Fully human monoclonal antibody of anti-human granulin precursor

Abstract

Described herein are antibodies, in particular monoclonal antibodies, that specifically bind to human granulin precursors and are useful in the treatment of cancer in a patient. Methods of making and using the same are also provided.

Description

[0001] Related Applications

[0002] This application claims priority to U.S. Provisional Application Serial No. 63 / 313,304, filed on February 24, 2022, the entire contents of which are incorporated herein. Technical Field

[0003] Provided herein are fully human monoclonal antibodies (mAbs). More specifically, described herein are mAbs that specifically bind to human progranulin (also known as GP88). Other names for progranulin include progranulin / epithelialin precursor, GEP, and acrogranin. Background Art

[0004] A promising new target for cancer therapy and diagnosis has been identified and is called PC-cell derived growth factor (PCDGF) (also known as granulin-epithelialin precursor [GEP or GP88] or progranulin). PCDGF (referred to herein as progranulin) is an 88 kDa glycoprotein containing a 20 kDa carbohydrate moiety and a 68 kDa core protein that is identical to the precursor of epithelin / granulin, a family of six dicysteine-rich polypeptides derived from proteolytic processing of progranulin (Bhandari, V. et al., Proc Natl Acad Sci USA 89(5): 1715-1719, 1992). The first identified members of this family were 6 kDa polypeptide growth regulators, epithelins 1 and 2 isolated from rat kidney (Shoyab, M. et al., Proc Natl Acad of Sci 87(20):7912, 1990) or granulocytes from human granulocyte extracts (Bateman et al., Biochem Biophys Res Commun 173(3):1161-1168, 1990).

[0005] Cloning of the complementary deoxyribonucleic acid (cDNA) of epithelin and granulin revealed that it encodes a 63 kDa protein containing a 7.5 kDa cysteine-rich epithelin / granulin repeat sequence, a signal peptide, and several putative glycosylation sites (Bhandari, V. et al., Proc Natl Acad Sci USA 89(5):1715-1719, 1992). Studies on the biological function of the cloned and expressed precursor showed that it was biologically inactive (Bhandari, V. et al., 1992 and Plowman, G. et al., J Biol Chem 267(18):13073-13078, 1992). This led to the hypothesis that the precursor needs to be processed into a 6 kDa molecular form in order to obtain biological activity. Based on this hypothesis, the large molecular weight form was defined as granulin / epithelin precursor or PGRN, now known as progranulin. Under normal physiological conditions, progranulin is not secreted but serves as a precursor of mature granulin / epithelin. Work from the Serrero laboratory reported the purification of an autocrine growth factor from the culture medium of a highly tumorigenic teratomatous PC cell line, providing the first demonstration of the physiological existence of the precursor as a biologically active growth factor as a glycoprotein of 88 kDa with a 17 amino acid signal peptide and a 20 kDa carbohydrate moiety (Zhou, G. et al., J Biol Chem 268(15):10863-10869, 1993). In other studies on several cell lines of epithelial or mesenchymal origin (Zanocco-Marani, T. et al., Cancer Res 59(20):5331-5340, 1999; He, Z. et al., Cancer Research 62(19):5590, 2002; He, Z. et al., J Mol Med 81(10):600-612, 2003; Serrero, G, Biochem Biophys Res Commun 308(3):409-413, 2003), it was found that the autocrine production and secretion of progranulin leads to growth dysregulation and causes tumor formation in vivo, indicating that progranulin acts as a tumorigenic growth factor.

[0006] High expression levels of autosecretory progranulin are associated with the most malignant breast cancer cells. For example, progranulin messenger RNA (mRNA) and protein expression are very low in immortalized, non-tumorigenic breast epithelial MCF-10A cells and estrogen receptor positive (ER+) MCF-7 cells. In contrast, significantly increased levels of progranulin were observed in more aggressive estrogen receptor negative (ER-) cell lines (e.g., MDA-MB-468, 453, and 231) (Lu, R. et al., Proc Natl Acad Sci USA 97 (8): 3993-3998, 2000). Further studies have confirmed that progranulin mediates the growth-promoting effects of 17-β estradiol (E2) in human breast cancer cells; wherein, in E2-dependent MCF-7 cells, overexpression of progranulin renders the cells independent of E2, while E2 receptor status and E2 responsiveness remain unchanged, causing the cells to become tamoxifen-resistant (Lu, R. et al., Proc Natl Acad Sci USA 98(1):142-147, 2001). In addition, it has been found that mice bearing progranulin-overexpressing tumors treated with tamoxifen develop larger tumors than mice not treated with tamoxifen, indicating that ER+ breast cancer cells overexpressing progranulin are not only resistant to the growth inhibitory effects of tamoxifen, but also respond adversely to tamoxifen, resulting in stimulation of tumor growth in vivo. In addition to conferring tamoxifen resistance, data also indicate that overexpression of progranulin in ER+ breast cancer cells leads to resistance to fulvestrant (FASLODEX), the aromatase inhibitor letrozole, and the chemotherapeutic agent doxorubicin (Tangkeangsirisin, W. et al., Carcinogenesis 25(9): 1587-1592, 2004; Abrhale; T. et al., BMC Cancer 11: 231, 2011; Tangkeangsirisin, W. et al., Advances in Breast Cancer Research Vol. 3 No. 3: 11, 2014). The latter finding is supported by a study by Kudoh et al. (Kudoh, R. et al., Cancer Research 60(15): 4161, 2000), in which microarray analysis identified progranulin as a gene significantly (>12-fold) upregulated in doxorubicin-resistant breast cancer cells MCF-7 / D40. Furthermore, progranulin was upregulated 17-fold in MCF-7 cells treated with doxorubicin for 15 h. These results suggest that increased progranulin expression in breast tumors is associated with chemoresistance.

[0007] Although little is known about the progranulin receptor because it has not been completely cloned, some laboratories have studied the progranulin-dependent signaling pathway that mediates proliferation and survival (Arechavaleta-Velasco, F. et al., MedOncol 34(12): 194, 2017). Regarding the receptor, it has been demonstrated that the specific, time- and temperature-dependent binding of progranulin to cells is saturable. Biotinylated progranulin was cross-linked to human breast cancer cells using the cross-linking agent DSS, showing the presence of a cross-linked band with an apparent molecular weight of approximately 190-200 kDa, indicating that the apparent molecular weight of the putative receptor is 100-110 kDa (Xia et al., Biochem Biophys Res Commun 245(2): 539-543, 1998). Cells that do not bind or respond to progranulin do not present this cross-linked protein complex. Progranulin-mediated intracellular signaling pathways include MAPK ERK 1 / 2, PI-3 kinase and FAK, which lead to the activation of cell cycle regulatory proteins Cyclin D1 and Cyclin B (Zanocco-Marani, T. et al., Cancer Res 59(20):5331-5340, 1999; Lu, R. et al., Proc Natl Acad Sci USA 98(1):142-147, 2001; He, Z. et al., J Mol Med 81(10):600-612, 2003; Jones, MB et al., Clin Cancer Res 9(1):44-51, 2003). It has been shown that in breast cancers overexpressing Her-2, progranulin and its receptor can interact with the Her-2 receptor rather than the EGF receptor, leading to Her-2 phosphorylation and subsequent Herceptin resistance (Kim, WE et al., Clin Cancer Res 12(14Pt1):4192-4199, 2006).

[0008] Regarding the survival function of progranulin, investigations into the mechanism of action have shown that progranulin blocks the apoptotic effect of tamoxifen by upregulating bcl-2 expression, and also strongly stimulates the expression of the angiogenic factors vascular endothelial growth factor (VEGF) and angiopoietin-2 (Tangkeangsirisin, W. et al., Carcinogenesis 25(9): 1587-1592, 2004). Since progranulin stimulates VEGF, investigations were conducted to determine whether progranulin also stimulates other processes involved in metastasis. Whether overexpressed or exogenously added in MCF-7 cells, progranulin stimulates migration, expression of matrix metalloproteinase-9, and invasion (Tangkeangsirisin, W. et al., Tangkeangsirisin, W. et al., Cancer Res 64(5): 1737-1743, 2004; Tangkeangsirisin, W. et al., Carcinogenesis 25(9): 1587-1592, 2004). In addition to MMP-9, activation of MMP-2 has also been reported (He, Z. et al., J Mol Med 81(10):600-612, 2003). Thus, progranulin stimulates angiogenesis and tumor cell invasiveness mediated at least in part by stimulating VEGF and matrix metalloproteinase expression, respectively (He, Z. et al., Cancer Research 62(19):5590, 2002; He et al., 2003; Tangkeangsirisin, W. et al., Cancer Res 64(5):1737-1743, 2004; Tangkeangsirisin, W. et al., Carcinogenesis 25(9):1587-1592, 2004). These data suggest that elevated progranulin levels are associated with increased metastasis.

[0009] Recently, a human / mouse chimeric monoclonal antibody named AG01 that specifically binds to human progranulin was demonstrated to have in vivo efficacy. Mice bearing MDA-MB-231 subcutaneous tumors treated with AG01 showed a significant reduction in tumor growth rate and tumor weight, while body and organ weights were not affected (Guha, R. et al., Breast Cancer Res Treat 186(3): 637-653, 2021).

[0010] Therefore, there is a need for mAbs against human progranulin, in particular fully human mAbs that specifically bind to human progranulin and can be used to treat human cancers in patients. Summary of the invention

[0011] Provided herein are monoclonal antibodies or antigen-binding fragments that bind to progranulin (PGRN or GP88). The isolated antibodies or antigen-binding fragments described herein have a K of less than or equal to 100 pM. D Binds to human progranulin (GP88). For example, the isolated antibodies or antigen-binding fragments described herein can bind to human progranulin with a K of less than or equal to 10 pM, less than or equal to 1 pM, less than or equal to 0.1 pM, or less than or equal to 0.01 pM. D More specifically, the isolated antibodies or antigen-binding fragments described herein can also bind to human progranulin with a K of less than or equal to 0.05 pM or less than or equal to 0.4 pM. D Binding, as measured by the Octet assay described below or as measured by any assay available to one of skill in the art. Certain mAbs herein do not compete with mAb AG0 1, a chimeric monoclonal antibody containing (a) a mouse variable region that specifically binds to human progranulin and (b) a human constant region (Guha, R. et al., 2021). Described herein is an isolated antibody or antigen-binding fragment thereof that binds to human progranulin and further competes for binding with mAb 16C11, 10B3, 10C8 or 14A6 described herein. Described herein is an isolated antibody or antigen-binding fragment thereof that binds to the same epitope as mAb 16C 11, 10B3, 10C8 or 14A6. Specifically, the mAbs herein include anti-human progranulin antibodies having the amino acid sequences described in Table 1 (shown using single-letter abbreviations standard in the art). These variable regions can be linked to IgG1 / κ isotype constant regions. The CDR residue numbering system of Chothia and Kabat is well known in the art and is further described in Dondelinger, M. et al., Front Immunol 9:2278, 2018, and was used to determine the amino acid sequences shown in Table 1 below. H and V L) are also shown below. The monoclonal antibody may further comprise the complete heavy chain and / or light chain of the antibody described in Table 1. In some embodiments, the monoclonal antibody may comprise the antigen binding domain of such heavy chain and / or light chain, such as the CDRs shown in Table 1. In some embodiments, the present disclosure provides nucleotide sequences encoding 10C8, 16C 11, 14A6 or 10B3 antibodies (or functional fragments thereof, such as CDRs and / or variable regions thereof), and / or specific amino acid sequences equivalent to the following, wherein equivalents can be readily obtained from the amino acid sequence of any one of SEQ ID NOs: 1-56 and the information shown in Table 4 and using the functional assays and other assays disclosed herein, and / or otherwise available to those of ordinary skill in the art. In some embodiments, the present disclosure also provides expression vectors, which include isolated nucleic acids comprising and / or consisting of such nucleotide sequences (in preferred embodiments, any one of SEQ ID NOs: 57-64 or derivatives thereof) and host cells (e.g., cell lines) containing such expression vectors. The content described herein relates to isolated nucleic acids encoding the heavy chain or light chain of the monoclonal antibody herein, expression vectors comprising the isolated nucleic acids, and cell lines containing the expression vectors. Also described herein is a method for treating cancer in a patient by identifying a patient with cancer cells expressing progranulin and administering an antibody or antigen-binding fragment as described herein to the patient. It is known that a variety of human cancers express progranulin, including ovarian cancer, breast cancer, multiple myeloma, lung cancer, kidney cancer, prostate cancer, hepatocellular carcinoma, uterine cancer, bladder cancer, bile duct cancer, esophageal cancer, gastric cancer, laryngeal cancer, brain cancer, leukemia, and glioblastoma. Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those generally understood by those of ordinary skill in the art. It should be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not limiting. Additional features will be described in the following specification section, or may be learned through the practice described herein. The aforementioned features and other features will become apparent to those skilled in the art after considering the description of the following exemplary embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] The accompanying drawings and photographs are incorporated in and constitute a part of this specification, which illustrate several embodiments and together with the description serve to explain the principles of the present invention. It is noteworthy that AG01 is the same mAb as c4F10 described herein and included in the figures.

