Anti-HSP70 Antibodies and Therapeutic Uses Thereof

Abstract

The present invention generally relates to the fields of medicine, immunology, and cancer biology. More particularly, the present invention relates to antibodies targeting HSP70 and their uses. Provided herein are agents, such as antibodies targeting HSP70. Provided are methods of treating cancer, comprising administering an effective amount of an agent targeting HSP70, such as an HSP70-specific antibody, to a patient in need thereof. The HSP70-specific antibody can enhance the uptake of HSP70 by antigen-presenting cells. TIFF2024536133000064.tif144145

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of and U.S. priority to Provisional Patent Application No. 63 / 249,909, filed September 29, 2021, the disclosure of which is incorporated herein by reference in its entirety for all purposes.

[0002] Sequence Listing Reference This application contains an electronically submitted Sequence Listing XML, which is incorporated herein by reference in its entirety. The Sequence Listing SML, created on September 27, 2022, is named UTFCP1512WO_ST26.xml and is 371,316 bytes in size.

[0003] 1. Field The present invention relates generally to the fields of medicine, immunology and cancer biology. More specifically, the present invention relates to antibodies that target HSP70 and methods of use thereof. [Background technology]

[0004] 2. Description of Related Technical Fields The development of an immune response against cancerous cells is thought to depend on a series of reinforcing events, termed the cancer-immune cycle (Chen & Mellman, 2013), which begins with the release of cancer cell antigens during cancer cell death. Dendritic cells (DCs) are thought to be key early components of this response due to their ability to capture, process, and then present these tumor antigens to T cells via presentation via histocompatibility complex (MHC) class I and II molecules, which then leads to the priming and activation of effector CD4+ and CD8+ T cell responses. The important role of DCs is demonstrated, in part, by the many mechanisms utilized by tumors to suppress DC activity, including hypoxia, adenosine, lactic acid, low pH, and the expression of interleukin (IL)-10 and PD-L1, among others (Veglia & Gabrilovich, 2017).

[0005] Heat shock proteins (HSPs) in general, and HSP70 in particular, are thought to play a key role in this process due to their ability to link innate and adaptive immune responses (Shevtsov & Multhoff, 2016). For example, extracellular HSP70 binds and chaperones tumor antigens, then targets antigen-presenting cells, including DCs, through binding to different cell surface receptors, including CD91, oxidized low-density lipoprotein receptor 1 (OLR1), and scavenger receptor expressed by endothelial cells (SREC)-1, among others (McNulty et al., 2013), thereby delivering the bound antigen to DCs for processing. Furthermore, extracellular HSP70 secreted from tumor cells induces proinflammatory cytokines, such as interleukin (IL)-6 and tumor necrosis factor (TNF)-α, from macrophages (Vega et al., 2008), thereby enabling cross-presentation and T cell activation, respectively. Therefore, HSP70 is considered an attractive target for cancer therapy due to its important intracellular role as a cytoprotective antiapoptotic factor that promotes cancer cell survival in the face of various stressors, including both radiation and various chemotherapeutic agents (Boudesco et al., 2018). Furthermore, HSP70 is also considered an attractive target for cancer therapy due to its ability to stimulate immune responses not only through DCs but also possibly through macrophages, NK cells, and T cells (Shvetsov & Multhoff, 2016; Zininga et al., 2018).

[0006] Approaches that enhance DC uptake of HSP70-tumor antigen complexes hold promise for enhancing antitumor immunity and breaking tolerance. Several pharmacological inhibitors have been developed that directly target intracellular HSP70 or, in part, target its co-chaperones, which may act as sensitizers to radiation or chemotherapy (Boudesco et al., 2018). Furthermore, membrane-bound HSP70, while typically absent or found at low levels in normal cells, often shows enhanced expression on the surface of tumor cells and is associated with a more aggressive phenotype and poor prognosis in some malignancies (Boudesco et al., 2018; Chatterjee & Burns, 2017). Furthermore, HSP70 - / - Tumors have emerged as less immunogenic and more invasive (Dodd et al., 2015). This has led to the development and testing of various approaches that rely on cell surface expression of HSP70 for their activity, including ferromagnetic and gold nanoparticle-based therapies, vaccine strategies (Shvetsov & Multhoff, 2016), and monoclonal antibodies such as cmHSP70.1 (Stangl et al., 2011).

[0007] In recent years, immune checkpoint inhibitors (ICIs), including monoclonal antibodies against cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and programmed cell death 1 (PD-1) and its ligand PD-L1, have revolutionized immunotherapy through their ability to induce durable remissions even in advanced malignancies. Tumors that respond to ICIs are generally considered to tend to have a higher immune cell infiltration and / or interferon gene signature, or a higher mutational burden (TMB), and are sometimes referred to as "hot" tumors (Maleki Vareki, 2018). In contrast, so-called "cold" tumors with low immune cell infiltration or low TMB tend not to respond to ICIs, including pancreatic and prostate cancers (Maleki Vareki, 2018). Despite advances in treating various malignancies, there remains a need for new approaches that can convert cold tumors into more immunogenic tumors and provide alternative approaches for the treatment of these tumors. Summary of the Invention

[0008] overview The present invention is based, in part, on the discovery of anti-HSP70 antibodies (e.g., anti-HSP70 monoclonal antibodies) or antigen-binding fragments thereof. In certain circumstances, the anti-HSP70 monoclonal antibodies or antigen-binding fragments thereof can, for example, direct extracellular or soluble HSP70 bound to tumor-derived antigens to immune cells (e.g., dendritic cells), thereby treating cancer and / or enhancing the effectiveness of cancer therapy (e.g., cancer immunotherapy). In certain circumstances, the anti-HSP70 monoclonal antibodies or antigen-binding fragments thereof can, for example, form higher-order (i.e., more than one-to-one) complexes with HSP70.

[0009] The present invention provides antibodies (eg, isolated antibodies) that bind to human HSP70, eg, extracellular or soluble human HSP70.

[0010] In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3) comprising the amino acid sequence of SEQ ID NO:242 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-GA) comprising the amino acid sequence of SEQ ID NO:243 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-GAALIE) comprising the amino acid sequence of SEQ ID NO:244 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-YTE) comprising the amino acid sequence of SEQ ID NO:245 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-LS) comprising the amino acid sequence of SEQ ID NO:246 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-DF215) comprising the amino acid sequence of SEQ ID NO:247 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-DF228) comprising the amino acid sequence of SEQ ID NO:248 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249.

[0011] In certain aspects, the antibody has a K of 50 nM or less, 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM, or 0.05 nM or less as measured by surface plasmon resonance or biolayer interferometry. D It binds to human HSP70.

[0012] In some aspects, the antibody can form a higher-order antibody:HSP70 complex. The antibody:HSP70 complex can have, for example, a molecular weight of at least about 290 kDa, at least about 300 kDa, at least about 580 kDa, at least about 1,450 kDa, or at least about 1,750 kDa, or a molecular weight of about 290 kDa, about 580 kDa, about 1,450 kDa, or about 1,750 kDa. In some aspects, the antibody:HSP70 complex can have a molecular weight of at least about 300 kDa or more than about 300 kDa. In some aspects, the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:2. For example, the antibody:HSP70 complex may contain (i) about 1 antibody molecule and about 2 HSP70 molecules; (ii) about 2 antibody molecules and about 4 HSP70 molecules; (iii) about 5 antibody molecules and about 10 HSP70 molecules; or (iv) about 6 antibody molecules and about 12 HSP70 molecules.

[0013] In certain aspects, the antibody:HSP70 complex has a K of 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM, or 0.05 nM or less as measured by surface plasmon resonance or biolayer interferometry. D and binds to human FcγR (e.g., FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b) at 1000 ng / mL.

[0014] In some aspects, the antibody enhances the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells. Uptake can be mediated, for example, by human FcγR1, human FcγR2a, human FcγR2b, human FcγR2c, human FcγR3a, and / or human FcγR3b.

[0015] In some aspects, the antibody:HSP70 complex activates immune effector cells, such as CD8+ T cells, CD4+ T cells, NK cells, and dendritic cells, eg, immature dendritic cells.

[0016] In another embodiment, the invention provides antibodies (e.g., isolated antibodies) capable of forming higher order (i.e., more than one to one) antibody:HSP70 complexes. In some embodiments, the antibody:HSP70 complex further comprises an HSP70-related peptide.

[0017] The antibody:HSP70 complex may have, for example, a molecular weight of at least about 290 kDa, at least about 300 kDa, at least about 580 kDa, at least about 1,450 kDa, or at least about 1,750 kDa, or a molecular weight of about 290 kDa, about 580 kDa, about 1,450 kDa, or about 1,750 kDa. For example, the antibody:HSP70 complex may have a molecular weight of at least about 300 kDa or greater than about 300 kDa. In one aspect, the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:2. For example, the antibody:HSP70 complex may contain (i) about 1 antibody molecule and about 2 HSP70 molecules; (ii) about 2 antibody molecules and about 4 HSP70 molecules; (iii) about 5 antibody molecules and about 10 HSP70 molecules; or (iv) about 6 antibody molecules and about 12 HSP70 molecules.

[0018] In certain aspects, the antibody:HSP70 complex has a K of 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM, or 0.05 nM or less as measured by surface plasmon resonance or biolayer interferometry. D and binds to human FcγR (e.g., FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b) at 1000 ng / mL.

[0019] In some aspects, the antibody enhances the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells. Uptake can be mediated, for example, by human FcγR1, human FcγR2a, human FcγR2b, human FcγR2c, human FcγR3a, and / or human FcγR3b.

[0020] In one aspect, the antibody binds to an epitope of HSP70 comprising K573 to Q601 of SEQ ID NO: 11. In one aspect, the antibody binds to an epitope of HSP70 comprising one, two, three, four, five, six, seven, or eight of the following residues: K573, E576, W580, H594, K595, R596, E598, and Q601 of SEQ ID NO: 11. In one aspect, the antibody binds to an epitope of HSP70 comprising KEWHKREQ (SEQ ID NO:250).

[0021] In one aspect, the antibody comprises an immunoglobulin heavy chain variable region comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO:1, a VHCDR2 comprising the amino acid sequence of SEQ ID NO:2, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO:3, and an immunoglobulin light chain variable region comprising a VLCDR1 comprising the amino acid sequence of SEQ ID NO:4, a VLCDR2 comprising the amino acid sequence of SEQ ID NO:5, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO:6. In one aspect, the antibody comprises an immunoglobulin heavy chain variable region (hVH-1) comprising the amino acid sequence of SEQ ID NO:12, and an immunoglobulin light chain variable region (hVL-1) comprising the amino acid sequence of SEQ ID NO:19.

[0022] In some aspects, the antibody enhances the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells. Uptake can be mediated, for example, by human FcγR1, human FcγR2a, human FcγR2b, human FcγR2c, human FcγR3a, and / or human FcγR3b.

[0023] In another embodiment, the present invention provides higher order (ie, more than one to one) complexes comprising an antibody and HSP70.

[0024] The antibody:HSP70 complex may have, for example, a molecular weight of at least about 290 kDa, at least about 300 kDa, at least about 580 kDa, at least about 1,450 kDa, or at least about 1,750 kDa, or a molecular weight of about 290 kDa, about 580 kDa, about 1,450 kDa, or about 1,750 kDa. In some aspects, the antibody:HSP70 complex may have a molecular weight of at least about 300 kDa or greater than about 300 kDa. In some aspects, the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:2. For example, the antibody:HSP70 complex may contain (i) about 1 antibody molecule and about 2 HSP70 molecules; (ii) about 2 antibody molecules and about 4 HSP70 molecules; (iii) about 5 antibody molecules and about 10 HSP70 molecules; or (iv) about 6 antibody molecules and about 12 HSP70 molecules.

[0025] In certain aspects, the antibody:HSP70 complex has a K of 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM, or 0.05 nM or less as measured by surface plasmon resonance or biolayer interferometry. D and binds to human FcγR (e.g., FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b) at 1000 ng / mL.

[0026] In one aspect, the antibody binds to an epitope of HSP70 comprising K573 to Q601 of SEQ ID NO: 11. In one aspect, the antibody binds to an epitope of HSP70 comprising one, two, three, four, five, six, seven, or eight of the following residues: K573, E576, W580, H594, K595, R596, E598, and Q601 of SEQ ID NO: 11. In one aspect, the antibody binds to an epitope of HSP70 comprising KEWHKREQ (SEQ ID NO:250).

[0027] In one aspect, the antibody comprises an immunoglobulin heavy chain variable region comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO:1, a VHCDR2 comprising the amino acid sequence of SEQ ID NO:2, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO:3, and an immunoglobulin light chain variable region comprising a VLCDR1 comprising the amino acid sequence of SEQ ID NO:4, a VLCDR2 comprising the amino acid sequence of SEQ ID NO:5, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO:6. In one aspect, the antibody comprises an immunoglobulin heavy chain variable region (hVH-1) comprising the amino acid sequence of SEQ ID NO:12, and an immunoglobulin light chain variable region (hVL-1) comprising the amino acid sequence of SEQ ID NO:19.

[0028] In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3) comprising the amino acid sequence of SEQ ID NO:242 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-GA) comprising the amino acid sequence of SEQ ID NO:243 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-GAALIE) comprising the amino acid sequence of SEQ ID NO:244 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-YTE) comprising the amino acid sequence of SEQ ID NO:245 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-LS) comprising the amino acid sequence of SEQ ID NO:246 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-DF215) comprising the amino acid sequence of SEQ ID NO:247 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249. In one aspect, the antibody comprises an immunoglobulin heavy chain (hVH-1-G1m3-DF228) comprising the amino acid sequence of SEQ ID NO:248 and an immunoglobulin light chain variable region (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249.

[0029] In another aspect, the invention provides an isolated nucleic acid comprising a nucleotide sequence encoding the immunoglobulin heavy chain of any of the aforementioned antibodies and / or a nucleotide sequence encoding the immunoglobulin light chain of any one of the aforementioned antibodies. In another aspect, the invention provides an expression vector comprising any of the aforementioned nucleic acids. In another aspect, the invention provides a host cell comprising any of the aforementioned expression vectors.

[0030] In another aspect, the invention provides a pharmaceutical composition comprising any of the foregoing antibodies.

[0031] In another embodiment, the present invention provides a method for treating cancer in a subject in need thereof. The method comprises administering any of the above-mentioned antibodies or pharmaceutical compositions to the subject. In some embodiments, the method further comprises radiation therapy. In some embodiments, the radiation therapy is selected from gamma-ray irradiation, X-ray irradiation and radioisotope therapy. In some embodiments, the radiation therapy comprises X-ray radiation therapy.

