domains.

10. NME7 ＡＢ The antibody of claim 1, which disrupts the binding between NME7-X1 and PSMGFR or between NME7-X1 and PSMGFR.

11. NME7 ＡＢ The antibody of claim 1, which disrupts the binding between NME7-X1 and N-10 or between NME7-X1 and N-10.

12. NME7 ＡＢ and MUC1 ＊ Between the extracellular domains or between NME7-X1 and MUC1 ＊ The NME7 does not disrupt the interaction between the extracellular domains. ＡＢ or the antibody of claim 1, wherein NME7-X1 binds to the N-10 peptide (SEQ ID NO: 170) but does not bind to the C-10 peptide (SEQ ID NO: 171).

13. The antibody of claim 2 , wherein the antibody is 5A1, 4A3 or 5D4.

14. An amino acid sequence in the heavy chain variable region comprising: YTFTNYGMN (SEQ ID NO:439) in the CDR1 region; WINTYTGEPTYVDDFKG (SEQ ID NO: 440) in the CDR2 region; and LRGIRPGPLAY (SEQ ID NO:441) in the CDR3 region; and An amino acid sequence in the light chain variable region comprising: SASSSVSYMN (SEQ ID NO:444) in the CDR1 region; GISNLAS (SEQ ID NO:445) in the CDR2 region; and QQRSSYPPT (SEQ ID NO: 446) in the CDR3 region The antibody of claim 2 .

15. An amino acid sequence in the heavy chain variable region comprising: NTFTEYTMH (SEQ ID NO:429) in the CDR1 region; GFNPNNGVTNYNQKFKG (SEQ ID NO:430) in the CDR2 region; and RYYHSTYVFYFDS (SEQ ID NO:431) in the CDR3 region; and An amino acid sequence in the light chain variable region comprising: SASQGISNYLN (SEQ ID NO:434) in the CDR1 region; YTSSLHS (SEQ ID NO:435) in the CDR2 region; and QQYSKLPYT (SEQ ID NO: 436) in the CDR3 region The antibody of claim 2 .

16. An amino acid sequence in the heavy chain variable region comprising: NTFTEYTMH (SEQ ID NO:388) in the CDR1 region; GFNPNNGVTNYNQKFKG (SEQ ID NO:389) in the CDR2 region; and RYYHSLYVFYFDY (SEQ ID NO: 390) in the CDR3 region; and An amino acid sequence in the light chain variable region comprising: ITSTDIDDDMN (SEQ ID NO:393) in the CDR1 region; EGNTLRP (SEQ ID NO:394) in the CDR2 region; and LQSDNLPLT (SEQ ID NO: 395) in the CDR3 region The antibody of claim 2 .

17. The antibody of claim 1 which is a human antibody, a humanized antibody or an engineered antibody mimetic.

18. The antibody of claim 1 which is non-human.

19. 19. The antibody of claim 18 which is murine or camelid.

20. A method for administering to a patient for the prevention or treatment of cancer, comprising administering to said patient a composition comprising an antibody according to claims 1 to 16.

21. A method for preventing or treating cancer metastasis in a patient, comprising administering to said patient a composition comprising an antibody according to claims 1 to 16.

22. A method for diagnosing cancer or cancer metastasis, comprising contacting a patient sample and a normal sample with an antibody according to claims 1-16 and comparing the results obtained from both samples, wherein the presence of positive binding to said antibody in said patient sample indicates the presence of cancer or cancer metastasis in said patient.

23. The method of claim 22, wherein the antibody is as defined in claim 12.

24. The method of claim 22, wherein the antibody is linked to an imaging agent.

25. 23. The method of claim 22, wherein the patient specimen is blood, body fluids, tissue, circulating cells, in vitro, including intraoperatively, in vivo.

26. anti-NME7 ＡＢ Cells engineered to express an antibody or fragment thereof.

27. The cell of claim 26 , wherein the cell is an immune cell.

28. The cell of claim 26, wherein the immune cell is a T cell or a NK cell.

29. 27. The cell of claim 26, wherein the cell is a stem or progenitor cell.

30. 30. The cell of claim 29, wherein the stem or progenitor cell subsequently differentiates to become a T cell.

31. The cell of claim 26, comprising a chimeric antigen receptor (CAR) that recognizes a tumor-associated antigen.

32. The cell of claim 26 , wherein expression of the anti-NME7 antibody is inducible.

33. anti-NME7 ＡＢ 27. The cell of claim 26, wherein the nucleic acid encoding the antibody is inserted into a Foxp3 enhancer or promoter.

34. anti-NME7 ＡＢ The cell of claim 26 , wherein the antibody is present in an NFAT induction system.

35. The cell of claim 34, wherein the NFATc1 response element is inserted upstream of the antibody sequence that is inserted into the Foxp3 enhancer or promoter region.

36. The anti-NME7 ＡＢ The antibody or fragment thereof binds to the NME7 B3 peptide or to the MUC1 ＊ NME7 to the PSMGFR peptide of the extracellular domain ＡＢ Or the cell according to claim 26, which disrupts the binding of NME7-X1.

37. The immune cell of claim 27, comprising expressing a CAR that recognizes a tumor-associated antigen and an anti-NME7 antibody.

38. The tumor-associated antigen is MUC1 ＊ The immune cell of claim 37 .

39. NME7, listed in Figures 6-9 as an immunogenic elicitor moiety ＡＢ 2. An anti-cancer vaccine comprising a composition comprising one or more peptides derived from or having at least 80%, 85%, 90%, 95%, 97% sequence identity thereto.

40. The anti-cancer vaccine of claim 39, wherein the peptide is a peptide of SEQ ID NO: 141-145 or a peptide having at least 80%, 85%, 90%, 95%, 97% sequence identity thereto.

41. 41. The anti-cancer vaccine of claim 40, wherein the peptide is a peptide of SEQ ID NO: 145 or a peptide having at least 80%, 85%, 90%, 95%, 97% sequence identity thereto.

42. A BiTE comprising an antibody according to any one of claims 1 to 16.

43. The cysteine ​​residues in the NME7 B3 peptide are mutated to avoid disulfide bonds. ＡＢ Methods for generating antibodies.

44. The cells were cultured at 100°C for 10 min. ＡＢ Or a method of generating cells with enhanced metastatic potential comprising culturing with NME7-X1.

45. NME7 ＡＢ Or cells engineered to express NME7-X1.

46. NME7 ＡＢ or a transgenic animal expressing NME7-X1.

47. NME7 ＡＢ Or the transgenic animal of claim 46, wherein NME7-X1 is human.

48. NME7 ＡＢ or the transgenic animal of claim 47, wherein expression of NME7-X1 is inducible.

Images