[0013] Figure 1 is a bar graph showing flow binding of various anti-progranulin (PRGN or GP88) mAbs to MDA-MB-231 cells at various mAb concentrations.

[0014] Figure 2 is a bar graph showing the percentage of MDA-MB-231 cell migration for different anti-progranulin mAbs.

[0015] Figure 3 is a bar graph showing tumor volume after treatment with the indicated antibodies in an in vivo tumor growth model. The model uses MDA-MB-231 cells injected into athymic mice. The term "ip" refers to intraperitoneal injection of the therapeutic mAb herein.

[0016] Figure 4 is a line graph showing tumor volume in mice treated with the novel fully human mAb 10C8. DETAILED DESCRIPTION OF THE INVENTION

[0018] The present disclosure relates to antibodies, including but not limited to fully human monoclonal antibodies, which specifically bind to human progranulin (hu PRGN, or PRGN, or GP88) and can be used to treat cancers, such as but not limited to breast cancer (e.g., triple negative breast cancer (which can be exemplified using, for example, the MDA-MB-231 cell line), adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer (which can be exemplified using, for example, the MDA-MB-468 cell line), basal breast cancer (which can be exemplified using, for example, the HS578T cell line), estrogen receptor (ER) luminal breast cancer (which can be exemplified using, for example, the MCF-7 cell line), ER positive tamoxifen resistant luminal breast cancer (which can be exemplified using, for example, the TamR MCF-7 cell line for illustration), letrozole-resistant luminal breast cancer (e.g., ACLRTUSM cell line for illustration), ovarian cancer, uterine cancer, prostate cancer, bladder cancer (e.g., T24 cell line for illustration), renal cancer, hepatocellular carcinoma, bile duct cancer, esophageal cancer, colorectal cancer, gastric cancer, laryngeal cancer, lung cancer (e.g., non-small cell lung cancer (e.g., H1299 cell line for illustration), lung adenocarcinoma (e.g., A549 cell line for illustration), brain cancer, myeloma / leukemia, mesothelioma (e.g., MSTO-H11 cell line for illustration), epidermoid or squamous cell carcinoma (e.g., A431 cell line for illustration), and cancers expressing progranulin (GP88) recognized or discovered by those skilled in the art, etc. In preferred embodiments, exemplary cell lines (e.g., cell lines that can exemplify cancer) are cell lines that are known and can be shown to express and / or bind progranulin (GP88) in standard assays, detected by antibodies of the present disclosure, and / or derivatives bind to or inhibit binding to progranulin (GP88) by the antibodies and / or derivatives disclosed herein, and / or react to the antibodies and / or derivatives in vitro (e.g., using migration assays) and / or in vivo (e.g., using xenografts). In some embodiments, the disclosure provides isolated antibodies, one or more antigen-binding fragments thereof, which bind to human progranulin (GP88) and further compete for binding with antibodies 10C8, 16C11, 14A6 or 10B3 antibodies (or functional fragments thereof, such as CDRs and / or variable regions thereof); heavy and light chain variable regions thereof (see Table 1); and / or polypeptides comprising the CDRs of such antibodies (see Table 1); and / or derivatives thereof (e.g., comprising conservative amino acid substitutions thereof (see, e.g., Table 4)), any of which are considered equivalents thereof. Specifically, the mAbs herein include anti-human progranulin (GP88) antibodies having the amino acid sequences of the respective CDRs described in Table 1, or variants thereof, as disclosed herein or as would be otherwise recognized by a person of ordinary skill in the art.

[0019] In preferred embodiments, antibodies of the present disclosure comprise the following amino acid and nucleotide sequences. Preferred antibodies of the present disclosure may comprise the complementary determining regions (CDRs) of 10C8, 16C11, 14A6 or 10B3 antibodies. Table 1 shows the CDRs of 10C8, 16C11, 14A6 or 10B3 antibodies using the Chothia and Kabat methods.

[0020] Table 1: Monoclonal Antibody CDRs

[0021]

[0022]

[0023] In some embodiments, the present disclosure provides fully human anti-progranulin antibodies that are internalized into cells expressing progranulin and compete with trastuzumab for binding to HER2, as well as methods of using such antibodies for their neutralization and internalization properties. In preferred embodiments, such antibodies include antibodies referred to herein as 10C8, 16C11, 14A6 or 10B3, and / or CDRs (see Table 1) contained in such antibodies; their heavy and light chain variable regions (see Table 1); and / or derivatives thereof (e.g., containing conservative amino acid substitutions (see, e.g., Table 4)). In some embodiments, the present disclosure provides fully human anti-progranulin antibodies that are internalized and do not compete with antibodies or derivatives disclosed herein for binding to progranulin, as well as methods of studying their internalization properties. In preferred embodiments, such antibodies include antibodies referred to herein as 10C8, 16C 11, 14A6 or 10B3; comprising heavy and light chain variable regions thereof (see Table 1); and / or comprising CDRs of Table 1; and / or derivatives thereof (e.g., comprising conservative amino acid substitutions thereof (see, e.g., Table 4)). The antibody may further include at least a portion of an antibody shown in Table 1 (most preferably including CDRs thereof) and / or a complete heavy chain and / or light chain, and / or derivatives thereof, comprising the CDRs shown in Table 1. In some preferred embodiments, such antibodies may have the amino acid sequence of a variable heavy chain ("VH") or variable light chain ("VL") polypeptide (VH or VL "chain", respectively) shown below for an antibody or its equivalent, shown in Table 1. In some embodiments, the isolated antibodies or antigen-binding fragments described herein are expressed in an amount of up to 3.6 x 10 -9 Molar (M) K DBinding to human progranulin as measured by the Octet assay described below (see, e.g., the Examples section herein), or as measured by any assay available to the skilled artisan. In some embodiments, the disclosure provides nucleotide sequences encoding specific amino acid sequences of equivalents of the 10C8, 16C 11, 14A6 or 10B3 antibodies, which can be readily obtained from any of the amino acid sequences in Table 1 (i.e., SEQ ID NOs: 1-56) and the information presented in Table 4, and / or equivalents / derivatives thereof. In preferred embodiments, the 10C8 antibody comprises SEQ ID NOs: 1-3 and 7-9; or SEQ ID NOs: 4-6 and 10-12; and / or SEQ ID NOs: 13 and SEQ ID NOs: 14; or variants thereof, as disclosed herein or as would otherwise be recognized by one of ordinary skill in the art. In preferred embodiments, the 16C 11 antibody comprises SEQ ID NOs: 15-17 and 21-23; or SEQ ID NOs: 18-20 and 24-26; and / or SEQ ID NO: 27 and SEQ ID NO: 28; or variants thereof, as disclosed herein or as would otherwise be recognized by one of ordinary skill in the art. In preferred embodiments, the 14A6 antibody comprises SEQ ID NOs: 29-31 and 35-37; or SEQ ID NOs: 32-34 and 38-40; and / or SEQ ID NO: 41 and SEQ ID NO: 42; or variants thereof, as disclosed herein or as would otherwise be recognized by one of ordinary skill in the art. In preferred embodiments, the 10B3 antibody comprises SEQ ID NOs: 43-45 and 49-51; or SEQ ID NOs: 46-48 and 52-54; and / or SEQ ID NO: 55 and SEQ ID NO: 56; or variants thereof, as disclosed herein or as would otherwise be recognized by one of ordinary skill in the art.

[0024] In some embodiments, the present disclosure also provides an expression vector, which includes an isolated nucleic acid comprising such a nucleotide sequence (in a preferred embodiment as shown below, or a derivative thereof) and / or consisting thereof, and a host cell (e.g., a cell line) containing such an expression vector. In some embodiments, the present disclosure provides a method of using the antibody and / or its fragment (e.g., CDR) (collectively referred to herein as "antibody" unless otherwise specified) to treat a cancer involving progranulin. In some embodiments, the present disclosure also provides a method of treating cancer in a patient by identifying a patient with cancer cells expressing progranulin and administering an antibody or antigen-binding fragment as described herein to the patient. It is known that a variety of human cancers express progranulin, including but not limited to ovarian cancer, breast cancer, multiple myeloma, lung cancer, kidney cancer, prostate cancer, hepatocellular carcinoma, uterine cancer, bladder cancer, bile duct cancer, esophageal cancer, gastric cancer, laryngeal cancer, brain cancer, leukemia, and glioblastoma. Therefore, in some preferred embodiments, a fully human monoclonal antibody is provided herein that specifically binds to human progranulin (PGRN) and can be used to treat cancer, such as breast cancer.

[0025] Combinations of antibodies are also contemplated herein, such as a combination of one or more antibodies described herein with another antibody available to a person of ordinary skill in the art. For example, in some embodiments, the combination may be identified, which provides statistically significant differences from the results obtained using only one or more antibodies without using other antibodies (e.g., neutralization assays). In some embodiments, the combination exhibits superimposed activity and / or preferably synergistic activity. In some embodiments, the combination may include 10C8, 16C11, 14A6, or 10B3 antibodies (or derivatives thereof) and other antibodies and / or conjugates. The combination may also include chemotherapeutic agents used in standard treatments, some of which are used in combination with anti-granule protein precursor therapy. The antibodies of such compositions may be different entities, such as two or more different monoclonal antibodies or derivatives thereof, or may be found on the same entity, such as bifunctional antibodies (single antibodies or derivatives thereof containing multiple binding specificities). Such combinations described herein may also be combined with one or more other agents (such as anti-CTLA-4 antibodies, etc.) that may affect immune cell function. One of ordinary skill in the art will recognize that many such combinations may be suitable for use as described herein.

[0026] As used herein, the term "antibody" refers to a complete antibody and any antigen-binding fragment (i.e., "antigen-binding portion") or single chain thereof. A complete antibody is a glycoprotein comprising at least two heavy chains (H) and two light chains (L), which are interconnected by disulfide bonds. Each heavy chain consists of a heavy chain variable region (abbreviated herein as V H) and a heavy chain constant region. The heavy chain constant region consists of three domains: CH1, CH2, and CH3. Each light chain consists of a light chain variable region (abbreviated herein as V L ) and a light chain constant region. The light chain constant region consists of one domain, CL. H and V L The V region can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDRs), interspersed with more conserved regions, termed framework regions (FRs). H and V L It is composed of three CDRs and four FRs, which are arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens. The constant region of an antibody can mediate the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system. The term "chimeric antibody" is an antibody molecule in which (a) the constant region or a portion thereof is changed, replaced or exchanged so that the antigen binding site (variable region) is connected to a constant region that is different or changed in category, effector function and / or species, or to a completely different molecule that gives the chimeric antibody new properties, such as an enzyme, toxin, hormone, growth factor, drug, etc.; or (b) the variable region or a portion thereof is changed, replaced or exchanged for a variable region with a different or changed antigen specificity. For example, a mouse antibody can be modified by replacing the constant region of a mouse antibody with a constant region from a human immunoglobulin. Due to the replacement of the human constant region, the chimeric antibody can retain its specificity for recognizing the antigen while having reduced human antigenicity compared to the original mouse antibody.