[0032] In some aspects, the cancer is multiple myeloma, breast cancer, melanoma, colon cancer, pancreatic cancer, or prostate cancer. In some aspects, the subject (or a sample from the subject) has a serum HSP70 level greater than 20 ng / mL.

[0033] In another aspect, the present invention provides a method for enhancing the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells, comprising contacting the cells with any of the aforementioned antibodies or pharmaceutical compositions.

[0034] In another embodiment, provided herein is the antibody molecule, pharmaceutical composition, cell, or pharmaceutical composition of any one of the present embodiments for use in treating cancer in a subject.

[0035] In another embodiment, provided herein is the use of an antibody molecule, pharmaceutical composition, cell, or pharmaceutical composition of any one of the present embodiments in the manufacture of a medicament for treating cancer in a subject.

[0036] Other objects, features, and advantages of the present invention will become apparent from the following drawings and detailed description. It should be understood, however, that the detailed description and specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. [Brief explanation of the drawings]

[0037] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein.

[0038] [Figure 1] BALB / c mice injected with MOPC315.BM-luc cells received 200 μg injections of the indicated HSP70 mAbs (squares) or their IgG2B isotype controls (circles) twice weekly during weeks 1 to 3. Disease burden was monitored using whole-animal in vivo imaging and confirmed by serum light chain levels. [Figure 2A] Octet analysis showing the affinity of 77A for mouse HSP70 (top panel), human HSP70 produced in Escherichia coli (E. coli) (middle panel), and human HSP70 produced in Sf9 insect cells (bottom panel) (Figure 2A). [Figure 2B] 77A shows preferential binding to the ADP-HSP70 complex (Fig. 2B). [Figure 2C]Binding of 77A to HSP70-GFP showed the greatest affinity in the presence of ADP and the peptide substrate (NRL) (Figure 2C). Within each group of bars in Figure 2C, the bars represent, from left to right, buffer, ATP, ADP, ATP NRL, and ADP NRL. [Figure 3-1] Figure 3A-F. Full-length (FL) HSP70 was expressed as an N-terminal GFP fusion protein in HSP70 KO 293T cells, along with the indicated deletion mutants, in which progressively more C-terminal amino acids were removed (Figure 3A). After IP of cell extracts with 77A, the proteins were detected by Western blotting (WB) with an anti-GFP antibody (Figure 3B). Smaller deletions were then created for finer mapping (Figure 3C), and the indicated IPs were performed from cell lysates (CL) or culture medium supernatants (CM). Loading was confirmed using an alpha light chain (LC) antibody. The putative binding domain of 77A is shown in a molecular model of HSP70, representing the relevant region of the protein (Figure 3D; the amino acid sequence shown corresponds to positions 595-614 of SEQ ID NO:11). Alanine scanning analysis was used to determine the exact amino acids that comprise the 77A epitope, and the results are shown in (Figure 3E), with a ribbon diagram depicting the primary and secondary critical sites shown in (Figure 3F). [Figure 3-2] See description of Figure 3-1. [Figure 3-3] See description of Figure 3-1. [Figure 4] Figure 4A-B. Luc-labeled MM1.S human myeloma cells were injected into nude mice and treated twice weekly for weeks 2-5 with either an IgG2B isotype control mAb or 77A at the indicated doses. Whole-animal live imaging data are shown at week 5 (Figure 4A). Dorsal (upper panel) and ventral (lower panel) views demonstrate significant tumor growth in IgG2B-treated mice, but not in 77A-treated mice, especially at higher dose levels. The same experiment was then performed in NSG mice, and tumor growth was measured both by imaging (Figure 4B) and by ELISA against the human light chain. [Figure 5] Immature mouse DC2.4 cells were incubated with either vehicle (left two panels) or 6x-His-tagged HSP70 (right two panels) in the presence of IgG2B or 77A as indicated for 6 hours at 37°C. Immature mouse DC2.4 cells were then stained with either control IgG (left panel) or α-6x-His-tagged mAb (right three panels), and HSP70 uptake was determined by flow cytometry. Significant uptake of HSP70 is only seen in the presence of 77A (rightmost panel). [Figure 6] DC2.4 cells exposed to Sf9-derived 6x-His-tagged human HSP70 in the presence of either an isotype control mAb or 77A mAb were stained with either wheat germ agglutinin (WGA) conjugated to Alexa Fluor 594 to stain the cell membrane, α-6x-His mAb to detect HSP70, or 4',6-diamidino-2-phenylindole (DAPI) to stain the nucleus. Individual and merged images were obtained. Representative fields are shown at 200x magnification. [Figure 7] Electron micrographs of DCs exposed to HSP70 and gold-labeled 77A. Magnification shown is 100,000x. [Figure 8A]Figures 8A-C. Murine DC2.4 cells were treated with either PBS or 5 μg of ADP-HSP70 purified from A-375 melanoma cells, which express high levels of HSP70 and are therefore an excellent source of HSP70, in combination with 10 μg of IgG2B or 77A for 48 hours. RNA was harvested, and cDNA was hybridized to qPCR arrays as described in the text. Genes activated or repressed by more than two-fold are shown for the comparison of IgG2B to PBS (Figure 8A; upper panel) and 77A to PBS (Figure 8A; lower panel). Data from three biological replicates are shown. Ingenuity Pathway Analysis was then performed to identify biological processes that may be affected by these changes (Figure 8B). Cytokines released by mature DCs were then analyzed using a BioRad Bio-Plex™ Pro Mouse Cytokine Array. The significant changes induced in cells exposed to HSP70 and 77A compared to cells exposed to HSP70 and IgG2B are shown in a bar graph (Figure 8C). Within each group of bars in (Figure 8C), the left bar represents the IgG control (CTRL), and the right bar represents HSP70 stimulation. [Figure 8B] See legend to Figure 8A. [Figure 8C] See legend to Figure 8A. [Figure 9-1]Figures 9A-G. BALB / c mice were injected with 7,500 luc-4T1 cells into the fourth right mammary gland. After 12 days, when all mice had palpable and measurable tumors, they were intravenously injected with 200 μg of either IgG2B (filled squares) or 77A (open squares) twice weekly for 3 weeks. Tumor volume was measured using both calipers and whole-animal imaging for primary tumors (Figure 9A), whereas imaging was used to assess lung metastases (Figure 9B). Peripheral blood was collected on day 32 and assessed for CD4+ and CD8+ T cells (Figure 9C) and, in duplicate, for CD11c+ / MHC class II+ cells and total MHC class II+ cells (Figure 9D). 77A induces HSP70 uptake into human primary CD4+ and CD8+ T cells (Figure 9E). 77A stimulates MHC-independent cytolytic CD4 T cell activity (FIG. 9F). 77A activity against the A375 melanoma model in nude mice (FIG. 9G). [Figure 9-2] See description of Figure 9-1. [Figure 9-3] See description of Figure 9-1. [Figure 9-4] See description of Figure 9-1. [Figure 10] HSP70 bound to ATP at the nucleotide-binding domain (NBD) has an open conformation to allow interaction with the substrate-binding domain (SBD). Substrate peptide interaction with the SBD, in concert with J proteins and nucleotide exchange factors (NEFs), stimulates HSP70 ATPase activity and leads to lid closure, thereby stabilizing HSP70 interaction with the substrate. (Source: Craig & Marszalek, 2017). The approximate location of the 77A bond on the HSP70 model is also shown diagrammatically in the right panel. [Figure 11]Figures 11A-C. Homogenates from a 10 mL pellet of 4T1 cells expressing HSP70-GFP were purified on an ADP-agarose column to isolate ADP-HSP70-peptide complexes, which were then injected intraperitoneally (10 μg) into BALB / c mice on day -24 and boosted subcutaneously on day -10. These mice were then injected with 4T1-luc cells expressing HSP70-GFP on day 0 as described in the legends of Figures 9A-D, and tumor growth was monitored by whole-animal imaging (Figure 11A). On day 37, animals were euthanized, at which time spleen cells were isolated and analyzed by fluorescence-activated cell sorting (FACS) that day or cultured in the presence of irradiated 4T1 cells expressing HSP70-GFP for 7 days and then analyzed by FACS. Comparisons are provided for CD4+ (Figure 11B) and CD8+ (Figure 11C) T cell content at the time of spleen isolation (day 0) and after 7 days of culture (left panel). Cytotoxic T cell activity was also tested by performing viability studies after exposing the indicated cell fractions to live wild-type 4T1 cells or 4T1 cells expressing HSP70-GFP (right panel). [Figure 12] Figure 12A-B. Cytokine release assays were performed on CD4+ (Figure 12A) and CD8+ (Figure 12B) T cells isolated from mouse spleens after exposure to either 4T1 cells or 4T1 cells expressing HSP70-GFP, as detailed above. IFNγ secretion is shown in the upper panel, and IL-2 secretion is shown in the lower panel. [Figure 13] 77A induces HSP70 uptake via FcγR2A and FcγR2B. The top row represents HSP70 knockout HEK293T cells expressing human FcγR2A. The bottom row represents HSP70 knockout HEK293T cells expressing the indicated human Fcγ receptors. [Figure 14]Figures 14A-B. The sequences of mouse HSP70 (SEQ ID NO:251; positions 594-614) and human HSP70 (SEQ ID NO:11; positions 594-614) at the proposed binding site for 77A are highlighted (Figure 14A). Amino acid differences are boxed. A potential phosphorylation site in mouse HSP70 is at K595; potential phosphorylation sites in human HSP70 are at K595 and K597. Potential ubiquitination sites in mouse HSP70 are at S604, S608, and Y611; potential ubiquitination sites in human HSP70 are at S608 and Y611. Mutation of three amino acids in human HSP70 to mimic the mouse version reduces the ability of 77A to recognize human HSP70 (Figure 14B). [Figure 15] Figure 15A-B. Tumor targeting of 77A. PLD plus IgG2B or 77A was evaluated in BALB / c mice orthotopically injected with 4T1 cells (A) and in a CT26-based immunocompetent colon cancer model (B). [Figure 16] Antibody 77A was tested in an epitope binning experiment for competition for binding to HSP70 protein with the indicated antibodies. The numbers in Figure 16 reflect the percent of maximal binding in the presence of potentially competing antibodies. [Figure 17] Binding of antibody 77A to ADP-HSP70 (top) and ATP-HSP70 (bottom) measured by biolayer interferometry (BLI). [Figure 18] Binding of antibody 77A to ADP-HSP70 and ATP-HSP70 measured by ELISA, where No. 1° and No. 2° represent negative controls in which no primary or secondary antibody, respectively, was present. [Figure 19] Tumor volume after treatment of CT-26 tumor-bearing mice with the indicated antibodies. Boxed days (14, 17, 21) indicate the days on which mice received treatment. *p=0.0023 versus isotype control (IgG2B) as determined by Dunnett's multiple comparison t-test. [Figure 20]Figures 20A-B. Sequence alignment of humanized variants hVH-1 to hVH-5 (Figure 20A) and hVL-1 to hVL-5 (Figure 20B). [Figure 21] HSP70 uptake after incubation of cells expressing the indicated human Fcγ receptors with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated antibodies, as measured by total MFI (left) and % GFP-positive cells (right). HC1 LC12001=h77A-1; HC2 LC1=h77A-6; HC3 LC1=h77A-11. [Figure 22] HSP70 uptake after incubation of cells expressing the indicated human Fcγ receptors with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated IgG2 antibodies, as measured by total MFI (left) and % GFP-positive cells (right). HC1 LC1=h77A; HC1 LC1=h77A-1; HC2 LC1=h77A-6; HC3 LC1=h77A-11. [Figure 23] HSP70 uptake after incubation of cells expressing the indicated mouse Fcγ receptors with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated antibodies, as measured by total MFI (left) and % GFP-positive cells (right). HC1 LC1=h77A; HC1 LC1=h77A-1; HC2 LC1=h77A-6; HC3 LC1=h77A-11. [Figure 24] HSP70 uptake after incubation of cells expressing the indicated mouse Fcγ receptors with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated IgG2 antibodies, as measured by total MFI (left) and % GFP-positive cells (right). HC1 LC1=h77A; HC1 LC1=h77A-1; HC2 LC1=h77A-6; HC3 LC1=h77A-11. [Figure 25]HSP70 uptake after incubation of DCs with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated antibodies, as measured by MFI (left) and % GFP-positive cells (right). Results are shown for all cells gated on HSP70GFP. HC1 LC1 = h77A; HC2 LC1 = h77A-6; HC3 LC1 = h77A-11. [Figure 26] HSP70 uptake after incubation of DCs with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated antibodies, as measured by % GFP-positive cells (left) and MFI (right). Results are shown for plasmacytoid DCs, CD303+ve, CD1C-ve cells: HC1 LC1 = h77A; HC1 LC1 = h77A-1; HC2 LC1 = h77A-6; HC3 LC1 = h77A-11. [Figure 27] HSP70 uptake after incubation of DCs with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated antibodies, as measured by % GFP-positive cells (left) and MFI (right). Results are shown for type 1 DCs, CD141+ve, CD1c-ve cells. HC1 LC1=h77A; HC1 LC1=h77A-1; HC2 LC1=h77A-6; HC3 LC1=h77A-11. [Figure 28] HSP70 uptake after incubation of DCs with HSP70GFP, GFP-Nanobody Alexa-488, and the indicated antibodies, as measured by % GFP-positive cells (left) and MFI (right). Results are shown for type 2 DCs, CD1C+ve, CD303-ve cells: HC1 LC1=h77A; HC1 LC1=h77A-1; HC2 LC1=h77A-6; HC3 LC1=h77A-11. [Figure 29A] Figures 29A-J show sequence alignments of humanized variants hVH-1.1 to hVH-1.78 (Figures 29A-F) and hVL-1.1 to hVL-1.53 ​​(Figures 29G-J). [Figure 29B]See legend to Figure 29A. [Figure 29C] See legend to Figure 29A. [Figure 29D] See legend to Figure 29A. [Figure 29E] See legend to Figure 29A. [Figure 29F] See legend to Figure 29A. [Figure 29G] See legend to Figure 29A. [Figure 29H] See legend to Figure 29A. [Figure 29I] See legend to Figure 29A. [Figure 29J] See legend to Figure 29A. [Figure 30] Size-exclusion chromatography (SEC) traces for h77A-1-G1m3 labeled with CF647 dye ("G1m3-CF647") and HSP70 fused to GFP ("HSP70-GFP"), each run alone and detected using the corresponding fluorescence channel. Peaks, corresponding retention times, and calculated approximate molecular weights (*MW) are shown. The molecular weight of each peak was calculated from a MW standard curve. [Figure 31] Size-exclusion chromatography (SEC) trace of h77A-1-G1m3 ("G1m3") labeled with CF647 dye and HSP70 fused to GFP ("HSP70-GFP") after co-incubation of antibody and HSP70. The same sample was run twice, first to detect fluorescence from GFP (left) and then again to detect fluorescence from CF647 (right). Peaks, corresponding retention times, and calculated approximate molecular weights (*MW) are shown. The molecular weight of each peak was calculated from a MW standard curve. The results demonstrate colocalization of antibody and HSP70 in the detected high molecular weight peaks. [Figure 32]Size-exclusion chromatography (SEC) trace of h77A-1-G1m3 labeled with CF647 dye ("G1m3-CF647") and HSP70 fused to GFP ("HSP70-GFP") after co-incubation at the indicated molar ratios. Results are shown for detection of fluorescence from CF647. Peaks, corresponding retention times, and calculated approximate molecular weights (*MW) are indicated. The molecular weight of each peak was calculated from a MW standard curve. [Figure 33] Size-exclusion chromatography (SEC) trace of h77A-1-G1m3 labeled with CF647 dye ("G1m3-CF647") and HSP70 fused to GFP ("HSP70-GFP") after co-incubation at the indicated molar ratios. Results are shown for detection of fluorescence from GFP. Peaks, corresponding retention times, and calculated approximate molecular weights (*MW) are indicated. The molecular weight of each peak was calculated from a MW standard curve. [Figure 34] Predicted composition of the high molecular weight complex containing h77A-1-G1m3 and HSP70. [Figure 35A] Figures 35A-C. Binding of mouse 77A antibody in mouse IgG1 ("mIgG1"; Figure 35A), mouse IgG2a ("mIgG2a"; Figure 35B), and mouse IgG2b ("mIgG2b"; Figure 35C) formats to mouse FcγR3 alone or in complex with human HSP70 ("huHSP70") as measured by ELISA. EC50 is shown. [Figure 35B] See legend to Figure 35A. [Figure 35C] See legend to Figure 35A. [Figure 36] Binding of humanized h77A-1-G1m3 antibody ("G1m(3)") to human FcγR2A alone or in complex with HSP70 as measured by ELISA. EC50 is shown. [Figure 37A]Figures 37A-C. Binding of the murine 77A antibody to murine FcγR3 (Figure 37A) and the humanized h77A-1-G1m3 antibody to human FcγR2a (Figure 37B), each alone ("antibody only") or in complex with HSP70 ("complex"), as measured by biolayer interferometry (BLI). Kinetics of h77A-1-G1m3 binding to human FcγR2a, alone ("G1m(3)") or in complex with HSP70 ("G1m(3) complex"), as measured by biolayer interferometry (BLI) (Figure 37C). [Figure 37B] See legend to Figure 37A. [Figure 37C] See legend to Figure 37A. [Figure 38] Binding of commercially available CM710.1 and N15F2-5 antibodies, each alone (top trace) or in complex with HSP70 (bottom trace), to mouse FcγR3 as measured by biolayer interferometry (BLI). [Figure 39A] Size-exclusion chromatography (SEC) traces after co-incubation of antibody and HSP70: (i) N15F2-5 ("N15") or mouse 77A in mouse IgG1 format ("mou IgG1"), each labeled with CF647 dye, and (ii) HSP70 fused to GFP ("HSP70-GFP"). The same sample was run twice, first to detect fluorescence from GFP (left) and then again from CF647 (right). Peaks, corresponding retention times, and calculated approximate molecular weights (*MW) are shown. The molecular weight of each peak was calculated from a MW standard curve. [Figure 39B]Size-exclusion chromatography (SEC) traces after co-incubation of antibody and HSP70: (i) CM710.1 or mouse 77A in mouse IgG1 format ("mou IgG1"), each labeled with CF647 dye, and (ii) HSP70 fused to GFP ("HSP70-GFP"). The same sample was run twice, first to detect fluorescence from GFP (left) and then again from CF647 (right). Peaks, corresponding retention times, and calculated approximate molecular weights (*MW) are shown. The molecular weight of each peak was calculated from a MW standard curve. [Figure 40] Uptake of antibody:HSP70-GFP complexes in 293 cells transfected with human FcγR2A as measured by FACS. Antibodies tested were humanized h77A-1 in human IgG2 format ("HC1-LC1") and mouse 77A in mouse IgG2b LALAPG format ("LALAPG"; reduced Fc receptor binding variant). HEL: isotype control antibody. y-axis: percent GFP-positive cells; x-axis: antibody concentration. The EC50 for humanized h77A-1 in human IgG2 format ("HC1-LC1") is shown. [Figure 41] Uptake of antibody:HSP70-GFP complexes in monocytic THP-1 cells as measured by FACS. Antibodies tested were humanized h77A-1 in human IgG2 format ("HC1-LC1") and humanized h77A-1 in IgG1 G1m3 format ("G1m(3)"). HEL: isotype control antibody. y-axis: percent GFP-positive cells, x-axis: antibody concentration. [Figure 42]Uptake of antibody:HSP70-GFP complexes in monocytic THP-1 cells as measured by FACS. Antibodies tested were h77A-1 in human IgG1 G1m3-GA format ("GA"), h77A-1 in human IgG1 G1m3-GAALIE format ("GAALIE"), h77A-1 in human IgG2 format ("HC1-LC1"), h77A-1 in human IgG1 G1m3 format ("G1m(3)"), and mouse 77A in mouse IgG2bLALAPG format ("LALAPG"). HEL: isotype control antibody. y-axis: percent GFP-positive cells; x-axis: antibody concentration. [Figure 43] Uptake of antibody:HSP70-GFP complexes in mouse RAW264.7 cells as measured by FACS. Antibodies tested were mouse IgG1 ("mIgG1"), IgG2a ("mIgG2a"), IgG2b ("mIgG2b"), and mouse 77A in IgG2bLALAPG ("LALAPG") format. HEL: isotype control antibody. y-axis: percent live GFP-positive cells, x-axis: antibody concentration. [Figure 44] Figures 44A-B. Uptake of antibody:HSP70-GFP complexes in mouse DC3.2 (Figure 44A) and DC2.4 (Figure 44B) cells as measured by FACS. Antibodies tested were mouse IgG1 ("mIgG1"), IgG2a ("mIgG2a"), IgG2B ("mIgG2b"), and mouse 77A in IgG2bLALAPG ("LALAPG") format. HEL: isotype control antibody. Y-axis: percent live GFP-positive cells, x-axis: antibody concentration. [Figure 45] Uptake of antibody:HSP70-GFP complexes in primary mouse monocytes derived from bone marrow cells as measured by FACS. Antibodies tested were mouse IgG1 ("mIgG1"), IgG2a ("mIgG2a"), IgG2B ("mIgG2b"), and mouse 77A in IgG2bLALAPG ("LALAPG") format. HEL: isotype control antibody. Y-axis: percent live GFP-positive cells, x-axis: antibody concentration. [Figure 46A]Figures 46A-C. Uptake of antibody:HSP70-GFP complexes in human macrophage cells (Figure 46A), monocyte cells (Figure 46B), and macrophage cells (Figure 46C) as measured by FACS. Antibodies tested were human IgG1 G1m3 ("G1m(3)"), IgG1 G1m3-GA ("GA"), and h77A-1 in IgG1 G1m3-GAALIE format. HEL: isotype control antibody. Y-axis: percent live GFP-positive cells, x-axis: antibody concentration. [Figure 46B] See legend to Figure 46A. [Figure 46C] See legend to Figure 46A. [Figure 47] Uptake of antibody:HSP70-GFP complexes in mouse DC3.2 dendritic cells as measured by FACS. Antibodies tested were N15F2-5, CM170.1, and mouse 77A ("253-77A") in mouse IgG1, IgG2a, and IgG2b formats. HEL: isotype control antibody. X-axis: Alexa Flour 488-A to detect GFP, y-axis: FSC-A to detect live cells. The percentage of live GFP-positive cells is shown. [Figure 48] Uptake of antibody:HSP70-GFP complexes in mouse MH-S macrophage cells measured by FACS. Antibodies tested were N15F2-5, CM170.1, and mouse 77A ("253-77A") in mouse IgG1, IgG2a, and IgG2b formats. HEL: isotype control antibody. X-axis: Alexa Flour 488-A to detect GFP; y-axis: FSC-A to detect live cells. The percentage of live GFP-positive cells is shown. [Figure 49A] Figures 49A-C. Binding of antibodies to human FcγR2A. Engineered Fc variants were assessed in a dose response titration for binding to FcγR2A by monomeric antibody or by antibody already complexed to antigen. [Figure 49B] See legend to Figure 49A. [Figure 49C] See legend to Figure 49A. [Figure 50] Uptake of HSP70-GFP complexed with engineered Fc variants by primary human monocytes. The number of cells positive for GFP was gated on and the percentage of positive cells in the live population was plotted. [Figure 51] Uptake of HSP70-GFP complexed with engineered Fc variants by primary human macrophages. The number of cells positive for GFP was gated on and the percentage of positive cells in the live population was plotted. [Figure 52] Uptake of HSP70-GFP complexed with engineered Fc variants by primary human dendritic cells. The number of cells positive for GFP was gated on and the percentage of positive cells in the live population was plotted. [Figure 53] Proliferation of CD8+ T cells determined by FACS after incubation with 77A complex. [Figure 54] FACS analysis of CD25 and HLA_DR activation markers on CD8+ T cells after incubation with the 77A complex. The complex formed between the 77A Fc variant and HSP70 induces the expression of phenotypic markers of activation in CD8+ T cells after co-culture. [Figure 55] FACS analysis of immature dendritic cells for activation markers. The complex of HSP70 formed with the 77A Fc variant induces the expression of CD83, a phenotypic marker of activation, in immature dendritic cells after incubation with the 77A complex. [Figure 56] Female C57BL / 6 mice transgenic for human FcγR were inoculated with the EG 7-luc tumor line. Tumor growth was monitored by imaging for luciferase signals. HSP70-OVA fusion protein was co-incubated with control isotype (IsoPAL), wild-type 77A IgG1 (77AWT), or engineered Fc variants (77AGA, 77AGAALIE) to form complexes. Mice were then immunized and boosted with the protein / complex. [Figure 57]Survival curves of female C57BL / 6 FcγR transgenic (Tg) mice by treatment group. [Figure 58] Female C57BL / 6 mice transgenic for human FcγR were inoculated with the E0771 tumor line. Tumor growth was monitored before and during treatment with either the wild-type humanized 77A IgG1 antibody or the engineered Fc variants. Tumor growth was compared to an isotype human IgG1 control during treatment. The "On Tx" field indicates the duration of treatment. [Figure 59] Survival curves of female C57BL / 6 FcγR Tg mice by treatment group. The "On Tx" region indicates the duration of treatment. [Figure 60] Male C57BL / 6 mice transgenic for human FcγR were inoculated with the PANO2-CRE2 tumor line. Tumor growth (volume) was measured using a caliper. Mice were then preconditioned with 10 Gy of direct tumor X-ray irradiation before treatment initiation. The following day, mice were treated with either a control isotype or the engineered Fc variant 77A antibody. DETAILED DESCRIPTION OF THE INVENTION