[0027] The term "isotype" refers to the antibody class provided by the heavy chain constant region gene (e.g., IgM, IgE, IgG, e.g., IgG1 or IgG4). Isotypes also include modified forms of one of these classes, wherein modifications are made to change the Fc function, e.g., to enhance or reduce effector function or binding to Fc receptors. Isotypes also refer to the antibody class provided by the light chain constant region (e.g., K, λ). The antibody may comprise an Fc region comprising one or more mutations that affect one or more antibody properties, such as stability, glycosylation pattern or other modifications, effector cell function, pharmacokinetics, etc. In some embodiments, the antibody has reduced or minimal glycosylation. In some embodiments, the antibody has eliminated or reduced effector function. Exemplary Fc mutations include, but are not limited to, (i) human IgG1 Fc region mutations L234A, L235A, G237A, and N297A; (ii) human IgG2 Fc region mutations A330S, P331S, and N297A; and (iii) human IgG4 Fc region mutations S228P, E233P, F234V, L235A, delG236, and N297A (EU numbering). In some embodiments, the human IgG2 Fc region comprises A330S and P331S mutations. In some embodiments, the human IgG4 Fc region comprises S288P mutations. In some embodiments, the human IgG4 Fc region comprises S288P and L235E mutations. Antibodies targeting cell surface antigens can trigger immune stimulation and effector functions associated with engagement of Fc receptors (FcRs) on immune cells. There are many Fc receptors that are specific for specific classes of antibodies, including IgG (gamma receptors), IgE (epsilon receptors), IgA (alpha receptors), and IgM (mu receptors). Binding of the Fc region to Fc receptors on the cell surface can trigger many biological responses, including phagocytosis of antibody-coated particles (antibody-dependent cell-mediated phagocytosis, or ADCP), clearance of immune complexes, lysis of antibody-coated cells by killer cells (antibody-dependent cell-mediated cytotoxicity, or ADCC), and release of inflammatory mediators, placental transfer, and control of immunoglobulin production. Additionally, binding of the C1 component of complement to antibodies can activate the complement system. Activation of complement is very important for the lysis of cellular pathogens. However, activation of complement can also stimulate inflammatory responses and may also be involved in autoimmune hypersensitivity reactions or other immune disorders. Variant Fc regions with reduced or eliminated ability to bind to certain Fc receptors can be used to develop therapeutic antibodies and Fc fusion polypeptide constructs that act by targeting, activating, or neutralizing ligand function without damaging or destroying local cells or tissues. An Fc domain monomer refers to a polypeptide chain comprising a second and a third antibody constant domain (eg, CH2 and CH3). In some embodiments, an Fc domain monomer further comprises a hinge domain.In some embodiments, the Fc domain monomer is any immunoglobulin antibody isotype, including IgG, IgE, IgM, IgA and IgD. Additionally, in some embodiments, the Fc domain monomer is any IgG subtype (e.g., IgG 1, IgG2, IgG2a, IgG2b, IgG2c, IgG3 and IgG4). Additional mutations in the Fc domain and the biological consequences of these mutations are well known in the art and can be applied to antibodies herein. See, for example, U.S. Patent Application Publication No. 20220002434.

[0028] As used herein, the term "antigen-binding portion" or "antigen-binding fragment" of an antibody refers to one or more fragments of an intact antibody that retain the ability to specifically bind to a given antigen (e.g., human granulin). The antigen-binding function of an antibody can be performed by a fragment of an intact antibody. Examples of binding fragments encompassed within the term antigen-binding portion or antigen-binding fragment of an antibody include Fab fragments, which are composed of V L 、V G A monovalent fragment consisting of the V, CL and CH1 domains; a F(ab)2 fragment, which is a bivalent fragment comprising two Fab fragments linked by a disulfide bond at the hinge region; G and CH1 domains; the Fd fragment consists of the V L and V G Domain composition F V Fragment; by V H Domain or V L domains (Ward et al., 1989 Nature 341: 544-546); and isolated complementarity determining regions (CDRs). Further, although the two domains V L and V G Encoded by different genes, they can be linked using recombinant methods, via artificial peptide linkers, to enable them to be made into a single protein chain, where V L and V GThe single-chain Fv (scFv) is a monovalent molecule that is paired with the antibody domains to form a monovalent molecule (called a single-chain Fv (scFv); see, e.g., Bird et al., 1988 Science 242: 423-426; and Huston et al., 1988 Proc. Natl. Acad. Sci. 85: 5879-5883). Such single-chain antibodies include one or more antigen-binding portions or fragments of an antibody. These antibody fragments are obtained using conventional techniques known to those skilled in the art, and the fragments are screened for utility in the same manner as intact antibodies. Antigen-binding fragments can also be incorporated into single-domain antibodies, maxibodies, minibodies, intrabodies, bispecific antibodies, trispecific antibodies, tetraspecific antibodies, v-NARs, and bis-scFv (see, e.g., Hollinger and Hudson, 2005, Nature Biotechnology, 23, 9, 1126-1136). The antigen-binding portion of the antibody can be implanted into a scaffold based on a polypeptide, such as type III fibronectin (Fn3) (see U.S. Pat. No. 6,703,199, which describes a fibronectin polypeptide monomer). The antigen-binding fragment can be incorporated into a scaffold comprising a pair of tandem Fv fragments (V H -CH1-V H -CH1) in a single chain molecule, it forms a pair of antigen binding regions together with the complementary light chain polypeptide (Zapata et al., 1995 Protein Eng. 8(10): 1057-1062; and U.S. Patent No. 5,641,870).

[0029] The term "epitope" refers to a protein determinant that can specifically bind to an antibody. An epitope is usually composed of chemically active surface groups of a molecule (e.g., amino acids or sugar side chains) and usually has specific three-dimensional structural characteristics as well as specific charge characteristics. The difference between a conformational epitope and a non-conformational epitope is that the binding of the former is lost in the presence of a denaturing solvent, while the latter is not. As used herein, the term "binding specificity" refers to the ability of a single antibody binding site to react with (e.g., have affinity for) only one antigenic determinant (e.g., epitope). The phrase "specific (or selective) binding" to an antibody (e.g., human progranulin binding antibody) refers to a binding reaction that determines the presence of a homologous antigen in a heterogeneous population of proteins and other biological molecules. The phrases "antibodies that recognize antigens" and "antibodies specific for antigens" are used interchangeably herein with the term "antibodies that specifically bind to antigens." As used herein, the term "affinity" refers to the strength of the interaction between an antibody and an antigen at a single antigenic site. Within each antigenic site, the variable region of the antibody "arm" interacts with the antigen at multiple sites through weak non-covalent forces; the more interactions there are, the stronger the affinity. As used herein, the term "K 结合" or "Ka" is intended to refer to the association rate of a particular antibody-antigen interaction, and as used herein, the term "K 解离 " or "Kd" is intended to refer to the dissociation rate of a particular antibody-antigen interaction. As used herein, the term "K D ” is intended to refer to the dissociation constant, which is obtained by the ratio of Kd to Ka (i.e., Kd / Ka) and is expressed in molar concentration (M). The K of an antibody D The value can be determined using methods recognized in the art. D Methods include measuring surface plasmon resonance using a biosensor system (e.g., a Biacore system), or measuring affinity in solution by solution equilibrium titration (SET). As used herein, the term "high affinity" for an antibody or an antigen-binding fragment thereof (e.g., a Fab fragment) generally refers to an affinity of 10 -9 M or lower K D Antibodies or antigen-binding fragments thereof.

[0030] The term "isolated antibody" refers to an antibody that is substantially free of other antibodies having different antigenic specificities (eg, an isolated antibody that specifically binds progranulin is substantially free of antibodies that specifically bind antigens other than progranulin). However, an isolated antibody that specifically binds progranulin may have cross-reactivity with other antigens (e.g., progranulin from species other than human). In addition, the isolated antibody may be substantially free of other cellular material and / or chemicals. The term "monoclonal antibody" or "monoclonal antibody composition" as used herein refers to a preparation of antibody molecules of single molecular composition. A monoclonal antibody composition exhibits a single binding specificity and affinity for a particular epitope. The term "human antibody" as used herein is intended to include antibodies having variable regions in which both the framework and CDR regions are derived from sequences of human origin. Further, if the antibody contains a constant region, the constant region is also derived from such human sequences, such as human germline sequences or mutant forms of human germline sequences. Human antibodies may include amino acid residues that are not encoded by human sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or somatic mutation in vivo). The term "human monoclonal antibody" refers to an antibody exhibiting a single binding specificity having a variable region in which both the framework and CDR regions are derived from human sequences. In one embodiment, the human monoclonal antibody is produced by a hybridoma comprising (i) a transgenic mouse ... B cells obtained from a non-human animal (e.g., a transgenic mouse) having a genome comprising a human heavy chain transgene and a light chain transgene (ii) are fused to immortalized cells. A "humanized" antibody is an antibody that retains the reactivity of a non-human antibody but is less immunogenic in humans. For example, this can be achieved by retaining the non-human CDR regions and replacing the remainder of the antibody with its human counterpart (i.e., the constant region and the framework portion of the variable region). See, e.g., Morrison et al., Proc. Natl. Acad. Sci. USA, 81: 6851-6855, 1984; Morrison and Oi, Adv. Immunol., 44:65-92, 1988; Verhoeyen et al., Science, 239:1534-1536, 1988; Padlan, Molec. Immun., 28:489-498, 1991; and Padlan, Molec. Immun., 31:169-217, 1994. Other examples of ergonomic technology include, but are not limited to, the Xoma technology disclosed in U.S. Pat. No. 5,766,886.

[0031] The term "amino acid" refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids. Naturally occurring amino acids are the amino acids encoded by the genetic code, as well as amino acids that are subsequently modified, such as hydroxyproline, γ-carboxyglutamate, and O-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as naturally occurring amino acids, i.e., an alpha carbon bound to a hydrogen, a carboxyl group, an amino group, and an R group, such as homoserine, norleucine, methionine sulfoxide, methionine methylsulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as naturally occurring amino acids. Amino acid mimetics refer to chemical compounds that have a structure that is different from the general chemical structure of an amino acid but that functions in a manner similar to a naturally occurring amino acid.

[0032] The term "conservatively modified variant" is applicable to both amino acids and nucleic acid sequences. For a particular nucleic acid sequence, a conservatively modified variant refers to a nucleic acid encoding the same or substantially the same amino acid sequence, or when the nucleic acid does not encode an amino acid sequence, it refers to a substantially identical sequence. Due to the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For example, the codons GCA, GCC, GCG, and GCU all encode the amino acid alanine. Therefore, at each position where the codon is designated as alanine, the codon can be changed to any corresponding codon described without changing the encoded polypeptide. Such nucleic acid variations are "silent variations", which are a type of conservatively modified variations. Each nucleic acid sequence encoding a polypeptide herein also describes each possible silent variation of the nucleic acid. The skilled person will recognize that each codon in the nucleic acid (except AUG, which is generally the only codon for methionine, and TGG, which is generally the only codon for tryptophan) can be modified to produce functionally identical molecules. Therefore, each silent variation of a nucleic acid encoding a polypeptide is implicit in each described sequence.

[0033] For polypeptide sequences, "conservatively modified variants" include single substitutions, deletions or additions to a polypeptide sequence that result in amino acids being replaced by chemically similar amino acids. Conservative substitution tables providing functionally similar amino acids are well known in the art. Such conservatively modified variants are in addition to polymorphic variants, interspecies homologs and alleles, and do not exclude them. The following eight groups include amino acids that are conservatively substituted for each other: 1) alanine (A), glycine (G); 2) aspartic acid (D), glutamic acid (E); 3) asparagine (N), glutamine (Q); 4) arginine (R), lysine (K); 5) isoleucine (I), leucine (L), methionine (M), valine (V); 6) phenylalanine (F), tyrosine (Y), tryptophan (W); 7) serine (S), threonine (T); and 8) cysteine ​​(C), methionine (M) (see, e.g., Creighton, Proteins (1984)). In some embodiments, the term "conservative sequence modifications" is used to refer to amino acid modifications that do not significantly affect or alter the binding properties of the antibody containing the amino acid sequence.

[0034] Amino acid substitutions that are considered conservative and non-conservative as understood by one skilled in the art are shown below in Table 2 using standard three letter or other amino acid abbreviations:

[0035] Table 2:

[0036]

[0037]

[0038] In the context of two or more nucleic acid or polypeptide sequences, the term "identical" or 100% "identity" percentage refers to two or more identical sequences or subsequences. When measured using one of the following sequence comparison algorithms or by manual alignment and visual inspection, to compare and compare on a comparison window or a specified region to obtain maximum consistency, if the two sequences have a specified percentage of identical amino acid residues or nucleotides (that is, in a specified region, or in the entire sequence when not specified, 60% identity, alternatively 65%, 70%, 75%, 80%, 85%, 90%, 95% or 99% identity), then the two sequences are "substantially identical". Alternatively, identity is present in a region of at least about 50 nucleotides (or 10 amino acids) in length, or in a region of 100 to 500 or 1000 or more nucleotides (or 20, 50, 200 or more amino acids) in length. For sequence comparison, a sequence usually serves as a reference sequence, and a test sequence is compared with it. When using a sequence comparison algorithm, test sequence and reference sequence are input to a computer, subsequence coordinates are specified if necessary, and sequence algorithm program parameters are specified. Default program parameters can be used, or alternative parameters can also be specified. Then, the sequence comparison algorithm calculates the percentage sequence identity of the test sequence relative to the reference sequence based on the program parameters. "Comparison window" as used herein comprises a fragment selected from any one of 20 to 600 with reference to the quantity of continuous positions, generally about 50 to about 200, more generally about 100 to about 150, wherein two sequences can be compared after a sequence is optimally compared with the reference sequence with the same number of continuous positions. The sequence alignment method used for comparison is well known in the art. Optimal alignment of sequences for comparison can be performed by, for example, the local homology algorithm of Smith and Waterman (1970) Adv. Appl. Math. 2:482c, the homology alignment algorithm of Needleman and Wunsch, J. Mol. Biol. 48:443, 1970, the similarity search method of Pearson and Lipman, Proc. Nat'l. Acad. Sci. USA 85:2444, 1988, by computer implementations of these algorithms (GAP, BESTFIT, FASTA and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.), or by manual alignment and visual inspection (see, e.g., Brent et al., Current Protocols in Molecular Biology, John Wiley & Sons, Inc. (Ringbou ed., 2003)).Two examples of algorithms suitable for determining percent sequence identity and sequence similarity are the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al., Nuc. Acids Res. 25:3389-3402, 1977 and Altschul et al., J. Mol. Biol. 215:403-410, 1990, respectively. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information. The algorithm first involves identifying high scoring sequence pairs (HSPs) by identifying short words of length W in the query sequence that either match or satisfy some positive-valued threshold score T when aligned with a word of the same length in a database sequence. T is called the neighborhood word score threshold (Altschul et al., supra). These initial neighborhood word hits act as seeds for initiating searches to find longer HSPs containing them. The word hits are extended in both directions along each sequence for as far as the cumulative alignment score can be increased. For nucleotide sequences, the cumulative score is calculated using the parameters M (reward score for matching residue pairs; always > 0) and N (penalty score for mismatched residues; always < 0). For amino acid sequences, the cumulative score is calculated using a scoring matrix. Extension of word hits in each direction is stopped when: the cumulative alignment score drops by the amount X from its maximum value reached; the cumulative score becomes zero or lower due to the accumulation of one or more negative-scoring residue alignments; or the end of either sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses a word length (W) of 11, an expectation value (E) of 10, M=5, N=-4, and a comparison of both chains by default. For amino acid sequences, the BLASTP program uses by default a word length of 3, an expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff and Henikoff, Proc. Natl. Acad. Sci. USA 89:10915, 1989), 50 alignments (B), 10 expectation (E), M=5, N=-4, and a comparison of both strands. The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin and Altschul, Proc. Natl. Acad. Sci. USA 90:5873-5787, 1993). One of the measures of similarity provided by the BLAST algorithm is the smallest sum probability (P(N)), which provides an indication of the probability that a match between two nucleotide or amino acid sequences would occur by chance. For example, a nucleic acid is considered similar to a reference sequence if the smallest sum probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.2, or less than about 0.01, or less than about 0.001.The percent identity between two amino acid sequences can also be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4: 11-17, 1988), which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. In addition, the percent identity between two amino acid sequences can also be determined using the algorithm of Needleman and Wunsch ((J. Mol, Biol. 48: 444-453, 1970), which has been incorporated into the GAP program in the GCG software package (available at the World Wide Web site gcg.com), using either a Blossom 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6.

[0039] In addition to the above sequence identity percentages, another indication that two nucleic acid sequences or polypeptides are substantially identical is that the polypeptide encoded by the first nucleic acid is immunologically cross-reactive with antibodies raised against the polypeptide encoded by the second nucleic acid, as described below. Thus, for example, when the two peptides differ only in conservative substitutions, a polypeptide is generally substantially identical to a second polypeptide. Another indication that two nucleic acid sequences are substantially identical is that the two molecules or their complements hybridize to each other under stringent conditions, as described below. Yet another indication that two nucleic acid sequences are substantially identical is that the same primers can be used to amplify the sequences.

[0040] Preferably, the antibody or antigen-binding fragment thereof comprises one or more amino acid sequences that are at least 70%, at least 75%, at least 80%, at least 85%, at least 88%, at least 90%, at least 92%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identical to at least one of SEQ ID NOs: 1-56 (i.e., CDR sequences, VH sequences and / or VL sequences shown in Table 1). In some embodiments, equivalents of the 10C8, 16C11, 14A6 or 10B3 antibodies include derivatives of one or more CDRs of the 10C8, 16C11, 14A6 or 10B3 antibodies, preferably including up to three (3) conservative amino acid substitutions of the CDRs thereof (see Table 2), provided that the derivatives retain their ability to bind to PGRN, preferably human PGRN (GP88). In some embodiments, equivalents of 10C8, 16C11, 14A6 or 10B3 antibodies include derivatives of one or more VH and / or VL chains of 10C8, 16C11, 14A6 or 10B3 antibodies, preferably including up to ten conservative amino acid substitutions outside their CDRs (see Table 2), provided that the derivatives retain the ability to bind to PGRN (preferably huPGRN). In preferred embodiments, any such substitutions allow the antibody to be conjugated to one or more detectable labels, one or more cytotoxic agents and / or other payloads (e.g., to provide a bispecific antibody), or do not interfere with the conjugation of the antibody.

[0041] The term "nucleic acid" is used interchangeably with the term "polynucleotide" in this article, and refers to deoxyribonucleotides or ribonucleotides and polymers thereof in either single-stranded or double-stranded form. The term encompasses nucleic acids containing known nucleotide analogs or modified backbone residues or bonds, which are synthetic, naturally occurring, and non-naturally occurring, have binding properties similar to reference nucleic acids, and are metabolized in a manner similar to reference nucleotides. Examples of such analogs include, but are not limited to, phosphorothioates, phosphoramidates, methylphosphonates, chiral methylphosphonates, 2-O-methyl ribonucleotides, peptide nucleic acids (PNAs). Unless otherwise indicated, a particular nucleic acid sequence also implicitly encompasses conservatively modified variants (e.g., degenerate codon substitutions) and complementary sequences, as well as sequences that are clearly specified. Specifically, as described in detail below, degenerate codon substitutions can be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with mixed-base and / or deoxyinosine residues (Batzer et al., Nucleic Acid Res. 19:5081, 1991; Ohtsuka et al., J. Biol. Chem. 260:2605-2608, 1985; and Rossolini et al., Mol. Cell. Probes 8:91-98, 1994).

[0042] In certain embodiments, nucleic acid molecules encoding one or more antibodies described herein may be inserted into one or more expression vectors, as discussed in more detail below. In such embodiments, the antibody may be encoded by nucleotides corresponding to an amino acid sequence. The specific combination (codon) of nucleotides encoding various amino acids (AA) is well known in the art, as described in various references used by those skilled in the art (e.g., Lewin, B. Genes V, Oxford University Press, 1994). The amino acid nucleotide sequence encoding the antibody may be determined with reference to, for example, Table 3. Nucleic acid variants may use any combination of nucleotides encoding the antibody.

[0043] Table 3: Codons encoding amino acids (AA)

[0044]

[0045] Encoding variable heavy chain (V H ) and variable light chain (V L An exemplary nucleotide (e.g., cDNA) sequence of an amino acid sequence of

[0046]

[0047]

[0048] and

[0049]

[0050] Variations of these nucleotide sequences are also contemplated herein. Compositions comprising these nucleotide sequences are also contemplated. Such variations encode antibodies or polypeptides comprising CDRs or variants thereof, as disclosed herein or as will be recognized by one of ordinary skill in the art. One of ordinary skill in the art will appreciate that nucleotide sequences encoding specific amino acid sequences of equivalents of 10C8, 16C 11, 14A6 or 10B3 antibodies can be readily derived from the amino acid sequences of any one of SEQ ID NOs: 57-64 and the information presented in Table 3. For example, it can be inferred from the amino acid sequence GYTLTSY (SEQ ID NO: 1) and the information presented in Table 3 that the amino acid sequence can be encoded by the nucleotide sequence GGC TAC ACC CTG ACC AGC TAC (SEQ ID NO: 65). One of ordinary skill in the art will appreciate that nucleotide sequences encoding SEQ ID NOs: 1-56 and derivatives thereof can be inferred in the same manner, and such nucleotide sequences are contemplated herein. The present disclosure also provides expression vectors, which include isolated nucleic acids comprising and / or consisting of such nucleotide sequences (in preferred embodiments, any one of SEQ ID NOs: 57-64 or derivatives thereof), and host cells (eg, cell lines) containing such expression vectors.

[0051] The term "operably connected" refers to the functional relationship between two or more polynucleotide (e.g., DNA) fragments. Typically, the term refers to the functional relationship between a transcriptional regulatory sequence and a transcribed sequence. For example, if a promoter or enhancer sequence stimulates or regulates the transcription of a coding sequence in an appropriate host cell or other expression system, the promoter or enhancer sequence is operably connected to the coding sequence. Typically, a promoter transcriptional regulatory sequence that is operably connected to a transcribed sequence is physically connected to the transcribed sequence, i.e., it is cis-acting. However, some transcriptional regulatory sequences such as enhancers do not need to be physically connected to or located near the coding sequence that enhances transcription.

[0052] As used herein, the term "optimization" refers to a nucleotide sequence that has been altered to encode an amino acid sequence using preferred codons in a production cell or organism, which is generally a eukaryotic cell, such as a Pichia cell, a Chinese hamster ovary cell (CHO) or a human cell. The optimized nucleotide sequence is engineered to retain the amino acid sequence originally encoded by the starting nucleotide sequence (also referred to as the "parent" sequence) completely or as much as possible. The optimized sequence herein has been engineered to have preferred codons in mammalian cells. However, it is also contemplated herein that these sequences are optimally expressed in other eukaryotic cells or prokaryotic cells. The amino acid sequence encoded by the optimized nucleotide sequence is also referred to as optimized.

[0053] The terms "polypeptide" and "protein" are used interchangeably herein to refer to a polymer of amino acid residues. The terms apply to amino acid polymers in which one or more amino acid residues are artificial chemical mimetics of the corresponding naturally occurring amino acids, as well as naturally occurring amino acid polymers and non-naturally occurring amino acid polymers. Unless otherwise indicated, a particular polypeptide sequence also implicitly encompasses conservatively modified variants thereof.

[0054] As used herein, the term "recombinant human antibody" includes all human antibodies prepared, expressed, produced or isolated by recombinant methods, such as antibodies isolated from transgenic or transchromosomal animals (such as mice) with human immunoglobulin genes or hybridomas prepared therefrom, antibodies isolated from host cells transformed to express human antibodies (such as from transfectomas), antibodies isolated from recombinant combinatorial human antibody libraries, and antibodies prepared, expressed, produced or isolated by any other method, the method involving splicing all or part of the human immunoglobulin gene sequence to other DNA sequences. Such recombinant human antibodies have variable regions in which the framework regions and CDR regions are derived from human germline immunoglobulin sequences. However, in certain embodiments, such recombinant human antibodies may be subjected to in vitro mutagenesis (or, when using transgenic animals for human Ig sequences, in vivo somatic mutagenesis), and thus the V H and V L The amino acid sequence of the region is as follows, which, although derived from human germline V H and V L Sequences that are not naturally present in the human antibody germline repertoire.

[0055] The term "recombinant host cell" (or simply "host cell") or "cell line" refers to a cell into which a recombinant expression vector has been introduced. It should be understood that such terms are intended to refer not only to the specific subject cell, but also to the progeny of such cells. Because certain modifications may occur in subsequent generations due to mutations or environmental influences, such progeny may not actually be identical to the parent cell, but are still included within the scope of the term "host cell" or "cell line" as used herein.

[0056] The term "subject" includes humans and non-human animals. Non-human animals include all vertebrates (e.g., mammals and non-mammals), such as non-human primates (e.g., cynomolgus monkeys), sheep, dogs, cows, chickens, amphibians, and reptiles. Unless otherwise indicated, the terms "patient" or "subject" are used interchangeably herein.

[0057] As used herein, the terms "treating" or "treatment" of any disease or pathology (e.g., breast cancer) refer, in one embodiment, to ameliorating the disease or pathology (i.e., slowing or arresting or reducing the development of the disease or at least one clinical symptom thereof). In another embodiment, "treating" or "treatment" refers to alleviating or improving at least one physical parameter, including parameters that may not be discernible by the patient. In yet another embodiment, "treating" or "treatment" refers to modulating the disease or pathology, whether physically (e.g., stabilizing discernible symptoms), physiologically (e.g., stabilizing physical parameters), or both. In yet another embodiment, "treating" or "treatment" refers to preventing or delaying the onset or development or progression of the disease or pathology. "Prevention," when referring to the indications described herein, includes conditions or pathologies associated with cancers expressing progranulin.