[0039] Detailed Description The present invention is based, in part, on the development of anti-HSP70 antibodies or antigen-binding fragments thereof that are useful in the treatment of certain indications, such as cancer. In certain circumstances, anti-HSP70 antibodies (e.g., anti-HSP70 monoclonal antibodies) or antibody fragments thereof can, for example, direct extracellular or soluble HSP70 bound to tumor-derived antigens to immune cells (e.g., dendritic cells), thereby treating cancer and / or enhancing the effectiveness of cancer therapy (e.g., cancer immunotherapy). In certain circumstances, anti-HSP70 antibodies or antibody fragments thereof can, for example, form higher-order (i.e., more than one-to-one) complexes with HSP70.

[0040] The data provided herein demonstrate that an anti-HSP70 antibody (mAb; designated clone 77A) exhibits activity independent of surface HSP70 expression and targets extracellular HSP70 associated with tumor-derived antigens to DCs. This antibody can be used to better understand the role of HSP70 in immunity and as a therapeutic agent to enhance the efficacy of cancer immunotherapy. In particular, clone 77A is a high-affinity HSP70 mAb that exhibits antitumor activity in both hematological malignancy and solid tumor models in immunocompetent and nude mice, but not in immunodeficient mice with a spontaneous protein kinase DNA-activated catalytic subunit (PRKDCSCID) mutation, also known as SCID mice. This antibody enhances the internalization of HSP70 by DCs in in vitro assays, leading to the upregulation of genes associated with DC maturation. When tested against orthotopically implanted 4T1 cells, an immunologically cold model of mouse triple-negative breast cancer that is unresponsive to ICI, 77A reduced primary tumor growth and inhibited the development of lung and liver metastases. In combination with pegylated liposomal doxorubicin (PLD), an agent that induces immunogenic cell death (ICD) and enhances the release of HSP70-tumor peptide complexes, 77A cured some mice in both 4T1 and colorectal cancer models. Finally, when purified ADP-HSP70 complexes from 4T1 cells were used as a vaccine with 77A, tumor growth following subsequent challenge with live 4T1 cells was inhibited compared with the mAb isotype control, and the abundance of 4T1-specific cytolytic CD4+ and CD8+ T cell activity was enhanced. Thus, enhancing HSP70 uptake by immune cells using the clonal 77A mAb enhances antitumor immunity, both alone and in several rationally designed combination regimens.

[0041] The data provided herein also show that the 77A antibody and its variants form high-molecular-weight immune complexes upon binding to HSP70. These high-molecular-weight immune complexes bind to Fc receptors more strongly than the antibody alone and exhibit significantly enhanced cellular uptake of HSP70 compared to the antibody alone. Because other commercially available anti-HSP70 antibodies do not appear to form such high-molecular-weight immune complexes, the ability to form high-molecular-weight immune complexes is not simply a function of HSP70 binding. Therefore, antibodies (e.g., 77A and its variants) that can form high-molecular-weight immune complexes upon binding to HSP70 are believed to be particularly useful in the treatment of certain indications, such as cancer.