[0058] The term "vector" is intended to refer to a polynucleotide molecule capable of transporting another polynucleotide connected thereto. One type of vector is a "plasmid", which refers to a circular double-stranded DNA loop to which additional DNA fragments can be connected. Another type of vector is a viral vector, such as an adeno-associated virus vector (AAV or AAV2), in which additional DNA fragments can be connected to the viral genome. Some vectors can replicate autonomously in the host cell into which they are introduced (e.g., bacterial vectors and additional mammalian vectors with bacterial replication origins). Other vectors (e.g., non-additional mammalian vectors) can be integrated into the genome of the host cell after being introduced into the host cell, and thus replicated together with the host genome. In addition, some vectors can guide the expression of the gene operably connected thereto. Such vectors are referred to herein as "recombinant expression vectors" (or simply "expression vectors"). Generally, the expression vectors utilized in recombinant DNA technology are often in the form of plasmids. In this specification, since plasmids are the most commonly used vector forms, "plasmids" and "vectors" are used interchangeably. However, it is also intended to include expression vectors of other forms that perform equivalent functions, such as viral vectors (e.g., replication-defective retroviruses, adenoviruses, and adeno-associated viruses).

[0059] Antibodies can be conjugated with drugs to form antibody-drug conjugates (ADCs). Generally, ADCs include joints between drugs and antibodies. Joints can be degradable or non-degradable joints. Degradable joints are usually easily degraded in the intracellular environment, such as joints that degrade at the target site, thereby releasing the drug from the antibody. Suitable degradable joints include, for example, joints that are enzymatically degraded, including peptidyl joints that can be degraded by intracellular proteases (such as lysosomal proteases or endosomal proteases), or sugar joints, such as joints containing glucuronic acid that can be degraded by glucuronidase. Peptidyl joints can include, for example, dipeptides, such as valine-citrulline, phenylalanine-lysine or valine-alanine. Other suitable degradable joints include, for example, pH-sensitive joints (such as joints that are hydrolyzed at a pH of less than 5.5, such as hydrazone joints) and joints that degrade under reducing conditions (such as disulfide joints). Non-degradable joints usually release drugs under conditions where the antibody is hydrolyzed by proteases.

[0060] Before being connected to the antibody, the linker has a reactive group that can react with certain amino acid residues, and the connection is achieved through the reactive group. Preferred sulfhydryl-specific reactive groups, and include, for example, maleimide compounds, halogenated amides (e.g., iodine, bromine or chlorine); halogenated esters (e.g., iodine, bromine or chlorine); halogenated methyl ketones (e.g., iodine, bromine or chlorine), benzyl halides (e.g., iodine, bromine or chlorine); vinyl sulfones, pyridine disulfides; mercury derivatives such as 3,6-bis(mercurymethyl)dioxane, and the counterions are acetate, chloride or nitrate; and polymethylene dimethyl sulfide thiosulfonate. The linker may include, for example, maleimide connected to the antibody via thiosuccinimide. The drug may be any cytotoxic, cell growth-inhibiting or immunosuppressive drug. In some embodiments, the linker connects the antibody and the drug, and the drug has a functional group that can be combined with the linker. For example, the drug may have an amino, carboxyl, sulfhydryl, hydroxyl or keto group that can form a bond with the linker. In the case where the drug is directly attached to the linker, the drug has a reactive group before being attached to the antibody. Useful drug classes include, for example, anti-tubulin drugs, DNA minor groove binding agents, DNA replication inhibitors, alkylating agents, antibiotics, folic acid antagonists, antimetabolites, chemosensitizers, topoisomerase inhibitors, vinca alkaloids, etc. Typical cytotoxic drugs include, for example, auristatins, camptothecins, duocar-mycins, etoposide, maytansine and maytansine compounds (e.g., DM1 and DM4), taxanes, benzodiazepines or benzodiazepine-containing drugs (e.g., pyrrolo[1,4]benzodiazepines, PBD, indolebenzodiazepine and oxazolidinone and vinca alkaloids).

[0061] As used herein, drug-linker can be used to form ADC in a simple step. In other embodiments, bifunctional linker compounds can be used to form ADC in two or more steps. For example, cysteine ​​residues react with the reactive moiety of the linker in the first step, and in subsequent steps, the functional group on the linker reacts with the drug to form ADC. Generally, the functional group on the linker is selected to promote the specific reaction with the appropriate reactive group on the drug part. As a non-limiting example, the part based on azide can be used to react specifically with the reactive alkynyl on the drug part. The drug is covalently bound to the linker by 1,3-dipolar cycloaddition between azide and alkynyl. Other useful functional groups include, for example, ketones and aldehydes (suitable for reacting with hydrazides and alkoxyamines), phosphines (suitable for reacting with azides), isocyanates and isothiocyanates (suitable for reacting with amines and alcohols) and activated esters, such as N-hydroxysuccinimide esters (suitable for reacting with amines and alcohols). These and other connection strategies are well known to those skilled in the art, such as those described in "Bioconjugation Technology", the second edition (Elsevier). It will be appreciated by those skilled in the art that when complementary pairs of reactive functional groups are selected for selective reaction between the drug moiety and the linker, each member of the complementary pair can be used for both the linker and the drug.

[0062] The antibodies can be used to treat cancer (e.g., breast cancer, including triple-negative breast cancer [TNBC]) or other diseases that show increased expression of progranulin. Triple-negative breast cancer is a situation where cancer cells do not have receptors for estrogen, progesterone, and HER2 proteins. The term "neutralizing" should be understood as the ability of the antibody to inhibit or block any biological activity of progranulin that causes tumorigenesis, including its ability to stimulate cell proliferation or induce tumor growth in experimental animals and humans. An effective amount of anti-progranulin antibodies is administered to mammals (including humans) by various routes.

[0063] In some preferred embodiments, the disclosure provides an isolated monoclonal antibody, such as a human monoclonal antibody, that binds to PGRN (preferably hu PGRN), and is preferably antibody 10C8, 16C 11, 14A6 or 10B3 (which can be used in combination). In some preferred embodiments, the antibody is derived from human antibodies, human IgG, human IgG1, human IgG2, human IgG2a, human IgG2b, human IgG3, human IgG4, human IgM, human IgA, human IgA 1, human IgA2, human IgD, human IgE, dog antibodies, dog IgGA, dog IgGB, dog IgGC, dog IgGD, chicken antibodies, chicken IgA, chicken IgD, chicken IgE, chicken IgG, chicken IgM, chicken IgY, goat antibodies, goat IgG, mouse antibodies, mouse IgG, pig antibodies, rat antibodies, llama antibodies, alpaca antibodies, shark antibodies and camel antibodies. In some preferred embodiments, the disclosure provides derivatives of antibodies disclosed herein, optionally selected from F ab 、F ab2 , Fab' single-chain antibody, F v , single chain, monospecific antibody, bispecific antibody, trimeric antibody, multispecific antibody, multivalent antibody, chimeric antibody, canine-human chimeric antibody, canine-mouse chimeric antibody, antibody containing canine Fc, humanized antibody, human antibody, canine antibody, CDR-grafted antibody, shark antibody and nanobody.

[0064] In some preferred embodiments, the present disclosure provides derivatives of the antibodies disclosed herein, comprising a detectable label fixedly attached thereto, optionally, wherein the detectable label is selected from fluorescein, DyLight, Cy3, Cy5, FITC, HiLyte Fluor 555, HiILyte Fluor 647, 5-carboxy-2,7-dichlorofluorescein, 5-carboxyfluorescein, 5-FAM, serotonin, 5-serotonin (5-HAT), 6-carboxyfluorescein (6-FAM), FITC, 6-carboxy-1,4-dichloro-2',7'-dichlorofluorescein (TET), 6-carboxy-1,4-dichloro-2',4',5',7'-tetrachlorofluorescein (HEX), 6-carboxy-4',5'-dichloro-2',7'-tetrachlorofluorescein (TEF ...7'-Dimethoxyfluorescein (6-JOE), Alexa fluor, Alexa fluor 350, Alexa fluor 405, Alexa fluor 430, Alexa fluor 488, Alexa fluor 500, Alexa fluor 514, Alexa fluor 532, Alexa fluor 546, Alexa fluor 555, Alexa fluor568, Alexa fluor 594. Alexa fluor610, Alexa fluor 633, Alexa fluor 635, Alexa fluor 647, Alexa fluor 660, Alexa fluor 680, Alexa fluor 700, Alexa fluor 750, BODIPY fluorophore, BODIPY 492 / 515, BODIPY493 / 503, BODIPY 500 / 510, BODIPY 505 / 515, BODIPY 530 / 550, BODIPY 542 / 563, BODIPY558 / 568, BODIPY 564 / 570, BODIPY 576 / 589, BODIPY 581 / 591, BODIPY 630 / 650-X, BODIPY650 / 665-X, BODIPY 665 / 676, FL, FL ATP, FI-ceramide, R6G SE, TMR, TMR-X conjugate, TMR-X, SE, TR, TR ATP, TR-X SE, Rhodamine, Rhodamine 110, Rhodamine 123, Rhodamine B, Rhodamine B 200, Rhodamine BB, Rhodamine BG, Rhodamine B extra, 5-carboxytetramethylrhodamine (5-TAMRA), 5GLD, 6-carboxyrhodamine 6G, Lissamine, Lissamine Rhodamine B, Phallicidine, Phalloidin, Rhodamine Red, Rhod-2, 6-Carboxy-X-Rhodamine (ROX), Carboxy-X-Rhodamine (5-ROX), Sulforhodamine B can C, Sulforhodamine G Extra, 6-Carboxytetramethylrhodamine (TAMRA), Tetramethylrhodamine (TRITC), Rhodamine WT, Texas Red, and Texas Red-X. ,

[0065] In some preferred embodiments, the disclosure provides derivatives of antibodies disclosed herein, comprising an effector portion attached thereto, optionally, wherein the effector portion is selected from cytotoxic drugs, toxins, diphtheria toxin A chain, exotoxin A chain, ricin A chain, abrin A chain, curcin, crotin, phenomycin, enomycin, and radiochemicals, optionally, comprising a cleavable linker between the antibody and the effector portion, wherein the cleavable linker releases the effector portion into or within the cell. In some preferred embodiments, the disclosure provides isolated polynucleotides encoding antibodies of the disclosure, optionally, wherein the nucleic acid sequences are those disclosed herein, and expression vectors and host cells comprising the polynucleotides. In some preferred embodiments, the disclosure provides compositions comprising at least one antibody or derivative disclosed herein; at least one isolated polynucleotide encoding such an antibody or derivative; or at least one expression vector comprising such a polynucleotide; and / or, at least one host cell comprising such a polynucleotide and / or an expression vector; or a combination thereof; and a pharmaceutically acceptable carrier.

[0066] In some preferred embodiments, the disclosure provides a method for detecting PGRN on a cell, the method comprising contacting a test biological sample with an antibody or derivative of the disclosure, and detecting an antibody that binds the biological sample or its components. In some embodiments, the method comprises comparing the binding amount of the test biological sample or its components with the binding amount of the control biological sample or its components, wherein the binding of the test biological sample or its components relative to the increase in the control biological sample or its components indicates that there are cells expressing PGRN (preferably hu PGRN) in the test biological sample (e.g., wherein the test biological sample is a mammalian cell, tissue, or blood). The method can be an in vivo method or an in vitro method.

[0067] In some preferred embodiments, the disclosure provides a method for treating, preventing and / or improving mammalian cancer, comprising administering to a mammal at least one effective dose of a pharmaceutical composition comprising an antibody or derivative of the disclosure. In some embodiments, the disclosure provides such antibodies, comprising a cytotoxic effector portion attached thereto, optionally wherein the effector portion is selected from a cytotoxic drug, a toxin, a diphtheria toxin A chain, an exotoxin A chain, a ricin A chain, abrin A chain, curcin, crotin, phenomycin, enomycin, and radiochemicals. In some embodiments, such antibodies may include a cleavable linker between the antibody and the effector portion, wherein the cleavable linker releases the effector portion into a cell or releases it within the cell. In some such embodiments, the antibody is administered as an antibody-drug conjugate. In some embodiments, multiple doses are administered to an animal; and / or, the antibody is administered at a dose of about 1 to 50 mg / kg.