[0042] I. Definition As used herein, "nucleic acid," "nucleic acid sequence," "oligonucleotide," "polynucleotide," or other grammatical equivalents refer to at least two nucleotides, either deoxyribonucleotides or ribonucleotides, covalently linked to each other, or their analogs. Polynucleotides are polymers of any length, including, for example, 20, 50, 100, 200, 300, 500, 1000, 2000, 3000, 5000, 7000, 10,000, etc. Polynucleotides described herein generally contain phosphodiester bonds, but in some cases include nucleic acid analogs that may have at least one different bond, such as a phosphoramidate, phosphorothioate, phosphorodithioate, or O-methylphosphoramidite bond, as well as peptide nucleic acid backbones and linkages. Mixtures of naturally occurring polynucleotides and analogs can be made, or mixtures of different polynucleotide analogs and naturally occurring polynucleotides and analogs can be made. The following are non-limiting examples of polynucleotides: genes or gene fragments, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, ribozymes, cDNA, cRNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, and primers. Polynucleotides may contain modified nucleotides, such as methylated nucleotides and nucleotide analogs. If present, modifications to the nucleotide structure may be imparted before or after assembly of the polymer. The sequence of nucleotides may be interrupted by non-nucleotide components. Polynucleotides may be further modified after polymerization, such as by conjugation with a labeling component. The term also includes both double-stranded and single-stranded molecules. Unless otherwise specified or required, the term polynucleotide encompasses both the double-stranded form and each of the two complementary single-stranded forms known or predicted to constitute the double-stranded form.When a polynucleotide is RNA, it is composed of a specific sequence of four nucleotide bases: adenine (A), cytosine (C), guanine (G), thymine (T), and uracil (U) instead of thymine. Thus, the term "polynucleotide sequence" is the alphabetical representation of a polynucleotide molecule. Unless otherwise indicated, a particular polynucleotide sequence implicitly encompasses not only the sequence explicitly indicated, but also its conservatively modified variants (e.g., degenerate codon substitutions) and complementary sequences. Specifically, 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.

[0043] As used herein, the terms "peptide," "polypeptide," and "protein" refer to a polymer of amino acid residues. These terms also apply to naturally occurring amino acid polymers, amino acid polymers containing modified residues, and non-naturally occurring amino acid polymers, as well as amino acid polymers in which one or more amino acid residues are artificial chemical mimetics of corresponding naturally occurring amino acids. In this instance, the term "polypeptide" encompasses antibodies or fragments thereof.

[0044] As used herein, other terms used in the fields of recombinant nucleic acid technology, microbiology, immunology, antibody engineering and molecular and cell biology will be commonly understood by those skilled in the art.

[0045] As used herein, the terms "a" and "an" mean "one or more" and include plurals unless the context is inappropriate.

[0046] Although the use of the term "or" in the claims is used to mean "and / or" unless expressly indicated to refer to alternatives only or the alternatives are not mutually exclusive, the present disclosure supports a definition that refers to alternatives only and "and / or." As used herein, "another" can mean at least a second or more.

[0047] As used herein, the terms "subject" and "patient" are used interchangeably and refer to an organism treated by the methods and compositions described herein. Such organisms preferably include, but are not limited to, mammals (e.g., murines, simians, equines, bovines, porcines, canines, felines, etc.), and more preferably include humans.

[0048] As used herein, the term "effective amount" refers to an amount of a compound (e.g., a compound of the present disclosure) sufficient to produce a beneficial or desired result. An effective amount can be administered in one or more administrations, applications, or dosages, and is not intended to be limited to a particular formulation or route of administration.

[0049] The terms "treatment" and "treating" refer to the administration or application of a therapeutic agent or the performance of a procedure or modality on a subject for the purpose of obtaining a therapeutic benefit for a disease or health-related condition. These terms also include any effect, e.g., alleviation, reduction, modulation, improvement, or elimination, that results in the improvement of a condition, disease, disorder, etc., or the amelioration of their symptoms. For example, treatment can include the administration of a pharmaceutically effective amount of an antibody targeting HSP70, alone or in combination with the administration of chemotherapy, immunotherapy, or radiation therapy, the performance of surgery, or any combination thereof.

[0050] The term "therapeutic benefit" or "therapeutically effective" refers to anything that promotes or enhances the well-being of a subject with respect to the medical treatment of this condition. This includes, but is not limited to, reducing the frequency or severity of signs or symptoms of the disease. For example, treating cancer can include, for example, reducing tumor size, reducing tumor invasiveness, reducing the rate of cancer growth, or preventing metastasis. Treating cancer can also refer to extending the survival of a subject with cancer.

[0051] As used herein, the term "pharmaceutical composition" refers to a combination of an active agent and a pharmaceutically acceptable carrier (inert or active) that makes the composition particularly suitable for diagnostic or therapeutic use in vivo or ex vivo.

[0052] As used herein, the term "pharmaceutically acceptable carrier" refers to any standard pharmaceutical carrier, such as phosphate buffered saline solution, water, emulsions (e.g., oil / water or water / oil emulsions), and various types of wetting agents. The composition may also contain stabilizers and preservatives. For examples of carriers, stabilizers, and adjuvants, see, for example, Adeboye Adejare, Remington: The Science and Practice of Pharmacy (23rd ed. 2020).

[0053] Throughout this application, the term "about" is used to indicate that a value includes the inherent variation of error for the device, the method being used to determine the value, the variation that exists between study subjects, or a value that is within 10% of the stated value.

[0054] As used herein, the term "essentially free" with respect to a specified component is used herein to mean that none of the specified components are intentionally formulated into the composition and / or are present only as contaminants or in trace amounts. Thus, the total amount of the specified component resulting from unintentional contamination of the composition is less than 0.5%, less than 0.1%, or less than 0.05%, preferably less than 0.01%. Most preferred are compositions in which the amount of the specified component is undetectable by standard analytical methods.

[0055] Throughout this specification, when compositions are described as having, including, or comprising particular ingredients, or when processes and methods are described as having, including, or comprising particular steps, it is further contemplated that there are compositions of the invention that consist essentially of or consist of the recited ingredients, and that there are processes and methods of the invention that consist essentially of or consist of the recited processing steps.

[0056] In this application, when an element or component is described as being included in and / or selected from a recited list of elements or components, it is understood that the element or component can be any one of the recited elements or components, or the element or component can be selected from a group consisting of two or more of the recited elements or components.

[0057] Furthermore, it should be understood that the elements and / or features of the compositions or methods described herein, whether expressly or implicitly stated herein, can be combined in various ways without departing from the spirit and scope of the invention. For example, if reference is made to a particular compound, that compound can be used in various aspects of the compositions and / or methods of the invention, unless otherwise understood from the context. In other words, within this application, although embodiments have been described and illustrated to allow a clear and concise application to be described and depicted, it is intended and will be understood that the embodiments can be combined or separated in various ways without departing from the present teachings and invention. For example, it should be understood that all features described and illustrated herein may be applicable to all aspects of the invention described and illustrated herein.

[0058] It should be understood that the order of steps or order for performing certain actions is immaterial so long as the invention remains operable. Moreover, two or more steps or actions may be conducted simultaneously.

[0059] Any examples herein, or the use of exemplary language, such as "such as" or "including," are intended merely to better illustrate the invention and do not pose a limitation on the scope of the invention unless claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0060] Use of the terms "include," "includes," "including," "have," "has," "having," "contain," "contains," or "containing," including grammatical equivalents thereof, should generally be understood as open-ended and non-limiting, and for example, does not exclude additional, unrecited elements or steps unless otherwise specified or understood from the context.

[0061] II. Antibodies and Antibody Modifications Provided herein are monoclonal antibodies having clones and paired CDRs from the heavy and light chains exemplified in Tables 1, 6, 10, and 11. Such antibodies can be made using the methods described herein.

[0062] The monoclonal antibody of the present invention has several uses, including the manufacture of diagnostic kits for detecting HSP70 and treating diseases associated with increased levels of HSP70.In these contexts, such antibodies can be linked to diagnostic or therapeutic agents, can be used as capture agents or competitors in competitive assays, or can be used individually without attaching additional agents to such antibodies.Antibodies can be mutated or modified, as will be further discussed below.Methods for preparing and characterizing antibodies are well known in the art (see, for example, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, 1988; U.S. Patent No. 4,196,265).

[0063] An "antibody" is an immunoglobulin molecule capable of specifically binding to a target, such as a carbohydrate, polynucleotide, lipid, or polypeptide, via at least one antigen recognition site located in the variable region of the immunoglobulin molecule. As used herein, the term encompasses not only intact polyclonal or monoclonal antibodies, but also fragments thereof (Fab, Fab', F(ab')2, Fv, Fd, Fd', single-chain antibodies (ScFv), diabodies, linear antibodies, etc.), mutants, naturally occurring variants thereof, fusion proteins comprising an antibody portion having an antigen recognition site of the required specificity, humanized antibodies, chimeric antibodies, and any other modified configuration of an immunoglobulin molecule containing an antigen recognition site of the required specificity.

[0064] An "isolated antibody" is an antibody that has been separated and / or recovered from a component of its natural environment. Contaminant components of its natural environment are materials that would interfere with diagnostic or therapeutic uses for the antibody, and may include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. In particular examples, the antibody is purified to (1) greater than 95% by weight, most particularly greater than 99% by weight, antibody, as determined by the Lowry method, or (2) homogeneity by SDS-PAGE under reducing or non-reducing conditions using Coomassie blue or silver staining. Isolated antibody includes the antibody in situ within recombinant cells, since at least one component of the antibody's natural environment will not be present. Ordinarily, however, isolated antibody will be prepared by at least one purification step.

[0065] The basic four-chain antibody unit is a heterotetrameric glycoprotein composed of two identical light (L) chains and two identical heavy (H) chains. As used herein, the term "heavy chain" refers to a larger immunoglobulin subunit that is associated through its amino-terminal region with an immunoglobulin light chain. The heavy chain contains a variable region (V H ) and constant region (C H The constant region contains C H 1. Hinge, C H 2 and C HIn the case of IgE, IgM and IgY, the heavy chain contains three domains: C H Heavy chains comprise four domains but lack a hinge domain. Those skilled in the art will understand that heavy chains are classified as gamma, mu, alpha, delta, or epsilon (γ, μ, α, δ, ε), with several subclasses within them (e.g., γ1-γ4, α1-α2). It is the nature of this chain that determines the "class" of an antibody as IgG, IgM, IgA IgD, or IgE, respectively. Immunoglobulin subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, etc., are well characterized and are known to confer functional specialization.

[0066] As used herein, the term "light chain" refers to the smaller immunoglobulin subunit associated with the amino-terminal region of a heavy chain. Like heavy chains, light chains contain a variable region (V L ) and constant region (C L The light chain contains its constant domain (C L ), which can associate with any pair of different heavy chains to form immunoglobulin molecules. Also included within the meaning of light chain is a light chain having a lambda variable region (V-lambda) linked to a kappa constant region (C-kappa) or a kappa variable region (V-kappa) linked to a lambda constant region (C-lambda).

[0067] IgM antibodies, for example, consist of five basic heterotetrameric units together with an additional polypeptide called the J chain and therefore contain 10 antigen-binding sites, whereas secreted IgA antibodies can polymerize to form multivalent assemblies containing two to five basic four-chain units together with the J chain. In the case of IgG, the four-chain unit is generally about 150,000 daltons. Each L chain is linked to an H chain by one covalent disulfide bond, and the two H chains are linked to each other by one or more disulfide bonds depending on the H chain isotype. Each H and L chain also has regularly spaced intrachain disulfide bridges. Each H chain contains a variable region (V) at its N-terminus. H ), followed by three constant domains (C H ) followed by four C for mu and isotype H Each L chain has a variable domain (V L ) followed by a constant domain (C L ) V L is V H Align with C L is the first constant domain of the heavy chain (C H 1). Certain amino acid residues are thought to form an interface between the light-chain variable region and the heavy-chain variable region. V H and V L Together, these pairings form a single antigen-binding site. For the structure and properties of various classes of antibodies, see, e.g., Basic and Clinical Immunology, 8th edition, Daniel P. Stites, Abba I. Terr and Tristram G. Parslow (eds.), Appleton & Lange, Norwalk, Conn., 1994, page 71, and Chapter 6.

[0068] The "variable region" of an antibody refers to the variable region of the antibody light chain or the variable region of the antibody heavy chain, either alone or in combination. The term "variable" refers to the fact that certain segments of the variable region differ significantly in sequence among antibodies.L ) and the heavy chain portion (V H The variable regions of both heavy and light chains mediate antigen binding and define the specificity of a particular antibody for its particular antigen. However, variability is not evenly distributed throughout the variable regions. Instead, variable regions consist of relatively invariant stretches called framework regions (FRs) separated by shorter regions of extreme variability called complementarity-determining regions (CDRs) or hypervariable regions. Naturally occurring heavy and light chain variable regions each contain four FRs that primarily adopt a beta-sheet conformation, connected by three CDRs that form loops connecting, and in some cases forming part of, the beta-sheet structure. The CDRs complement the shape of the antigen and determine the affinity and specificity of the antibody for the antigen. V L and V H There are six CDRs in both the ribosomal and ribosomal amino acids. The CDRs in each chain are held in close proximity by FRs and, together with the CDRs of the other chain, contribute to the formation of the antigen-binding site of antibodies (see Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991)).

[0069] As used herein, the term "hypervariable region" refers to the amino acid residues of an antibody that are responsible for antigen binding. Hypervariable regions generally consist of amino acid residues from the "complementarity determining regions" or "CDRs" (e.g., V, VL ... L Approximately residues 24-34 (L1), 50-56 (L2), and 89-97 (L3), as well as V HApproximately residues 31-35 (H1), 50-65 (H2), and 95-102 (H3) in the H1 sequence; Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991)); and / or residues from the "hypervariable loops" (e.g., V, when numbered according to the Chothia numbering system). L Residues 24–34 (L1), 50–56 (L2), and 89–97 (L3) in V H residues 26-32 (H1), 52-56 (H2), and 95-101 (H3); Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)); and / or residues from the "hypervariable loops" / CDRs (e.g., V, when numbered according to the IMGT numbering system). L Residues 27–38 (L1), 56–65 (L2), and 105–120 (L3) in V H 27-38 (H1), 56-65 (H2), and 105-120 (H3); Lefranc, MP et al. Nucl. Acids Res. 27:209-212 (1999), Ruiz, M. et al. Nucl. Acids Res. 28:219-221 (2000)). Optionally, the antibody, when numbered according to AHo, includes V L The following points in the figure are 28, 36 (L1), 63, 74-75 (L2) and 123 (L3), as well as V H The CDRs have symmetric insertions at one or more of positions 28, 36 (H1), 63, 74-75 (H2), and 123 (H3); Honneger, A. and Plunkthun, AJ Mol. Biol. 309:657-670 (2001)). As used herein, CDRs can refer to CDRs defined by any of these numbering approaches, or by a combination of approaches, or by any other desired approach. Additionally, new definitions of highly conserved core, boundary, and hypervariable regions can be used.