[0068] The antibodies (e.g., polypeptides) and nucleic acids described herein may also be combined with one or more pharmaceutically acceptable carriers prior to administration to a host. A pharmaceutically acceptable carrier is a biologically or otherwise benign material, e.g., one that can be administered to a subject without causing any adverse biological effects or interacting in a deleterious manner with any other component of a pharmaceutical composition containing the material. The carrier will naturally be selected to minimize any degradation of the active ingredient and to minimize any deleterious side effects in the subject, as is well known to those skilled in the art. Suitable pharmaceutical carriers and their formulations are described, e.g., in Remington's: The Science and Practice of Pharmacy, 21 stEdition, David B.Troy, ed., described in Lippicott Williams & Wilkins (2005). Generally, an appropriate amount of pharmaceutically acceptable salt is used in the preparation to make the preparation isotonic. Examples of pharmaceutically acceptable carriers include, but are not limited to, sterile water, saline, buffered solutions such as Ringer's solution, and glucose solutions. The pH of the solution is generally from about 5 to about 8 or from about 7 to about 7.5. Other carriers include sustained-release preparations, such as semipermeable matrices of solid hydrophobic polymers containing polypeptides or fragments thereof. The matrix can be in the form of a molded article, such as a film, liposome, or microparticle. It is obvious to those skilled in the art that certain carriers will be more preferred, depending on, for example, the route of administration and concentration of the composition being administered. The carrier is a carrier suitable for administering polypeptides and / or fragments thereof to humans or other subjects. In addition to the immunogenic polypeptide, the pharmaceutical composition may also include a carrier, a thickener, a diluent, a buffer, a preservative, a surfactant, an adjuvant, an immunostimulant. The pharmaceutical composition may also include one or more active ingredients, such as antimicrobial agents, anti-inflammatory agents, and anesthetics. Pharmaceutical composition can be in the dosage unit preparation containing conventional pharmaceutically acceptable carrier, adjuvant and carrier, orally, parenterally, by inhalation spray, rectum, knot (intranodal) or topically applied.Term " pharmaceutically acceptable carrier " or " physiologically acceptable carrier " as used herein refers to one or more preparation materials suitable for completing or enhancing nucleic acid, polypeptide or peptide as the delivery of pharmaceutical composition. " pharmaceutical composition " is the composition comprising the nucleic acid or polypeptide of therapeutically effective amount.Term " effective amount " and " therapeutically effective amount " refer to the amount of antibody, nucleic acid etc. for observing required therapeutic effect (for example, eliminating the cell expressing PGRN, for example, the cancerous cell expressing PGRN) separately.

[0069] Also provided are methods for treating one or more disease symptoms (e.g., cancer) in a mammalian host, comprising administering to the mammal at least one or more effective doses of one or more antibodies (and / or derivatives thereof) described herein. In some embodiments, the antibody is a monoclonal antibody or a fragment or derivative thereof comprising one or more of a combination of CDRs and / or variable regions of antibodies 10C8, 16C11, 14A6, or 10B3; an amino acid sequence shown in Table 1 and / or an amino acid sequence encoded by a nucleotide sequence disclosed herein or recognizable by a person of ordinary skill in the art; and / or substituted derivatives thereof and / or fragments thereof; and in some embodiments, conservatively substituted variants thereof. The one or more antibodies can be administered at a dose of about 1 to about 50 mg / kg, about 1 to about 30 mg / kg, or about 5 to about 30 mg / kg (e.g., any of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, or 40 mg / kg). In certain embodiments, the one or more antibodies can be administered to a mammal once or multiple times (e.g., intradermally, intravenously, orally, rectally) at about 10 mg / kg. When multiple doses are administered, the doses can contain approximately the same or different amounts of the antibody in each dose. The doses can also be separated from each other in time by the same or different intervals. For example, the dosage can be separated by about 6, 12, 24, 36, 48, 60, 72, 84 or 96 hours, any one of a week, two weeks, three weeks, one month, two months, three months, four months, five months, six months, seven months, eight months, nine months, ten months, eleven months, twelve months, 1.5 years, two years, three years, four years, five years or any time period before, after and / or between any of these time periods. In some embodiments, the antibody can be administered in combination with other agents (e.g., anti-infective agents and / or chemotherapeutic agents). Such other agents can be administered simultaneously with the antibody, or at different times and / or frequencies. As can be easily determined by a person of ordinary skill in the art, other embodiments of such methods will also be suitable.

[0070] In some preferred embodiments, the present disclosure provides an isolated antibody or antigen-binding fragment thereof, comprising: a) a heavy chain variable region comprising CDR sequences of SEQ ID NOs: 1, 2 and 3, respectively, and a light chain variable region comprising CDR sequences of SEQ ID NOs: 4, 5 and 6, respectively; b) a heavy chain variable region comprising CDR sequences of SEQ ID NOs: 7, 8 and 9, respectively, and a light chain variable region comprising CDR sequences of SEQ ID NOs: 10, 11 and 12, respectively; c) a heavy chain variable region comprising CDR sequences of SEQ ID NOs: 13, 14 and 15, respectively, and a light chain variable region comprising CDR sequences of SEQ ID NOs: 16, 17 and 18, respectively; d) a heavy chain variable region comprising CDR sequences of SEQ ID NOs: 19, 20, 21, respectively, and a light chain variable region comprising CDR sequences of SEQ ID NOs: 22, 23 and 24, respectively; e) a heavy chain and a light chain variable region comprising CDR sequences of SEQ ID NOs: NO: 1-3 and 7-9; f) heavy and light chain variable regions comprising CDR sequences SEQ ID NO: 4-6 and 10-12, respectively; g) heavy and light chain variable regions comprising SEQ ID NO: 13 and SEQ ID NO: 14, respectively; h) heavy and light chain variable regions comprising CDR sequences SEQ ID NO: 15-17 and 21-23, respectively; i) heavy and light chain variable regions comprising CDR sequences SEQ ID NO: 18-20 and 24-26, respectively; j) heavy and light chain variable regions comprising SEQ ID NO: 27 and SEQ ID NO: 28, respectively; k) heavy and light chain variable regions comprising CDR sequences SEQ ID NO: 29-31 and 35-37, respectively; 1) heavy and light chain variable regions comprising CDR sequences SEQ ID NO: 32-34 and 38-40, respectively; m) heavy and light chain variable regions comprising CDR sequences SEQ ID NO: 41 and SEQ ID NO: 42; n) heavy chain and light chain variable regions, which respectively contain CDR sequences SEQ ID NO: 43-45 and 49-51; o) heavy chain and light chain variable regions, which respectively contain CDR sequences SEQ ID NO: 46-48 and 52-54; p) heavy chain and light chain variable regions, which respectively contain SEQ ID NO: 55 and SEQ ID NO: 56; or a derivative of any one of a)-p); wherein the antibody or antigen-binding fragment thereof specifically binds to human progranulin.In some preferred embodiments, the disclosure provides an isolated antibody or antigen-binding fragment thereof comprising: a) a heavy chain variable region comprising SEQ ID NO: 25 and a light chain variable region comprising SEQ ID NO: 27, respectively; b) a heavy chain variable region comprising SEQ ID NO: 29 and a light chain variable region comprising SEQ ID NO: 31, respectively; c) a heavy chain variable region comprising SEQ ID NO: 33 and a light chain variable region comprising SEQ ID NO: 35, respectively; or d) a heavy chain variable region comprising SEQ ID NO: 37 and a light chain variable region comprising SEQ ID NO: 39, respectively. In some preferred embodiments, the antibody is internalized into cells expressing huPGRN in vitro and / or in vivo. In some preferred embodiments, the antibody competes with trastuzumab for binding to the hu PGRN receptor on the cell. In some preferred embodiments, the antibody does not compete with trastuzumab for binding to the hu PGRN receptor on the cell. In some preferred embodiments, the antibody is 10C8, 16C11, 14A6 and 10B3, or a derivative thereof. In some preferred embodiments, the disclosure provides a combination of such antibodies. In some preferred embodiments, the antibody is a separated monoclonal antibody. In some preferred embodiments, the antibody is a human monoclonal antibody. In some preferred embodiments, the antibody is derived from human antibodies, human IgG, human IgG1, human IgG2, human IgG2a, human IgG2b, human IgG3, human IgG4, human IgM, human IgA, human IgA1, human IgA2, human IgD, human IgE, dog antibodies, dog IgGA, dog IgGB, dog IgGC, dog IgGD, chicken antibodies, chicken IgA, chicken IgD, chicken IgE, chicken IgG, chicken IgM, chicken IgY, goat antibodies, goat IgG, mouse antibodies, mouse IgG, pig antibodies, rat antibodies, llama antibodies, alpaca antibodies, shark antibodies and camel antibodies. In some preferred embodiments, the antibody derivative is selected from F. ab 、F ab2 , Fab' single-chain antibody, F v, single chain, monospecific antibody, bispecific antibody, trimeric antibody, multispecific antibody, multivalent antibody, chimeric antibody, canine-human chimeric antibody, canine-mouse chimeric antibody, antibody containing canine Fc, humanized antibody, human antibody, canine antibody, CDR-grafted antibody, shark antibody and nanobody. In some preferred embodiments, the antibody or derivative comprises a detectable label fixedly attached thereto, optionally wherein the detectable label is selected from fluorescein, DyLight, Cy3, Cy5, FITC, HiLyteFluor 555, HiLyteFluor 647, 5-carboxy-2,7-dichlorofluorescein, 5-carboxyfluorescein, 5-FAM, serotonin, 5-serotonin (5-HAT), 6-carboxyfluorescein (6-FAM), FITC, 6-carboxy-1,4-dichloro-2',7'-dichlorofluorescein (TET), 6-carboxy-1,4-dichloro-2',4',5',7'-tetrachlorofluorescein (HEX), 6-carboxy-4',5'-dichloro-2',7'-Dimethoxyfluorescein (6-JOE), Alexa fluor, Alexa fluor 350, Alexa fluor 405, Alexa fluor 430, Alexa fluor 488, Alexa fluor 500, Alexa fluor 514, Alexa fluor 532, Alexa fluor546, Alexa fluor 555, Alexa fluor 568, Alexa fluor594, Alexa fluor 610, Alexa fluor 633, Alexa fluor 635, Alexa fluor 647, Alexa fluor 660, Alexa fluor 680, Alexa fluor 700, Alexa fluor 750, BODIPY fluorophore, BODIPY 492 / 515, BODIPY 493 / 503, BODIPY 500 / 510, BODIPY 505 / 515, BODIPY 530 / 550, BODIPY 542 / 563, BODIPY 558 / 568, BODIPY 564 / 570, BODIPY 576 / 589, BODIPY 581 / 591, BODIPY 630 / 650-X, BODIPY 650 / 665-X, BODIPY 665 / 676, FL, FL ATP, FI-ceramide, R6G SE, TMR, TMR-X conjugate, TMR-X, SE, TR, TRATP, TR-X SE, Rhodamine, Rhodamine 110, Rhodamine 123, Rhodamine B, Rhodamine B 200, Rhodamine BB, Rhodamine BG, Rhodamine B extra, 5-carboxytetramethylrhodamine (5-TAMRA), 5GLD, 6-carboxyrhodamine 6G, lissamine, lissamine rhodamine B, Phallicidine, phalloidin, rhodamine red, Rhod-2,6-Carboxy-X-Rhodamine (ROX), Carboxy-X-Rhodamine (5-ROX), Sulforhodamine B can C, Sulforhodamine G Extra, 6-Carboxytetramethylrhodamine (TAMRA), Tetramethylrhodamine (TRITC), Rhodamine WT, Texas Red and Texas Red-X. In some preferred embodiments, the antibody or derivative comprises an effector portion attached thereto, optionally, wherein the effector portion is selected from a cytotoxic drug, a toxin, a diphtheria toxin A chain, an exotoxin A chain, a ricin toxin A chain, abrin A chain, curcin, crotin, phenomycin, enomycin and a radiochemical. In some preferred embodiments, the antibody or derivative may comprise a cleavable linker between the antibody and the effector portion, wherein the cleavable linker releases the effector portion into or within the cell. In some preferred embodiments, the present disclosure provides an isolated polynucleotide encoding an antibody disclosed herein, optionally, wherein the nucleic acid sequence is at least one of SEQ ID NOs: 57-64. In some preferred embodiments, the polynucleotide may be an expression vector comprising one or more such polynucleotides. In some preferred embodiments, the disclosure provides host cells comprising such isolated polynucleotides and / or expression vectors. In some preferred embodiments, the disclosure provides compositions comprising any such at least one antibody and / or derivative; at least one isolated polynucleotide encoding thereof; at least one expression vector comprising and / or encoding these; and / or, at least one host cell comprising these; or any combination thereof; and a pharmaceutically acceptable carrier. ,

[0071] In some preferred embodiments, the disclosure provides a method for detecting hu PGRN (GP88) on a cell, the method comprising contacting a test biological sample with an antibody or derivative of the disclosure, and detecting the antibody or derivative bound to the biological sample or a component thereof. In some preferred embodiments, the cell may be selected from breast cancer, adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer, basal breast cancer, estrogen receptor (ER) luminal breast cancer, ER positive tamoxifen resistant luminal breast cancer, letrozole resistant luminal breast cancer, bile duct cancer, bladder cancer, brain cancer, glioblastoma, colorectal cancer, epidermoid carcinoma, squamous cell carcinoma, esophageal cancer, gastric cancer, hepatocellular carcinoma, kidney / renal cancer, laryngeal cancer, lung cancer, non-small cell lung cancer, lung adenocarcinoma, mesothelioma, myeloma / leukemia, ovarian cancer, prostate cancer, and uterine cancer cells. In some preferred embodiments, the method comprises comparing the binding amount of the test biological sample or its component and the binding amount of the control biological sample or its component, wherein the binding of the test biological sample or its component relative to the increase of the control biological sample or its component indicates that there are cells expressing hu PGRN in the test biological sample. In some preferred embodiments, the test biological sample can be a mammalian cell, tissue or blood. In some preferred embodiments, the method is an in vivo method or an in vitro method.