[0070] The "constant region" of an antibody refers to the constant region of the antibody light chain or the constant region of the antibody heavy chain, either alone or in combination. L ) and heavy chains (C in the case of IgM and IgE). H 1. C H 2 or C H 3 or C H The constant region of 4) confers important biological properties such as secretion, transplacental transfer, Fc receptor binding, and complement fixation. By convention, the numbering of constant region domains increases as the constant region domain becomes more distal from the antigen-binding site or amino-terminus of the antibody. The constant region is not directly involved in binding of the antibody to the antigen, but exhibits various effector functions, such as antibody participation in antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), antibody-dependent neutrophil phagocytosis (ADNP), and antibody-dependent complement deposition (ADCD).

[0071] An antibody may be an antibody fragment. An "antibody fragment" comprises only a portion of an intact antibody, generally including the antigen-binding site of an intact antibody, and thus retains the ability to bind to antigen. Examples of antibody fragments encompassed by this definition include: (i) V L , C L , V H and C H1 (ii) a Fab fragment having a C domain; H1 (iii) a Fab' fragment, which is a Fab fragment having one or more cysteine ​​residues at the C-terminus of the domain; H and C H (iv) Fd fragment having one domain; H and C H 1 domain and C H (v) a single antibody V fragment having one or more cysteine ​​residues at the C-terminus of one domain; L and V H (vi) Fv fragment having a V domain; H(vii) isolated CDR regions; (viii) F(ab')2 fragments, which are bivalent fragments containing two Fab' fragments linked by a disulfide bridge at the hinge region; (ix) single-chain antibody molecules (e.g., single-chain Fv; scFv); (x) light chain variable domains (V) in the same polypeptide chain. L ) linked to a heavy chain variable domain (V H (xi) a "diabody" having two antigen-binding sites, each comprising a tandem Fd segment (V) that, together with a complementary light chain polypeptide, forms a pair of antigen-binding regions; H -C H 1-V H -C H 1) is a "linear antibody" containing the pair

[0072] The antibody may be a chimeric antibody. A "chimeric antibody" refers to an antibody in which a portion of each of the heavy and light chain amino acid sequences is homologous to the corresponding sequence in antibodies from a particular species or class, while the remaining segments of the chain are homologous to the corresponding sequence in another antibody. For example, a chimeric antibody may contain an antigen-binding sequence from a nonhuman donor grafted onto heterologous nonhuman, human, or humanized sequences (e.g., framework and / or constant domain sequences). Typically, in these chimeric antibodies, the variable regions of both the light and heavy chains mimic the variable regions of antibodies from one mammalian species, while the constant regions are homologous to sequences in antibodies from another species. For example, methods have been developed to replace the light and heavy chain constant domains of a monoclonal antibody with similar domains of human origin, leaving the variable regions of the foreign antibody intact. Alternatively, "fully human" monoclonal antibodies can be produced in mice transgenic for human immunoglobulin genes. Methods have also been developed to convert the variable domains of monoclonal antibodies into more human forms by recombinantly constructing antibody variable domains with both rodent (e.g., mouse) and human amino acid sequences. In "humanized" monoclonal antibodies, only the hypervariable CDRs are derived from mouse monoclonal antibodies, while the framework and constant regions are derived from human amino acid sequences (see U.S. Patent Nos. 5,091,513 and 6,881,557, incorporated herein by reference). Replacing amino acid sequences in antibodies characteristic of rodents with amino acid sequences found at the corresponding positions in human antibodies is believed to reduce the likelihood of adverse immune reactions during therapeutic use. Hybridomas or other cells producing antibodies may also undergo genetic mutations or other changes that may or may not alter the binding specificity of the antibodies produced by the hybridoma.

[0073] A. Monoclonal Antibodies As used herein, the term "monoclonal antibody" refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations, which include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. In addition to their specificity, monoclonal antibodies are advantageous in that they may be synthesized uncontaminated by other antibodies. The modifier "monoclonal" should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies useful in the present disclosure can be prepared by the hybridoma method first described by Kohler et al., Nature, 256:495 (1975), or can be produced using recombinant DNA methods in bacteria, eukaryotic animal, or plant cells (e.g., U.S. Patent No. 4,816,567) after single-cell sorting of antigen-specific B cells, antigen-specific plasmablasts in response to infection or immunization, or capturing linked heavy and light chains from single cells in bulk-sorted antigen-specific collections. Monoclonal antibodies can also be isolated from phage antibody libraries using the techniques described, for example, in Clackson et al., Nature 352:624-628 (1991) and Marks et al., J. Mol. Biol. 222:581-597 (1991).

[0074] Methods for producing various types of monoclonal antibodies, including humanized, chimeric, and fully human, are well known in the art and are highly predictable. For example, the following U.S. patents and patent applications provide enabling descriptions of such methods: U.S. Patent Application Nos. 2004 / 0126828 and 2002 / 0172677; and U.S. Patent Nos. 3,817,837; 3,850,752; 3,939,350; 3,996,345; 4,196,265; 4,275,149; 4,277,437; No. 66,241; No. 4,469,797; No. 4,472,509; No. 4,606,855; No. 4,703,003; No. 4,742,159; No. 4,767,720; No. 4,8 No. 16,567; No. 4,867,973; No. 4,938,948; No. 4,946,778; No. 5,021,236; No. 5,164,296; No. 5,196,066; No. 5,22 3,409; 5,403,484; 5,420,253; 5,565,332; 5,571,698; 5,627,052; 5,656,434; 5,77 No. 0,376; No. 5,789,208; No. 5,821,337; No. 5,844,091; No. 5,858,657; No. 5,861,155; No. 5,871,907; No. 5,969 ,108; 6,054,297; 6,165,464; 6,365,157; ​​6,406,867; 6,709,659; 6,709,873; 6,753,407; 6,814,965; 6,849,259; 6,861,572; 6,875,434; 6,891,024, each of which is incorporated herein by reference.

[0075] B. Single chain antibody Single-chain variable fragments (scFvs) are fusions of the variable regions of the heavy and light chains of an immunoglobulin linked together by a short linker. This chimeric molecule retains the specificity of the original immunoglobulin despite the removal of the constant region and the introduction of a linker peptide. This modification usually leaves the specificity unchanged. scFvs can be generated directly from subcloned heavy and light chains derived from hybridomas or B cells. Single-chain variable fragments lack the constant Fc region found in intact antibody molecules and therefore lack the common binding sites (e.g., protein A / G) used to purify antibodies. These fragments can often be purified / immobilized using protein L, as protein L interacts with the variable region of the kappa light chain.

[0076] Flexible linkers are generally composed of helix- and turn-promoting amino acid residues, such as alanine, serine, and glycine. However, other residues can function as well. For example, a linker can be V H It has a proline residue two residues after the C-terminus and may have abundant arginine and proline at other positions.

[0077] Single-chain antibodies can also be produced by linking the receptor light and heavy chains using a non-peptide linker or chemical unit. Generally, the light and heavy chains are produced in separate cells, purified, and then linked together in an appropriate manner (i.e., the N-terminus of the heavy chain is attached to the C-terminus of the light chain via a suitable chemical bridge).

[0078] Cross-linking reagents are used to form molecular bridges that connect the functional groups of two different molecules, such as stabilizers and coagulants.However, it is conceivable that heteromeric complexes can be made that are composed of dimers or multimers of the same analog or different analogs.To link two different compounds stepwise, heterobifunctional cross-linking agents can be used, which eliminates the formation of undesired homopolymers.

[0079] Exemplary heterobifunctional crosslinkers contain two reactive groups, one that reacts with primary amine groups (e.g., N-hydroxysuccinimide) and the other that reacts with thiol groups (e.g., pyridyl disulfide, maleimide, halogen, etc.). Through the primary amine-reactive group, the crosslinker can react with a lysine residue of one protein (e.g., a selected antibody or fragment), and the crosslinker already attached to the first protein reacts with a cysteine ​​residue (free sulfhydryl group) of another protein (e.g., a selected agent) through the thiol-reactive group.

[0080] Preferably, the crosslinker used has reasonable stability in blood.Many kinds of disulfide bond-containing linkers are known, which can be used successfully to conjugate targeting agent and therapeutic / prophylactic agent.Containing sterically hindered disulfide bond linkers may prove to provide greater stability in vivo, and prevent targeting peptide from being released before reaching the site of action.Therefore, these linkers are a group of linking agents.

[0081] For example, SMPT is a bifunctional crosslinker containing a disulfide bond "sterically hindered" by adjacent benzene rings and methyl groups. The steric hindrance of the disulfide bond is thought to protect the bond from attack by thiolate anions, such as glutathione, which may be present in tissues and blood, thereby helping to prevent disjugation of the conjugate before the linked agent is delivered to the target site. Like many other known crosslinking reagents, the SMPT crosslinking reagent confers the ability to crosslink functional groups such as the SH of cysteine ​​or primary amines (e.g., the ε-amino group of lysine). Another possible type of crosslinker includes heterobifunctional photoreactive phenyl azides containing a cleavable disulfide bond, such as sulfosuccinimidyl-2-(p-azidosalicylamido)ethyl-1,3'-dithiopropionate. The N-hydroxy-succinimidyl group reacts with primary amino groups, and the phenyl azide (upon photolysis) reacts nonselectively with any amino acid residue.

[0082] In addition to hindered crosslinkers, unhindered linkers can also be used in accordance with the present invention.Other useful crosslinkers that are not considered to contain or generate protected disulfides include SATA, SPDP, and 2-iminothiolane.The use of such crosslinkers is well understood in the art.Flexible linkers can also be used.

[0083] U.S. Patent No. 4,680,338 describes bifunctional linkers useful for generating conjugates of ligands with amine-containing polymers and / or proteins, particularly for forming antibody conjugates with chelators, drugs, enzymes, detectable labels, etc. U.S. Patent Nos. 5,141,648 and 5,563,250 disclose cleavable conjugates containing labile bonds that are cleavable under various mild conditions. This linker is particularly useful in that the active substance of interest can be directly bound to the linker, and cleavage results in the release of the active substance. Specific applications include adding free amino or free sulfhydryl groups to proteins such as antibodies or drugs.

[0084] US Patent No. 5,856,456 provides a peptide linker for use in linking polypeptide components to create fusion proteins, such as single-chain antibodies.The linker is up to about 50 amino acids long and contains at least one proline followed by a charged amino acid (preferably arginine or lysine), and is characterized by greater stability and reduced aggregation.US Patent No. 5,880,270 discloses aminooxy-containing linkers useful in various immunodiagnostic and separation techniques.

[0085] C. Bispecific and Multispecific Antibodies Antibodies can be bispecific or multispecific. A "bispecific antibody" is an antibody that has binding specificities for at least two different epitopes. Exemplary bispecific antibodies can bind to two different epitopes of a single antigen. Other such antibodies may combine a first antigen-binding site with a second antigen-binding site. Alternatively, to focus and localize cellular defense mechanisms to infected cells, an antigen-specific arm can be combined with an arm that binds to a trigger molecule on leukocytes, such as a T cell receptor molecule (e.g., CD3), or an Fc receptor for IgG (FcγR), such as FcγRI (CD64), FcγRII (CD32), and FcγRIII (CD16). Bispecific antibodies can also be used to localize cytotoxic agents to infected cells. These antibodies have an antigen-binding arm and an arm that binds to a cytotoxic agent (e.g., saporin, anti-interferon-α, vinca alkaloid, ricin A chain, methotrexate, or a radioisotope hapten). Bispecific antibodies can be prepared as full-length antibodies or antibody fragments (e.g., F(ab')2 bispecific antibodies). Taki et al. (2015) describe a bispecific anti-HSP70 / anti-CD3 antibody.

[0086] Methods for producing bispecific antibodies are known in the art. Traditional production of full-length bispecific antibodies is based on the co-expression of two immunoglobulin heavy chain-light chain pairs, with the two chains having different specificities. Due to the random combination of immunoglobulin heavy chains and immunoglobulin light chains, these hybridomas (quadromas) produce a potential mixture of 10 different antibody molecules, of which only one has the correct bispecific structure. Purification of the correct molecule, usually performed by affinity chromatography steps, is quite laborious and results in low product yields.

[0087] According to a different approach, antibody variable domains with the desired binding specificities (antibody-antigen combining sites) are fused to immunoglobulin constant domain sequences. Preferably, the fusion comprises a hinge, C H2 and C H3The first heavy-chain constant region (C) containing the site necessary for light chain binding is present in at least one of the fusions. H1 ) is preferred. DNA encoding the immunoglobulin heavy chain fusion and, if desired, the immunoglobulin light chain are inserted into separate expression vectors and co-transfected into a suitable host cell. This provides greater flexibility in adjusting the mutual proportions of the three polypeptide fragments when unequal ratios of the three polypeptide chains used in the construction provide the optimal yield of the desired bispecific antibody. However, if expression of at least two polypeptide chains in equal ratios results in high yields, or if the ratio does not significantly affect the yield of the desired chain combination, it is possible to insert the coding sequences for two or all three polypeptide chains into a single expression vector.

[0088] Bispecific antibodies can be composed of a hybrid immunoglobulin heavy chain with a first binding specificity in one arm and a hybrid immunoglobulin heavy chain-light chain pair (providing a second binding specificity) in the other arm. This asymmetric structure facilitates separation of the desired bispecific compound from undesired immunoglobulin chain combinations, since the presence of an immunoglobulin light chain in only one half of the bispecific molecule provides a facile separation method. This approach is disclosed in WO 94 / 04690. For further details on generating bispecific antibodies, see, e.g., Suresh et al., Methods in Enzymology, 121:210 (1986).

[0089] According to another approach described in U.S. Pat. No. 5,731,168, the interface between a pair of antibody molecules can be engineered to maximize the percentage of heterodimers which are recovered from recombinant cell culture. A preferred interface is C H3The antibody comprises at least a portion of a domain. In this method, one or more small amino acid side chains from the interface of a first antibody molecule are replaced with larger side chains (e.g., tyrosine or tryptophan). By replacing the large amino acid side chains with smaller ones (e.g., alanine or threonine), a compensatory "cavity" of identical or similar size to the large side chain is created on the interface of a second antibody molecule. This provides a mechanism for increasing the yield of heterodimers over other unwanted end-products, such as homodimers.

[0090] Bispecific antibodies include cross-linked or "heteroconjugate" antibodies. For example, one of the antibodies in the heteroconjugate can be bound to avidin, and the other to biotin. Such antibodies are intended, for example, to target immune system cells to unwanted cells (U.S. Pat. No. 4,676,980). Heteroconjugate antibodies can be produced using any convenient cross-linking method. Suitable cross-linking agents are well known in the art and are disclosed in U.S. Pat. No. 4,676,980, along with several cross-linking techniques.

[0091] Techniques for generating bispecific antibodies from antibody fragments have also been described in the literature. For example, bispecific antibodies can be prepared using chemical linkage. Brennan et al., Science, 229:81 (1985) describes a procedure in which intact antibodies are proteolytically cleaved to generate F(ab')2 fragments. To stabilize vicinal dithiols and prevent intermolecular disulfide formation, these fragments are reduced in the presence of the dithiol complexing agent sodium arsenite. The resulting Fab' fragments are then converted to thionitrobenzoate (TNB) derivatives. One of the Fab'-TNB derivatives is then reconverted to the Fab'-thiol by reduction with mercaptoethylamine and mixed with an equimolar amount of the other Fab'-TNB derivative to form the bispecific antibody. The resulting bispecific antibody can be used as an agent for the selective immobilization of enzymes.

[0092] Technology exists that facilitates the direct recovery from E. coli of Fab'-SH fragments, which can be chemically coupled to form bispecific antibodies. Shalaby et al., J. Exp. Med., 175:217-225 (1992) describe the production of humanized bispecific antibody F(ab')2 molecules. Each Fab' fragment was separately secreted from E. coli and subjected to directed chemical coupling in vitro to form the bispecific antibody. The bispecific antibody thus formed was capable of binding to cells overexpressing the ErbB2 receptor and normal human T cells, as well as triggering the lytic activity of human cytotoxic lymphocytes against human breast tumor targets.

[0093] Various techniques for producing and isolating bispecific antibody fragments directly from recombinant cell culture have also been described (Merchant et al., Nat. Biotechnol. 16, 677-681 (1998)). For example, bispecific antibodies have been produced using leucine zippers (Kostelny et al., J. Immunol., 148(5):1547-1553, 1992). Leucine zipper peptides from Fos and Jun proteins were linked to the Fab' portions of two different antibodies by gene fusion. Antibody homodimers were reduced at the hinge region to form monomers and then reoxidized to form antibody heterodimers. This method can also be used to produce antibody homodimers. The "diabody" technology described by Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993) provided an alternative mechanism for producing bispecific antibody fragments. This fragment is separated from the V by a linker that is too short to allow pairing between the two domains on the same chain. L V connected to H Therefore, one fragment of V H and V L The domain is a complementary V L and V HThe Fv domains are paired with each other to form two antigen-binding sites. Another strategy for making bispecific antibody fragments by the use of single-chain Fv (sFv) dimers has also been reported. See Gruber et al., J. Immunol., 152:5368 (1994).

[0094] Bispecific or multispecific antibodies can be formed as DOCK-AND-LOCK™ (DNL™) complexes (see, e.g., U.S. Patent Nos. 7,521,056; 7,527,787; 7,534,866; 7,550,143; and 7,666,400). Generally, this technology utilizes the specific and high-affinity binding interaction between the dimerization and docking domain (DDD) sequence of the regulatory (R) subunit of cAMP-dependent protein kinase (PKA) and the anchor domain (AD) sequence from any of various AKAP proteins (Baillie et al., FEBS Letters. 2005;579:3264; Wong and Scott, Nat. Rev. Mol. Cell Biol. 2004;5:959). The DDD and AD peptides can be conjugated to any protein, peptide, or other molecule. Because the DDD sequence spontaneously dimerizes and binds to the AD sequence, this technique allows the formation of a complex between any selected molecule that can be bound to the DDD or AD sequence.

[0095] Antibodies with more than two binding valencies are contemplated.For example, trispecific antibodies can be prepared (Tutt et al., J.Immunol.147:60,1991; Xu et al., Science,358(6359):85-90,2017).These antibodies can also comprise sequences or moieties that allow receptor dimerization or multimerization.Such sequences include sequences derived from IgA that allow the formation of multimers with J chain.Another multimerization domain is the Gal4 dimerization domain.

[0096] Multivalent antibodies may be internalized (and / or catabolized) more quickly than bivalent antibodies by cells expressing the antigen to which the antibody binds. The antibodies of the present disclosure may be multivalent antibodies having three or more antigen-binding sites (e.g., tetravalent antibodies), which can be easily produced by recombinant expression of nucleic acids encoding the polypeptide chains of the antibody. Multivalent antibodies can comprise a dimerization domain and three or more antigen-binding sites. A preferred dimerization domain comprises (or consists of) an Fc region or hinge region. In this scenario, the antibody comprises an Fc region and three or more antigen-binding sites amino-terminal to the Fc region. Multivalent antibodies can comprise (or consist of) three to about eight, e.g., four, antigen-binding sites. Multivalent antibodies comprise at least one polypeptide chain (preferably two polypeptide chains), and the polypeptide chains comprise two or more variable regions. For example, the polypeptide chains can be VD1-(X1). n -VD2-(X2) n -Fc, where VD1 is a first variable region and VD2 is a second variable region, Fc is one polypeptide chain of the Fc region, X1 and X2 represent amino acids or polypeptides, and n is 0 or 1. For example, the polypeptide chain may comprise a VH-CH1-flexible linker-VH-CH1-Fc region chain or a VH-CH1-VH-CH1-Fc region chain. The multivalent antibody herein may further comprise at least two (preferably four) light chain variable region polypeptides. The multivalent antibody herein may comprise, for example, about two to about eight light chain variable region polypeptides. The light chain variable region polypeptide contemplated herein comprises a light chain variable region, and optionally a C L Further includes domains.

[0097] Charge modifications are particularly useful in the context of multispecific antibodies, where amino acid substitutions in Fab molecules result in a reduction of mispairing of light chains with mismatched heavy chains (Bence-Jones by-products) that can occur in the generation of Fab-based bi / multispecific antigen-binding molecules involving VH / VL exchange in one (or more than one, in the case of molecules comprising more than two antigen-binding Fab molecules) of the binding arms of the Fab-based bi / multispecific antigen-binding molecule (see also WO 2015 / 150447, especially the Examples therein, which are incorporated herein by reference in their entirety).

[0098] D. Antibody Conjugates The antibodies of the present disclosure can be linked to at least one agent to form an antibody conjugate. The conjugate can be, for example, an antibody conjugated to another proteinaceous molecule, a carbohydrate molecule, a lipid molecule, or a mixed moiety molecule. Such antibody conjugates include, but are not limited to, modifications that involve linking the antibody to one or more polymers. For example, the antibody can be linked to one or more water-soluble polymers. Attachment to a water-soluble polymer reduces the likelihood that the antibody will precipitate in an aqueous environment, such as a physiological environment. Those skilled in the art can select an appropriate water-soluble polymer based on considerations including, but not limited to, whether the polymer / antibody conjugate will be used in patient treatment and, if so, the pharmacological profile of the antibody (e.g., half-life, dosage, activity, antigenicity, and / or other factors).

[0099] To increase the effectiveness of antibody molecules as diagnostic or therapeutic agents, it is conventional to link, covalently bond, or complex at least one desired molecule or moiety to the antibody. Such a molecule or moiety can be, but is not limited to, at least one effector or reporter molecule. Effector molecules include molecules with desired activity, such as cytotoxic activity. Non-limiting examples of effector molecules that have been conjugated to antibodies include toxins, antitumor agents, therapeutic enzymes, radionuclides, antiviral agents, chelators, cytokines, growth factors, and oligo- or polynucleotides. In contrast, a reporter molecule is defined as any moiety that can be detected using an assay. Non-limiting examples of reporter molecules that have been conjugated to antibodies include enzymes, radiolabels, haptens, fluorescent labels, phosphorescent molecules, chemiluminescent molecules, chromophores, photoaffinity molecules, colored particles or ligands, enzymes (e.g., that catalyze colorimetric or fluorometric reactions), substrates, solid matrices, such as biotin. An antibody can contain one, two, or more of any of these labels.

[0100] Antibody conjugates can be used to deliver cytotoxic agents to target cells. This type of cytotoxic agent can improve antibody-mediated cytotoxicity and can include moieties such as cytokines, radioisotopes, chemotherapeutic drugs (including prodrugs), bacterial toxins (e.g., Pseudomonas exotoxin, diphtheria toxin, etc.), plant toxins (e.g., ricin, gelonin, etc.), chemical conjugates (e.g., maytansinoid toxins, auristatins, α-amanitin, anthracyclines, calicheamicin, etc.), radioconjugates, enzyme conjugates (e.g., RNase conjugates, granzyme antibody-directed enzyme / prodrug therapy), etc., which directly or indirectly stimulate cell death.

[0101] Antibody conjugates are also used as diagnostic agents.Antibody diagnostics generally fall into two categories: those used in in vitro diagnostics such as various immunoassays, and those used in in vivo diagnostic protocols generally known as "antibody-directed imaging".Many suitable imaging agents are known in the art, as are the methods for binding imaging agents to antibodies (see, for example, U.S. Patent Nos. 5,021,236, 4,938,948 and 4,472,509).The imaging moiety used can be paramagnetic ions, radioisotopes, fluorescent dyes, NMR detectable substances and X-ray imaging agents.

[0102] Paramagnetic ions contemplated for use as conjugates include chromium(III), manganese(II), iron(III), iron(II), cobalt(II), nickel(II), copper(II), neodymium(III), samarium(III), ytterbium(III), gadolinium(III), vanadium(II), terbium(III), dysprosium(III), holmium(III), and / or erbium(III), with gadolinium being particularly preferred. Ions useful in other contexts, such as X-ray imaging, include, but are not limited to, lanthanum(III), gold(III), lead(II), and bismuth(III).

[0103] Radioisotopes envisioned for use as conjugates include astatine 211 , 14 carbon, 51 chromium, 36 chlorine, 57 cobalt, 58 Cobalt, Copper 67 , 152 Eu, gallium 67 , 3 Hydrogen, iodine 123 , iodine 125 , iodine 131 ,indium 111 , 59 iron, 32 Phosphorus, rhenium 186 ,rhenium 188 ,75 selenium, 35 Sulfur, Technetium 99m and / or yttrium 90 Examples include: 125 I is often preferred. Technetium 99m and / or indium 111 Radiolabeled monoclonal antibodies of the present disclosure can be prepared according to methods well known in the art. For example, monoclonal antibodies can be iodinated by contact with a chemical oxidizing agent, such as sodium iodide and / or potassium iodide and sodium hypochlorite, or an enzymatic oxidizing agent, such as lactoperoxidase. Monoclonal antibodies according to the present disclosure can be iodinated with technetium by a ligand exchange process, for example, by reducing pertechnate with a stannous solution, chelating the reduced technetium onto a Sephadex column, and applying the antibody to the column. 99m Alternatively, direct labeling techniques can be used, for example, by incubating the antibody with a reducing agent such as pertechnate or SNCl2, a buffer such as sodium-potassium phthalate solution, and the antibody. Intermediate functional groups often used to attach radioisotopes that exist as metal ions to antibodies are diethylenetriaminepentaacetic acid (DTPA) or ethylenediaminetetraacetic acid (EDTA).

[0104] Fluorescent labels contemplated for use as conjugates include Alexa 350, Alexa 430, AMCA, BODIPY 630 / 650, BODIPY 650 / 665, BODIPY-FL, BODIPY-R6G, BODIPY-TMR, BODIPY-TRX, Cascade Blue, Cy3, Cy5,6-FAM, fluorescein isothiocyanate, HEX, 6-JOE, Oregon Green 488, Oregon Green 500, Oregon Green 514, Pacific Blue, REG, rhodamine green, rhodamine red, renographin, ROX, TAMRA, TET, tetramethylrhodamine, and / or Texas Red.

[0105] A further type of antibody contemplated in the present disclosure is one intended primarily for in vitro use, in which the antibody is linked to a secondary binding ligand and / or to an enzyme (enzyme tag) that generates a colored product upon contact with a chromogenic substrate. Examples of suitable enzymes include urease, alkaline phosphatase, (horseradish) hydrogen peroxidase, or glucose oxidase. Preferred secondary binding ligands are biotin and avidin and streptavidin compounds.

[0106] Several methods for binding or conjugating antibodies to their conjugate moieties are known in the art. Some conjugation methods include, for example, the use of diethylenetriaminepentaacetic anhydride (DTPA); ethylenetriaminetetraacetic acid; N-chloro-p-toluenesulfonamide; and / or metal chelate complexes using organic chelators such as tetrachloro-3α-6α-diphenylglycouril-3 (U.S. Pat. Nos. 4,472,509 and 4,938,948) bound to antibodies. Monoclonal antibodies can also be reacted with enzymes in the presence of coupling agents such as glutaraldehyde or periodate. Conjugates with fluorescein markers are prepared in the presence of these coupling agents or by reaction with isothiocyanates. In U.S. Pat. No. 4,938,948, imaging of breast tumors is achieved using monoclonal antibodies, and a detectable imaging moiety is attached to the antibody using a linker such as methyl-p-hydroxybenzimidate or N-succinimidyl-3-(4-hydroxyphenyl)propionate.

[0107] Another known method for site-specific conjugation of molecules to antibodies involves reacting the antibody with a hapten-based affinity label. Essentially, the hapten-based affinity label reacts with amino acids in the antigen-binding site, thereby disrupting this site and blocking specific antigen reaction. However, this can be disadvantageous, as it results in loss of antigen binding by the antibody conjugate.

[0108] Molecules containing azide groups can also be used to form covalent bonds to proteins via reactive nitrene intermediates generated by low-intensity ultraviolet light. In particular, 2- and 8-azido analogs of purine nucleotides have been used as site-specific optical probes to identify nucleotide-binding proteins in crude cell extracts. 2- and 8-azido nucleotides have also been used to map nucleotide-binding domains in purified proteins and can be used as antibody binders.