[0072] In some preferred embodiments, the disclosure provides a method for treating, preventing and / or improving mammalian cancer, comprising administering to a mammal at least one effective dose of a pharmaceutical composition comprising an antibody or derivative of the disclosure. In some preferred embodiments, the cancer is selected from breast cancer, adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer, basal breast cancer, estrogen receptor (ER) luminal breast cancer, ER positive tamoxifen resistant luminal breast cancer, letrozole resistant luminal breast cancer, bile duct cancer, bladder cancer, brain cancer, glioblastoma, colorectal cancer, epidermoid carcinoma, squamous cell carcinoma, esophageal cancer, gastric cancer, hepatocellular carcinoma, kidney cancer, laryngeal cancer, lung cancer, non-small cell lung cancer, lung adenocarcinoma, mesothelioma, myeloma / leukemia, ovarian cancer, prostate cancer and uterine cancer. In some preferred embodiments, the antibody comprises a cytotoxic effector portion attached thereto, optionally wherein the effector portion is selected from a cytotoxic drug, a toxin, a diphtheria toxin A chain, an exotoxin A chain, a ricin A chain, abrin A chain, curcin, crotin, phenomycin, enomycin, and a radiochemical. In some preferred embodiments, the antibody further comprises a cleavable linker between the antibody and the effector portion, wherein the cleavable linker releases the effector portion into or within the cell. In some preferred embodiments, the antibody is administered as an antibody-drug conjugate. In some preferred embodiments, multiple doses are administered to the animal; and / or, the antibody is administered at a dose of about 1 to 50 mg / kg.

[0073] In some preferred embodiments, the disclosure provides a kit for detecting hu PGRN (GP88) expression in or on a cell, the kit comprising an antibody or derivative and instructions for use of any one of the preceding claims. In some preferred embodiments, the cell is selected from breast cancer, adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer, basal breast cancer, estrogen receptor (ER) luminal breast cancer, ER positive tamoxifen resistance luminal breast cancer, letrozole resistance luminal breast cancer, bile duct cancer, bladder cancer, brain cancer, glioblastoma, colorectal cancer, epidermoid carcinoma, squamous cell carcinoma, esophageal cancer, gastric cancer, hepatocellular carcinoma, kidney / renal cancer, laryngeal cancer, lung cancer, non-small cell lung cancer, lung adenocarcinoma, mesothelioma, myeloma / leukemia, ovarian cancer, prostate cancer and uterine cancer cells. In some preferred embodiments, the antibody or derivative is in lyophilized form.

[0074] The following examples relate to the monoclonal antibodies described herein.

[0075] Example 1: Discovery and isolation of a high affinity fully human monoclonal antibody against human progranulin

[0076] Humanized mouse TC-mAb was expressed using human recombinant histidine-tagged full-length progranulin TM Immunization. The characteristics of these mice have been published (Moriwak1, A. et al., Exp Cell Res 390(2): 111914, 2020). Tc mice that produce human Abs (TC-mAb mice) stably maintain an engineered chromosome derived from mice in a mouse Ig knockout background, the chromosome containing complete human Ig heavy chain and K chain loci. Transchromosome (Tc) mice carrying mini-chromosomes with human immunoglobulin (Ig) loci can help develop fully human therapeutic monoclonal antibodies (Abs) when immunized with an antigen of interest. In this case, TC-mAb mice were immunized with human recombinant progranulin. The titer of sera from immunized mice was checked by EIA with human progranulin fixed on a nickel plate, and (progranulin) EIA was performed with an HRP-conjugated goat anti-human Fc secondary antibody. Mice with the highest anti-progranulin titers were used to collect spleen and lymph node B cells, which were fused with mouse myeloma HL-1 cells by electroporation. The fused hybridoma single cells were inoculated in semi-solid hybridoma medium D in 10 cm tissue culture plates. After 11 days, 1,726 single hybridoma clones were picked from the semi-solid medium plates and transferred to 96-well plates in hybridoma medium E (one clone per well). After 3 days, the culture medium of the hybridoma clones was assayed with progranulin EIA. The assay provided a large number of clones. The OD of 184 clones was 0.144. 650 >3.0, OD of 387 clones 650 OD of 423 clones was between 3.0 and 2.0 650 Between 2.0 and 1.0, and OD of 334 clones 650 Between 1 and 0.5. 650 The first 184 clones with p values ​​>3.0 were transferred to 48-well plates for confirmation screening by progranulin EIA, as described above. 170 clones were confirmed as strongly positive. These clones were transferred in duplicate to 6-well plates in medium E. The cells were cryopreserved under appropriate culture conditions for long-term storage in liquid nitrogen, and the culture medium containing secreted Ig was collected and stored for future evaluation and selection of hybridomas of interest. From these clones, 120 clones producing antibodies were prepared.

[0077] Example 2. Further antigen binding characterization and selection of anti-progranulin monoclonal antibody clones

[0078] In vitro proliferation assay. The ability of culture media from selected clones to inhibit proliferation of TNBC cells MDA-MB-231 was determined by tracking inhibition of phosphorylation of ERK1 / 2 (p-ERK1 / 2) and AKT (p-AKT), which are well-known signaling molecules in the proliferation and survival pathways of cells, including cancer cells. This assay was described by Guha et al. in 2021. Using this assay, the results showed that the addition of AG01 inhibited phosphorylation of ERK1 / 2 (MAP kinase or MAPK) and the survival signaling molecule p-AKT in a dose-dependent manner. All fully human hybridoma clones that tested positive for binding to progranulin by EIA were tested in this signaling molecule phosphorylation assay with MDA-MB-231 triple-negative breast cancer cells. According to these assays, 45 clones showed levels of inhibitory activity against p-ERK1 / 2, p-AKT, or both of them >50% when compared to control cells treated with human IgG as a control. Repeated assays further narrowed the number of clones showing p-ERK1 / 2 inhibition to 38. The results are shown in Table 4.

[0079] Table 4: Activity of cloned monoclonal antibodies

[0080]

[0081]

[0082]

[0083]

[0084] Clones producing antibodies that inhibited pERK1 / 2 and / or p-AKT by >50% are highlighted in grey. These 45 clones were then investigated for their ability to inhibit progranulin binding to MDA-MB-231 cells in a dose-dependent manner.

[0085] Progranulin binding by flow cytometry Binding assays were performed on an INTELLICYT flow cytometer using the instructions included therein. Additional binding assays were performed, including the ability of anti-progranulin antibodies to inhibit progranulin binding to two cell lines (human embryonic kidney HEK-293 and TNBC MDA-MB-231) as well as several other cancer cells (e.g., non-small cell carcinoma cell line H1299). Of the 45 mAbs assayed, 11 clones (indicated in bold in Table 5) produced antibodies that were able to inhibit progranulin binding in a dose-dependent manner.

[0086] Table 5: Inhibition of progranulin flow cytometric binding on MDA-MB-231 cells

[0087]

[0088] Several antibodies that were able to block p-ERK1 / 2 in in vitro assays had little inhibitory activity on progranulin binding, or even stimulated progranulin binding to cells. After further screening by combining several assays, four antibodies were tested. Figure 1 It was shown that human anti-progranulin antibodies inhibited the binding of progranulin to TNBCMDA-MB-231 cells in a dose-dependent manner and had greater activity than AG0 1, with the exception of 14A6, the addition of which resulted in an increase in progranulin binding in this assay.

[0089] Binding affinity determination by Octet . OctetRed96 BLI technology was used to examine the binding properties of selected fully human antibodies to progranulin. The method is briefly described herein. The assays were performed at 25°C. Pre-hydrated (1 hour) anti-human Fc capture (AHC) sensors were pre-treated by repeating 3 cycles of immersing the sensor in regeneration buffer followed by immersion in neutralization buffer (PBS). Anti-progranulin mAb was captured on the pre-treated biosensor at 20 μg / mL for 360 seconds. The sensor was then immersed in a baseline buffer similar to the progranulin diluent (PBS). The sensor was subsequently immersed in a well containing progranulin as shown for 420 seconds. The dissociation step was performed in assay buffer (PBS) for 600 seconds. Using the experimental conditions described above, based on K on and K off Determine K D Table 6 shows the K values ​​of selected mAbs. D Specifically, mAbs 16C11, 10B3, 10C8, and 14A6 had less than 10 -9 M of K D , among which 10C8 has the highest affinity, and its K D For 4×10 -12 M. These four mAbs were determined to contain the IgG1 / κ isotype based on Iso-Gold Rapid Isotyping.

[0090] Table 6: K values ​​of selected monoclonal antibodies determined using Octet D *

[0091]

[0092] *Sensor type: AHC; Antibody concentration: 50nM; Analyte: huPGRN

[0093] Monoclonal antibodies binding to the AG01 epitope. Experiments were performed to determine whether any of the selected fully human antibodies compete with AG01 for binding to progranulin, in this way determining whether these mAbs have epitopes that are similar or overlapping with the epitope of mAb AG0 1. This was done using biofilm interferometry on OctetRed96 according to the method described in Dafferner, A. et al., Chem Res Toxicol 30:1897-1910, 2017. Each clone was tested against AG01 on OctetRed96 and the tandem monoclonal co-binding with progranulin was tested. These assays were essentially performed by capturing AG01 on the AHC sensor or progranulin on the HIS1K sensor and testing the pairing or blocking ability of the tested antibodies. The results showed that the four selected fully human progranulin antibody clones bound to the progranulin that was previously occupied by AG01 without any interference. These data indicate that the four selected antibodies 16C11, 11C8, 10B3 and 14A6 bind to epitopes that are distinct from those bound by AG01. These results were confirmed by detailed surface plasmon resonance analysis of AG01 and mAb 16C11 using Biacore using the method described by Myszka, D. et al. (Biophys J 75:583-59, 1998).

[0094] Example 3: Functional activity of monoclonal antibodies against progranulin

[0095] Migration assay The migration assay described by Guha et al., 2021 was followed. Eleven clones were examined in the Transwell chamber migration assay to select the clone that showed the highest inhibition of TNBC MDA-MB-231 cell migration in a dose-dependent manner compared to AG0 1 as a positive control. Six clones were further retained for their ability to inhibit the migration of MDA-MB-231 cells in a dose-dependent manner. Figure 2 The results in the study showed that these mAbs could inhibit the migration of MDA-MB-231 cells at levels comparable to or even higher than AG01. Specifically, fully human mAbs added at 10 μg / mL inhibited the migration of MDA-MB-231 cells by 30% to 50%, while AG01 required a concentration of 50 μg / mL (Guha et al., 2021) to achieve similar levels of migration inhibition.

[0096] Example 4: Effect of anti-progranulin monoclonal antibodies on tumorigenesis in vivo

[0097] Subsequently, these mAbs with the highest affinity (16C11, 10C8, 10B3, and 14A6) were examined in vivo for their ability to block tumor growth of MDA-MB-231 cells injected subcutaneously into the flank of female athymic nude mice. It has been shown that progranulin expression is associated with increased tumorigenesis ((Tangkeangsirisin, Hayashi et al., 2004), and mice bearing TNBC tumors exhibit inhibition of tumor growth when treated with the anti-progranulin antibody AG01 (Guha et al., 2021). Therefore, we examined the ability of fully human anti-progranulin antibodies to inhibit tumor growth of the TNBC cell line MDA-MB-231 using the methods described in Guha et al., 2021. Three antibodies were tested in these nude mice. All antibodies were assayed at a concentration of 5 mg / kg. Athymic female mice were injected subcutaneously with 10 6 MDA-MB-231 cells. When the tumor reaches 100 mm 3 At 1 , mice were randomly divided into each experimental group with seven animals in each group. Antibodies were injected intraperitoneally at a dose of 5 mg / kg twice a week. Tumor size was determined using a caliper twice a week before injection of antibodies, and tumor volume was determined from the size of the tumor.

[0098] Figure 3 The effects of intraperitoneal injection of fully human antibodies on tumor volume determined at treatment day 25 are shown when compared to AG01 positive control and human IgG negative control. The data show that the selected antibodies exhibited varying degrees of tumor inhibition when compared to AG0 1 used as a positive control. These data indicate that the four selected antibodies are able to inhibit tumor growth of TNBC cell lines with similar or greater efficacy than mAb AG0 1.