[0109] Derivatization of immunoglobulins by selectively introducing sulfhydryl groups into the Fc region of the immunoglobulin using reaction conditions that do not alter the antibody binding site is also contemplated. Antibody conjugates produced according to this methodology have been disclosed to exhibit improved longevity, specificity, and sensitivity (U.S. Patent No. 5,196,066). Site-specific attachment of effector or reporter molecules, in which the reporter or effector molecule is conjugated to a carbohydrate residue in the Fc region, has also been disclosed in the literature. This approach has been reported to produce antibodies with diagnostic and therapeutic potential that are currently undergoing clinical evaluation.

[0110] E. Antibody-Drug Conjugates Antibody-drug conjugates (ADCs) are a class of highly potent biopharmaceuticals designed as targeted therapies for treating people with disease. ADCs are conjugated molecules composed of an antibody (whole mAb or antibody fragment such as scFv) linked to a biologically active cytotoxic / antiviral payload or drug via a stable chemical linker with a labile bond. Antibody-drug conjugates are examples of bioconjugates and immunoconjugates.

[0111] By combining the unique targeting capabilities of monoclonal antibodies with the cancer-killing potential of cytotoxic drugs, antibody-drug conjugates enable highly sensitive discrimination between healthy and diseased tissue, meaning that, in contrast to traditional systemic approaches, antibody-drug conjugates target and attack diseased cells while healthy cells are less severely affected.

[0112] In the development of ADC-based antitumor therapies, anticancer drugs (e.g., cytotoxins or cytotoxins) are conjugated to antibodies that specifically target certain cell markers (e.g., proteins ideally found only in or on diseased cells). The antibodies track these proteins throughout the body and attach to the surface of diseased cells. A biochemical reaction between the antibody and the target protein (antigen) triggers a signal in the targeted cells, which then absorb or internalize the antibody along with the cytotoxin. After the ADC is internalized, the cytotoxic drug is released, killing the cell or impairing cell replication. Due to this targeting, the drug ideally has fewer side effects than other agents and offers a wider therapeutic window.

[0113] A stable linkage between the antibody and the cytotoxic agent is a key aspect of ADCs. Linkers are based on chemical motifs, including disulfides, hydrazones, or peptides (cleavable) or thioethers (non-cleavable), to control the distribution and delivery of the cytotoxic agent to target cells. Both cleavable and non-cleavable linkers have been proven safe in preclinical and clinical studies. Brentuximab vedotin contains an enzyme-sensitive cleavable linker that delivers the potent and highly toxic anti-microtubule agent monomethyl auristatin E (MMAE), a synthetic anti-neoplastic agent, to human specific CD30-positive malignant cells. Due to its high toxicity, MMAE, which inhibits cell division by blocking tubulin polymerization, cannot be used as a single-agent chemotherapy drug. However, the combination of MMAE linked to an anti-CD30 monoclonal antibody (cAC10, a cell membrane protein of the tumor necrosis factor or TNF receptor), has been proven to be stable in extracellular fluids, cleavable by cathepsins, and safe for treatment. Another approved ADC, trastuzumab emtansine, is a combination of the microtubule inhibitor mertansine (DM-1), a derivative of maytansine, and the antibody trastuzumab (Herceptin® / Genentech / Roche) linked by a stable, non-cleavable linker.

[0114] The availability of better and more stable linkers has changed the function of chemical bonds. The type of cleavable or non-cleavable linker confers specific properties to cytotoxic (e.g., anti-cancer) drugs. For example, a non-cleavable linker retains the drug intracellularly. As a result, the entire antibody, linker, and cytotoxic agent enter the targeted cell, where the antibody is degraded to the amino acid level. The resulting complex—amino acid, linker, and cytotoxic agent—at this point becomes the active drug. In contrast, a cleavable linker is catalyzed by an enzyme in the host cell, thereby releasing the cytotoxic agent.

[0115] Another type of cleavable linker adds an additional molecule between the cytotoxic drug and the cleavage site. This linker technology allows researchers to create more flexible ADCs without worrying about altering the cleavage kinetics. Researchers are also developing new methods for peptide cleavage based on Edman degradation. Future directions in ADC development also include site-specific conjugation (TDC) and the development of alpha-releasing immunoconjugates and antibody-conjugated nanoparticles to further improve stability and therapeutic index. Antibody Production and Purification

[0116] Methods for generating monoclonal antibodies generally begin in the same way as methods for preparing polyclonal antibodies. The first step in both of these methods is immunization of a suitable host. As is well known in the art, the immunogenicity of a given composition for immunization can vary. Therefore, it is often necessary to enhance the host immune system, which can be achieved by coupling a peptide or polypeptide immunogen to a carrier. Exemplary and preferred carriers are keyhole limpet hemocyanin (KLH) and bovine serum albumin (BSA). Other albumins, such as ovalbumin, mouse serum albumin, or rabbit serum albumin, can also be used as carriers. Means for conjugating polypeptides to carrier proteins are well known in the art and include glutaraldehyde, m-maleimidobenocoyl-N-hydroxysuccinimide ester, carbodiimide, and bis-biazotized benzidine. As is also well known in the art, the immunogenicity of a particular immunogen composition can be enhanced by the use of nonspecific stimulators of the immune response, known as adjuvants. Exemplary and preferred adjuvants in animals include complete Freund's adjuvant (a non-specific stimulator of immune responses containing killed Mycobacterium tuberculosis), incomplete Freund's adjuvant, and aluminum hydroxide adjuvant, while in humans, alum, CpG, MFP59, and combinations of immunostimulatory molecules ("adjuvant systems" such as AS01 or AS03) are available. Additional experimental forms of inoculation to induce antigen-specific B cells are possible, such as nanoparticle vaccines, or gene-encoded antigens delivered as DNA or RNA genes in physical delivery systems (such as lipid nanoparticles or gold biolistic beads), delivered using needles, gene guns, or transcutaneous electroporation devices. Antigen genes can also be encoded and carried by replication-competent or replication-deficient viral vectors, such as adenovirus, adeno-associated virus, poxvirus, herpesvirus, or alphavirus replicons, or virus-like particles.

[0117] Methods for generating hybrids of antibody-producing cells and myeloma cells typically involve mixing somatic cells with myeloma cells at a 2:1 ratio in the presence of one or more agents (chemical or electrical) that promote cell membrane fusion; however, this ratio can vary from approximately 20:1 to approximately 1:1. In some cases, transformation of human B cells with Epstein-Barr virus (EBV) as an initial step increases the size of the B cells and enhances fusion with the relatively large myeloma cells. The efficiency of EBV transformation is enhanced by using CpG and Chk2 inhibitors in the transformation medium. Alternatively, human B cells can be activated by co-culturing them with a transfected cell line expressing CD40 ligand (CD154) in a medium containing additional soluble factors, such as IL-21 and human B-cell activating factor (BAFF), a type II member of the TNF superfamily. Fusion methods using Sendai virus or polyethylene glycol (PEG) are also known. Electrically induced fusion is also suitable. The fusion procedure typically involves approximately 1 × 10 -6 ~Approx. 1×10 -8Although viable hybrids are produced at a low frequency of 1 / 200, optimized procedures can achieve fusion efficiencies approaching 1 / 200. However, the relatively low efficiency of fusion does not pose a problem, since viable fused hybrids differentiate from the injected parent cells (especially the injected myeloma cells, which usually continue to divide indefinitely) by culturing in selective medium. Selective media generally contain agents that block de novo synthesis of nucleotides in tissue culture media. Exemplary and preferred agents are aminopterin, methotrexate, and azaserine. Aminopterin and methotrexate block de novo synthesis of both purines and pyrimidines, while azaserine blocks only purine synthesis. When aminopterin or methotrexate is used, the medium is supplemented with hypoxanthine and thymidine as a source of nucleotides (HAT medium). When azaserine is used, the medium is supplemented with hypoxanthine. If the B cell source is an EBV-transformed human B cell line, ouabain is added to eliminate EBV-transformed cells that have not fused to myeloma.

[0118] The preferred selective medium is HAT or HAT containing ouabain. Only cells capable of operating the nucleotide salvage pathway can survive in HAT medium. Myeloma cells lack key enzymes in the salvage pathway, such as hypoxanthine phosphoribosyltransferase (HPRT), and cannot survive. B cells can operate this pathway, but their lifespan in culture is limited and they generally die within about two weeks. Therefore, the only cells that can survive in selective medium are hybrids formed from myeloma and B cells. If the source of B cells used for fusion is an EBV-transformed B cell line, as in this case, ouabain can also be used for drug selection of the hybrids, since the EBV-transformed B cells are sensitive to drug killing, while the myeloma partner used is selected to be ouabain-resistant.

[0119] Culture provides a population of hybridomas from which specific hybridomas are selected. Hybridoma selection is typically performed by culturing the cells by single-clone dilution in microtiter plates and then testing individual clonal supernatants (after approximately 2–3 weeks) for the desired reactivity. The assay should be sensitive, simple, and rapid, such as radioimmunoassay, enzyme immunoassay, cytotoxicity assay, plaque assay, or dot immunobinding assay. Selected hybridomas are then serially diluted or single-cell sorted by flow cytometry sorting and cloned into individual antibody-producing cell lines, which can then be propagated indefinitely to produce monoclonal antibodies. Cell lines can be utilized for monoclonal antibody production in two basic ways: A sample of the hybridoma can be injected (often intraperitoneally) into an animal (e.g., a mouse). Optionally, the animal is primed with a hydrocarbon, particularly an oil such as pristane (tetramethylpentadecane), prior to injection. When human hybridomas are used in this manner, they are best injected into immunocompromised mice, such as SCID mice, to prevent tumor rejection. The injected animals develop tumors secreting the specific monoclonal antibodies produced by the fused cell hybrids. The animal's body fluids, such as serum or ascites, can then be extracted to provide high concentrations of the monoclonal antibodies. Individual cell lines can also be cultured in vitro, where the monoclonal antibodies are naturally secreted into the culture medium, from which they can be easily obtained in high concentrations. Alternatively, human hybridoma cell lines can be used in vitro to produce immunoglobulins in the cell supernatant. To optimize the ability to recover highly pure human monoclonal immunoglobulins, the cell lines can be adapted for growth in serum-free medium.

[0120] The hybridomas are cultured, the cells are then lysed, and total RNA is extracted. Random hexamers are used with RT to generate cDNA copies of the RNA, followed by PCR using a multiplexed mixture of PCR primers predicted to amplify all human variable gene sequences. The PCR products are cloned into the pGEM-T Easy vector and then sequenced by automated DNA sequencing using standard vector primers. Binding and neutralization assays can be performed using antibodies collected from hybridoma supernatants and purified by FPLC using a protein G column.

[0121] Recombinant full-length IgG antibodies can be produced by subcloning the heavy and light chain Fv DNA from the cloning vector into an IgG plasmid vector and transfecting it into 293 (e.g., Freestyle) or CHO cells, and the antibody can be collected and purified from the 293 or CHO cell supernatant. Other suitable host cell systems include bacteria such as E. coli, insect cells (S2, Sf9, Sf29, High Five), plant cells (e.g., tobacco, with or without engineering for human-like glycans), algae, or various non-human transgenic contexts such as mouse, rat, goat, or cow.

[0122] Expression of nucleic acids encoding antibodies is also contemplated, both for subsequent antibody purification and for host immunization. The antibody coding sequence can be RNA, such as natural or modified RNA. Modified RNA contemplates certain chemical modifications that confer increased stability and reduced immunogenicity to mRNA, thereby facilitating the expression of therapeutically important proteins. For example, N1-methyl-pseudouridine (N1mΨ) outperforms several other nucleoside modifications and their combinations in terms of translational potency. In addition to abolishing immune / eIF2α phosphorylation-dependent inhibition of translation, the incorporated N1mΨ nucleotide dramatically alters the dynamics of the translation process by increasing ribosome pausing and density on mRNA. The increased ribosome loading of modified mRNA makes the modified mRNA more susceptible to initiation by favoring either ribosome recycling or de novo ribosome recruitment on the same mRNA. Such modifications can be used to enhance antibody expression in vivo after RNA inoculation. RNA, whether natural or modified, can be delivered as naked RNA or in a delivery vehicle such as a lipid nanoparticle.

[0123] Alternatively, DNA encoding an antibody can be used for the same purpose. The DNA is contained in an expression cassette that contains a promoter that is active in the host cell for which the expression cassette is designed. The expression cassette is advantageously contained in a replicable vector, such as a conventional plasmid or minivector. The vector includes viral vectors, such as poxvirus, adenovirus, herpesvirus, adeno-associated virus, and lentivirus. Replicons encoding antibody genes, such as alphavirus replicons based on VEE virus or Sindbis virus, are also contemplated. Such vectors can be delivered by needle via intramuscular, subcutaneous, or intradermal route, or by transcutaneous electroporation if in vivo expression is desired.

[0124] Alternatively, molecular cloning approaches can be used to generate monoclonal antibodies. Single B cells labeled with the antigen of interest can be physically sorted using paramagnetic bead selection or flow cytometry sorting, followed by isolation of RNA from the single cells and amplified antibody genes by RT-PCR. Alternatively, bulk-sorted antigen-specific cell populations can be separated into microvesicles, and matched heavy and light chain variable genes can be recovered from single cells using physical linkage of heavy and light chain amplicons or common barcoding of heavy and light chain genes from the vesicles. Matched heavy and light chain genes from single cells can also be obtained from a population of antigen-specific B cells by treating the cells with RT-PCR primers and cell-permeable nanoparticles bearing barcodes to mark transcripts with one barcode per cell. Antibody variable genes can also be isolated by RNA extraction of hybridoma lines, and antibody genes can be obtained by RT-PCR and cloned into immunoglobulin expression vectors. Alternatively, combinatorial immunoglobulin phagemid libraries are prepared from RNA isolated from cell lines, and phagemids expressing suitable antibodies are selected by panning using viral antigens. The advantage of this approach over traditional hybridoma technology is that it allows for approximately 10 4 The advantages of this approach are that twice as many antibodies can be generated and screened in a single round, and that new specificities can be generated by combining heavy and light chains, further increasing the chances of finding a suitable antibody.