[0099] Based on these data and the binding data of mAb 10C8, the antitumor activities of mAb 10C8 and AG01 were further compared throughout the treatment course. Figure 4 The graph in shows the inhibition of tumor growth by 10C8 throughout the treatment. At the same dose of 5 mmg / kg, 10C8 had twice the tumor inhibitory activity of AG01 (P<0.006). When compared to human IgG as a control, twice weekly injections of 10C8 inhibited tumor growth by 50%.

[0100] Other embodiments will be apparent to those skilled in the art in view of the specification and teachings provided herein.It is intended that the specification and examples be considered exemplary only, with the true scope and spirit of the disclosure being indicated by the following claims.

Claims

1. An isolated antibody or antigen-binding fragment thereof comprising: a) a heavy chain variable region comprising the CDR sequences of SEQ ID NOs: 1, 2 and 3, respectively, and a light chain variable region comprising the CDR sequences of SEQ ID NOs: 4, 5 and 6, respectively; b) a heavy chain variable region comprising the CDR sequences of SEQ ID NOs: 7, 8 and 9, respectively, and a light chain variable region comprising the CDR sequences of SEQ ID NOs: 10, 11 and 12, respectively; c) a heavy chain variable region comprising CDR sequences of SEQ ID NOs: 13, 14 and 15, respectively, and a light chain variable region comprising CDR sequences of SEQ ID NOs: 16, 17 and 18, respectively; d) a heavy chain variable region comprising CDR sequences of SEQ ID NOs: 19, 20, 21, respectively, and a light chain variable region comprising CDR sequences of SEQ ID NOs: 22, 23, and 24, respectively; e) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 1-3 and 7-9, respectively; f) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 4-6 and 10-12, respectively; g) heavy and light chain variable regions comprising SEQ ID NO: 13 and SEQ ID NO: 14, respectively; h) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 15-17 and 21-23, respectively; i) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 18-20 and 24-26, respectively; j) heavy and light chain variable regions comprising SEQ ID NO: 27 and SEQ ID NO: 28, respectively; k) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 29-31 and 35-37, respectively; 1) heavy chain and light chain variable regions comprising CDR sequences SEQ ID NOs: 32-34 and 38-40, respectively; m) heavy and light chain variable regions comprising SEQ ID NO:41 and SEQ ID NO:42, respectively; n) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 43-45 and 49-51, respectively; o) heavy and light chain variable regions comprising CDR sequences of SEQ ID NOs: 46-48 and 52-54, respectively; p) heavy and light chain variable regions comprising SEQ ID NO: 55 and SEQ ID NO: 56, respectively; or a derivative of any one of a) to p); The antibody or antigen-binding fragment thereof specifically binds to human progranulin.

2. The isolated antibody or antigen-binding fragment thereof of claim 1, comprising: a) a heavy chain variable region comprising SEQ ID NO: 25 and a light chain variable region comprising SEQ ID NO: 27; b) a heavy chain variable region comprising SEQ ID NO: 29 and a light chain variable region comprising SEQ ID NO: 31; c) a heavy chain variable region comprising SEQ ID NO: 33 and a light chain variable region comprising SEQ ID NO: 35; or d) a heavy chain variable region comprising SEQ ID NO: 37 and a light chain variable region comprising SEQ ID NO:

39.

3. The antibody of any one of the preceding claims, wherein the antibody is internalized into cells expressing huPGRN in vitro and / or in vivo.

4. The antibody of any one of the preceding claims, which competes with trastuzumab for binding to the hu PGRN receptor on a cell.

5. The antibody of claim 4, which is selected from the group consisting of antibodies 10C8, 16C11, 14A6 and 10B3.

6. The antibody of any one of the preceding claims which does not compete with trastuzumab for binding to the hu PGRN receptor on cells.

7. The antibody of claim 6, which is selected from the group consisting of antibodies 10C8, 16C11, 14A6 and 10B3.

8. A combination of antibodies according to claim 6 or 7.

9. The antibody of any one of the preceding claims which is an isolated monoclonal antibody.

10. The antibody of claim 9, wherein the monoclonal antibody is a human monoclonal antibody.

11. The antibody of any one of the preceding claims, wherein the antibody is derived from a human antibody, human IgG, human IgG1, human IgG2, human IgG2a, human IgG2b, human IgG3, human IgG4, human IgM, human IgA, human IgA1, human IgA2, human IgD, human IgE, a canine antibody, canine IgGA, canine IgGB, canine IgGC, canine IgGD, a chicken antibody, chicken IgA, chicken IgD, chicken IgE, chicken IgG, chicken IgM, chicken IgY, a goat antibody, goat IgG, a mouse antibody, mouse IgG, a pig antibody, a rat antibody, a llama antibody, an alpaca antibody, a shark antibody, and a camel antibody.

12. A derivative of the antibody according to any one of the preceding claims, optionally selected from F ab 、F ab2 , Fab' single-chain antibody, F v , single chain, monospecific antibody, bispecific antibody, trimeric antibody, multispecific antibody, multivalent antibody, chimeric antibody, canine-human chimeric antibody, canine-mouse chimeric antibody, antibody containing canine Fc, humanized antibody, human antibody, canine antibody, CDR-grafted antibody, shark antibody and nanobody.

13. A derivative of the antibody of any of the preceding claims, comprising a detectable label fixedly attached thereto, optionally wherein the detectable label is selected from the group consisting of fluorescein, DyLight, Cy3, Cy5, FITC, HiLyte Fluor 555, HiLyte Fluor 647, 5-carboxy-2,7-dichlorofluorescein, 5-carboxyfluorescein, 5-FAM, serotonin, 5-serotonin (5-HAT), 6-carboxyfluorescein (6-FAM), FITC, 6-carboxy-1,4-dichloro-2′,7′-dichlorofluorescein (TET), 6-carboxy-1,4-dichloro-2′,4′,5′,7′-tetrachlorofluorescein (HEX), 6-carboxy-4′,5′-dichloro-2′,7′-dimethoxyfluorescein (6-JOE), Alexa fluor, Alexa fluor 350, Alexa fluor 405, Alexa fluor 430, Alexa fluor488, Alexa fluor 500, Alexa fluor 514, Alexa fluor 532, Alexa fluor 546, Alexa fluor 555, Alexa fluor 568, Alexa fluor 594, Alexa fluor 610, Alexa fluor 633, Alexa fluor 635, Alexa fluor 647, Alexa fluor 660, Alexa fluor 680, Alexa fluor700, Alexa fluor 750, BODIPY fluorophore, BODIPY 492 / 515, BODIPY 493 / 503, BODIPY 500 / 510, BODIPY 505 / 515, BODIPY 530 / 550, BODIPY 542 / 563, BODIPY 558 / 568、BODIPY 564 / 570、BODIPY 576 / 589, BODIPY 581 / 591, BODIPY 630 / 650-X, BODIPY 650 / 665-X, BODIPY665 / 676, FL, FL ATP, FI-ceramide, R6G SE, TMR, TMR-X conjugate, TMR-X, SE, TR, TR ATP, TR-XSE, Rhodamine, Rhodamine 110, Rhodamine 123, Rhodamine B, Rhodamine B 200, Rhodamine BB, Rhodamine BG, Rhodamine Bextra, 5-carboxytetramethylrhodamine (5-TAMRA), 5GLD, 6-carboxyrhodamine 6G, Lissamine, Lissamine Rhodamine B, Phallicidine, Phalloidin, Rhodamine Red, Rhod-2,6-Carboxy-X-rhodamine (ROX), Carboxy-X-rhodamine (5-ROX), Sulforhodamine B can C, Sulforhodamine G Extra, 6-Carboxytetramethylrhodamine (TAMRA), Tetramethylrhodamine (TRITC), Rhodamine WT, Texas Red and Texas Red-X. , 14. The antibody of any of the preceding claims, comprising an effector moiety attached thereto, optionally wherein the effector moiety is selected from a cytotoxic drug, a toxin, diphtheria toxin A chain, exotoxin A chain, ricin A chain, abrin A chain, curcin, crotin, phenomycin, enomycin and a radiochemical.

15. The antibody of claim 14, further comprising a cleavable linker between the antibody and the effector moiety, wherein the cleavable linker releases the effector moiety into or within a cell.

16. An isolated polynucleotide encoding the antibody of any one of the preceding claims, optionally wherein the nucleic acid sequence is at least one of SEQ ID NOs: 57-64.

17. An expression vector comprising one or more polynucleotides of claim 16.

18. A host cell comprising the isolated polynucleotide of claim 16 and / or the expression vector of claim 17.

19. A composition comprising: at least one antibody or derivative of any one of claims 1-15; at least one isolated polynucleotide of claim 16; or at least one expression vector of claim 17; and / or, at least one host cell of claim 18; or a combination thereof; and a pharmaceutically acceptable carrier.

20. A method of detecting hu PGRN on a cell, the method comprising contacting a test biological sample with the antibody or derivative of any one of claims 1-15, and detecting the antibody that binds to the biological sample or a component thereof.

21. The method of claim 20, wherein the cell is selected from the group consisting of breast cancer, adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer, basal breast cancer, estrogen receptor (ER) luminal breast cancer, ER positive tamoxifen resistant luminal breast cancer, letrozole resistant luminal breast cancer, bile duct cancer, bladder cancer, brain cancer, glioblastoma, colorectal cancer, epidermoid carcinoma, squamous cell carcinoma, esophageal cancer, gastric cancer, hepatocellular carcinoma, kidney / renal cancer, laryngeal cancer, lung cancer, non-small cell lung cancer, lung adenocarcinoma, mesothelioma, myeloma / leukemia, ovarian cancer, prostate cancer, and uterine cancer cells.

22. The method of claim 20 or 21, further comprising comparing the amount of binding of the test biological sample or a component thereof with the amount of binding of a control biological sample or a component thereof, wherein an increase in binding of the test biological sample or a component thereof relative to the control biological sample or a component thereof indicates the presence of cells expressing hu PGRN in the test biological sample.

23. The method of any one of claims 20-22, wherein the test biological sample is a mammalian cell, tissue or blood.

24. The method of any one of claims 20-23, wherein the method is an in vivo method or an in vitro method.

25. A method for treating, preventing and / or ameliorating cancer in a mammal, comprising administering to the mammal at least one effective dose of a pharmaceutical composition comprising the antibody or derivative according to any one of claims 1 to 15.

26. The method of claim 25, wherein the cancer is selected from breast cancer, adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer, basal breast cancer, estrogen receptor (ER) luminal breast cancer, ER-positive tamoxifen-resistant Lumina 1 breast cancer, letrozole-resistant luminal breast cancer, bile duct cancer, bladder cancer, brain cancer, glioblastoma, colorectal cancer, epidermoid carcinoma, squamous cell carcinoma, esophageal cancer, gastric cancer, hepatocellular carcinoma, kidney / renal cancer, laryngeal cancer, lung cancer, non-small cell lung cancer, lung adenocarcinoma, mesothelioma, myeloma / leukemia, ovarian cancer, prostate cancer, and uterine cancer.

27. The method of claim 25 or 26, wherein the antibody comprises a cytotoxic effector moiety attached thereto, optionally wherein the effector moiety is selected from a cytotoxic drug, a toxin, diphtheria toxin A chain, exotoxin A chain, ricin A chain, abrin A chain, curcin, crotin, phenomycin, enomycin, and a radiochemical.

28. The method of any one of claims 25-27, wherein the antibody further comprises a cleavable linker between the antibody and the effector moiety, wherein the cleavable linker releases the effector moiety into or within the cell.

29. The method of any one of claims 25-28, wherein the antibody is administered as an antibody-drug conjugate.

30. The method of any one of claims 25-29, wherein multiple doses are administered to the animal; and / or, wherein the antibody is administered at a dose of about 1 to 50 mg / kg.

31. A kit for detecting hu PGRN expression in or on a cell, the kit comprising the antibody or derivative of any preceding claim, and instructions for use.

32. The kit of claim 31, wherein the cell is selected from the group consisting of breast cancer, adenocarcinoma epidermal growth factor receptor (EGF-R) breast cancer, basal breast cancer, estrogen receptor (ER) luminal breast cancer, ER positive tamoxifen resistant luminal breast cancer, letrozole resistant luminal breast cancer, bile duct cancer, bladder cancer, brain cancer, glioblastoma, colorectal cancer, epidermoid carcinoma, squamous cell carcinoma, esophageal cancer, gastric cancer, hepatocellular carcinoma, kidney / renal cancer, laryngeal cancer, lung cancer, non-small cell lung cancer, lung adenocarcinoma, mesothelioma, myeloma / leukemia, ovarian cancer, prostate cancer, and uterine cancer cells.

33. The kit of any one of claims 30 to 32, wherein the antibody or derivative is in lyophilized form.

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