[0125] Other U.S. patents that teach the production of antibodies useful in the present disclosure, each of which is incorporated herein by reference, include U.S. Pat. No. 5,565,332, which describes the production of chimeric antibodies using a combinatorial approach; U.S. Pat. No. 4,816,567, which describes recombinant immunoglobulin preparations; and U.S. Pat. No. 4,867,973, which describes antibody-therapeutic agent conjugates.

[0126] Monoclonal antibodies produced by any means can be purified, if desired, using filtration, centrifugation, and various chromatographic methods, such as FPLC or affinity chromatography. Fragments of the monoclonal antibodies of the present disclosure can be obtained from the purified monoclonal antibodies by methods including digestion with enzymes, such as pepsin or papain, and / or by cleavage of disulfide bonds by chemical reduction. Alternatively, monoclonal antibody fragments encompassed by the present disclosure can be synthesized using an automated peptide synthesizer.

[0127] The antibodies disclosed herein can be isolated or purified. As used herein, the terms "isolated" and "purified" are intended to refer to a composition that can be isolated from other components, and the protein is purified to any degree compared to its naturally obtainable state. Thus, a purified protein also refers to a protein that has been removed from the environment in which it may naturally exist. When the term "substantially purified" is used, this designation refers to a composition in which the protein or peptide forms the majority of the composition, such as constituting about 50%, about 60%, about 70%, about 80%, about 90%, about 95% or more of the protein in the composition.

[0128] Protein purification techniques are well known to those skilled in the art. At one level, these techniques involve crude fractionation of the cellular environment into polypeptide and non-polypeptide fractions. After separating the polypeptide from other proteins, the polypeptide of interest can be further purified using chromatography and electrophoresis techniques to achieve partial or complete purification (or purification to homogeneity). Analytical methods particularly suitable for preparing pure peptides include ion exchange chromatography, exclusion chromatography; polyacrylamide gel electrophoresis; and isoelectric focusing. Other methods for protein purification include precipitation or heat denaturation with ammonium sulfate, PEG, antibodies, etc., followed by centrifugation; gel filtration, reverse phase, hydroxyapatite, and affinity chromatography; and combinations of these techniques with other techniques.

[0129] When purifying the antibodies of the present disclosure, it may be desirable to express the polypeptide in a prokaryotic or eukaryotic expression system and extract the protein using denaturing conditions. The polypeptide can be purified from other cellular components using an affinity column that binds to the tagged portion of the polypeptide. As is generally known in the art, the order of performing various purification steps can be changed, or certain steps can be omitted, and still result in a suitable method for preparing substantially purified proteins or peptides.

[0130] Generally, intact antibodies are fractionated using an agent that binds to the Fc portion of the antibody (i.e., protein A). Alternatively, antigens can be used to simultaneously purify and select suitable antibodies. Such methods often utilize a selection agent bound to a support, such as a column, filter, or beads. The antibody is bound to the support, contaminants are removed (e.g., washed away), and the antibody is released by applying conditions (salt, heat, etc.).

[0131] Various methods for quantifying the degree of purification of a protein or peptide will be known to those of skill in the art in light of the present disclosure. These include, for example, determining the specific activity of an active fraction or assessing the amount of polypeptide within a fraction by SDS / PAGE analysis. Another method for assessing the purity of a fraction is to calculate the specific activity of the fraction and compare it to the specific activity of the initial extract, thereby calculating the purity. The actual units used to express the amount of activity will, of course, depend on the particular assay technique chosen to follow purification and whether the expressed protein or peptide exhibits detectable activity.

[0132] It is known that polypeptide migration can vary, sometimes significantly, under different conditions of SDS / PAGE, and therefore it will be understood that the apparent molecular weight of purified or partially purified expression products may change under different electrophoretic conditions.

[0133] F. Antibody Modification The sequence of an antibody may be modified for a variety of reasons, such as improved expression, improved cross-reactivity, or reduced off-target binding. Modified antibodies may be made by any technique known to those of skill in the art, including expression through standard molecular biology techniques or chemical synthesis of polypeptides.

[0134] For example, one may desire to make modifications, such as introducing conservative changes into an antibody molecule. In making such changes, the hydropathic amino acid index can be considered. The importance of the hydropathic amino acid index in conferring interactive biological function on a protein is generally understood in the art (Kyte and Doolittle, 1982). It is recognized that the relative hydropathic properties of amino acids contribute to the secondary structure of a resulting protein, which in turn determines the interaction of the protein with other molecules, such as enzymes, substrates, receptors, DNA, antibodies, antigens, etc.

[0135] Substitution of similar amino acids can be effectively made based on hydrophilicity. U.S. Pat. No. 4,554,101, incorporated herein by reference, states that the greatest local average hydrophilicity of a protein, as governed by the hydrophilicity of its neighboring amino acids, correlates with the biological properties of the protein. As detailed in U.S. Pat. No. 4,554,101, the following hydrophilicity values ​​are assigned to amino acid residues: basic amino acids: arginine (+3.0), lysine (+3.0), and histidine (-0.5); acidic amino acids: aspartic acid (+3.0±1), glutamic acid (+3.0±1), asparagine (+0.2), and glutamine (+0.2); hydrophilic non-ionic amino acids: serine (+0.3), asparagine (+0.2 ), glutamine (+0.2) and threonine (-0.4), sulfur-containing amino acids: cysteine ​​(-1.0) and methionine (-1.3); hydrophobic non-aromatic amino acids: valine (-1.5), leucine (-1.8), isoleucine (-1.8), proline (-0.5±1), alanine (-0.5) and glycine (0); hydrophobic aromatic amino acids: tryptophan (-3.4), phenylalanine (-2.5) and tyrosine (-2.3).

[0136] An amino acid can be substituted for another amino acid with a similar hydrophilicity to produce a biologically or immunologically altered protein. Such changes are preferably made with amino acids whose hydrophilicity values ​​are within ±2, particularly preferably within ±1, and even more particularly preferably within ±0.5.

[0137] Amino acid substitutions are generally based on the relative similarity of the amino acid side-chain substituents, e.g., their hydrophobicity, hydrophilicity, charge, size, etc. Exemplary substitutions that take into consideration the various aforementioned characteristics are well known to those of skill in the art and include: arginine and lysine; glutamic acid and aspartic acid; serine and threonine; glutamine and asparagine; valine, leucine, and isoleucine.

[0138] The present disclosure also contemplates isotype modification. By modifying the Fc region to have a different isotype, different functionalities can be achieved. For example, changing to IgG1 can increase antibody-dependent cellular cytotoxicity, switching to class A can improve tissue distribution, and switching to class M can improve binding valency.

[0139] For example, the Fc region of an antibody can be engineered with altered effector functions by modifying C1q binding and / or FcγR binding, thereby altering CDC activity and / or ADCC activity. "Effector function" serves to activate or attenuate biological activity (e.g., in a subject). Examples of effector functions include, but are not limited to, C1q binding; complement-dependent cytotoxicity (CDC); Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; and downregulation of cell surface receptors (e.g., B cell receptors; BCRs). Such effector functions may require the Fc region to be combined with a binding domain (e.g., an antibody variable domain) and can be evaluated using various assays (e.g., Fc binding assays, ADCC assays, CDC assays, etc.).

[0140] For example, a variant Fc region of an antibody can be created that has improved C1q binding and improved FcγRIII binding (e.g., both improved ADCC activity and improved CDC activity). Alternatively, if reduced or eliminated effector function is desired, the variant Fc region can be engineered to have reduced CDC activity and / or reduced ADCC activity. In other embodiments, only one of these activities can be increased, and optionally the other activity can be decreased (e.g., to create an Fc region variant that has improved ADCC activity but reduced CDC activity, or vice versa).

[0141] In certain embodiments, the Fc domain can contain one or more mutations or modifications in either or both of the Fc polypeptide chains that alter binding to an Fcγ receptor (e.g., FcγRI / CD64, FcγRIIA / CD32A, FcγRIIB / CD32B, FcγRIIIIA / CD16, or FcγRIIIB). In some embodiments, the modified heavy chain constant region comprises a CH2 domain that is a wild-type CH2 domain of an IgG isotype (e.g., IgG1). The CH2 domain used herein can also be a variant of the wild-type CH2 domain, e.g., a variant of the wild-type IgG1 CH2 domain. Exemplary variants of the CH2 domain include variants that modulate the biological activity of the Fc region of an antibody, such as ADCC or CDC, or modulate the half-life / antibody stability of the antibody. The CH2 domain can have an enhanced effector function. The CH2 domain may contain one or more mutations at the following amino acids: E233, G236, G237, P238, H268, P271, L328, A330 and I332.

[0142] In some embodiments, the antibody can be modified to increase binding to FcγR2a and / or FcγR3a. Exemplary modifications that increase binding to FcγR2a include substitution at G236, for example, G236A. Exemplary modifications that increase binding to FcγR3a include substitution at G236, for example, G236A, substitution at A330, for example, A330L, and substitution at I332, for example, I332E.

[0143] The isolated monoclonal antibody or antigen-binding fragment thereof can contain substantially uniform glycans that are free of sialic acid, galactose, or fucose. The substantially uniform glycans can be covalently attached to the heavy chain constant region.

[0144] Monoclonal antibodies may have novel Fc glycosylation patterns. Glycosylation of the Fc region is typically either N-linked or O-linked. N-linked refers to the attachment of a carbohydrate moiety to the side chain of an asparagine residue. O-linked glycosylation refers to the attachment of one of the sugars N-acetylgalactosamine, galactose, or xylose to a hydroxyamino acid, most commonly serine or threonine, although 5-hydroxyproline or 5-hydroxylysine may also be used. The recognition sequences for enzymatic attachment of carbohydrate moieties to asparagine side chain peptide sequences are asparagine-X-serine and asparagine-X-threonine, where X is any amino acid except proline. Thus, the presence of either of these peptide sequences in a polypeptide creates a potential glycosylation site.

[0145] The glycosylation pattern can be altered, for example, by deleting one or more glycosylation sites found in the polypeptide and / or adding one or more glycosylation sites that are not present in the polypeptide. Addition of a glycosylation site to the Fc region of an antibody is conveniently achieved by modifying the amino acid sequence to contain one or more of the above tripeptide sequences (in the case of N-linked glycosylation sites). An exemplary glycosylation variant has an amino acid substitution of residue Asn297 of the heavy chain. Modification can also be made by adding or substituting one or more serine or threonine residues into the original polypeptide sequence (in the case of O-linked glycosylation sites). Furthermore, changing Asn297 to Ala can remove one of the glycosylation sites.

[0146] The isolated monoclonal antibody or antigen-binding fragment thereof may be present in a substantially homogeneous composition represented by the GNGN or G1 / G2 glycoforms and exhibit increased binding affinity to FcγRI and FcγRIII compared to the same antibody lacking substantially homogeneous GNGN glycoforms and possessing glycoforms containing G0, G1F, G2F, GNF, GNGNF, or GNGNFX. Fc glycosylation plays an important role in the antiviral and anticancer properties of therapeutic mAbs. Removal of core fucose dramatically improves the ADCC activity of mAbs mediated by natural killer (NK) cells, but appears to have the opposite effect on ADCC activity of polymorphonuclear cells (PMNs).

[0147] The isolated monoclonal antibody or its antigen-binding fragment can be expressed in cells that express β(1,4)-N-acetylglucosaminyltransferase III (GnTIII), so that GnTIII adds GlcNAc to the antibody.Methods for producing antibodies in this manner are provided in International Publication Nos. 9954342 and 03011878.Cell lines can be modified using genome editing techniques such as clustered regularly interspaced short palindromic repeats (CRISPR) to enhance, reduce, or eliminate certain post-translational modifications, such as glycosylation.For example, CRISPR technology can be used to eliminate the gene encoding the glycosylation enzyme in 293 cells or CHO cells used to express monoclonal antibodies.

[0148] Antibody variable gene sequences obtained from human B cells can be engineered to increase their manufacturability and safety. Potential protein sequence liabilities can be identified by searching for sequence motifs with sites containing: 1) an unpaired Cys residue, 2) N-linked glycosylation, 3) Asn deamidation, 4) Asp isomerization, 5) SYE cutting, 6) Met oxidation, 7) Trp oxidation, 8) N-terminal glutamic acid, 9) integrin binding, 10) CD11c / CD18 binding, or 11) Fragmentation Such motifs can be eliminated by modifying the synthetic gene containing the cDNA encoding the antibody.

[0149] Antibodies can be engineered to enhance solubility. For example, some hydrophilic residues, such as aspartic acid, glutamic acid, and serine, contribute significantly more favorably to protein solubility than other hydrophilic residues, such as asparagine, glutamine, threonine, lysine, and arginine.

[0150] Deep sequencing of the B cell repertoire of human B cells derived from blood donors has been extensively performed. Sequence information for a significant portion of the human antibody repertoire facilitates statistical evaluation of common antibody sequence features in healthy humans. Knowledge of antibody sequence features in the Human Recombinant Antibody Variable Gene Reference Database can be used to estimate the position-specific degree of "human similarity" (HL) of an antibody sequence. HL has been shown to be useful in developing antibodies for clinical use, such as therapeutic antibodies or antibodies as vaccines. The ultimate goal is to increase the human similarity of antibodies to reduce potential adverse effects and anti-antibody immune responses that can result in significantly reduced efficacy of antibody drugs or induce serious health consequences. We were able to evaluate the antibody characteristics of the combined antibody repertoire of three healthy human blood donors, totaling approximately 400 million sequences, and created a novel "relative human similarity" (rHL) score that focuses on the hypervariable regions of antibodies. The rHL score allows for easy distinction between human sequences (positive scores) and non-human sequences (negative scores). Antibodies can be engineered to eliminate residues that are uncommon in the human repertoire.

[0151] Methods for reducing or eliminating the antigenicity of antibodies and antibody fragments are known in the art. When antibodies are administered to humans, the antibodies are preferably "humanized" to reduce or eliminate their antigenicity in humans. Preferably, each humanized antibody has the same or substantially the same affinity for the antigen as the non-humanized murine antibody from which it is derived.

[0152] In one humanization approach, mouse immunoglobulin constant region is replaced with human immunoglobulin constant region to create chimeric protein.See for example, Morrison et al., 1984, PROC.NAT.ACAD.SCI. 81:6851-6855, Neuberger et al., 1984, Nature 312:604-608; U.S. Patent No. 6,893,625 (Robinson); U.S. Patent No. 5,500,362 (Robinson); and U.S. Patent No. 4,816,567 (Cabilly).

[0153] In the approach known as CDR grafting, the CDR of light chain variable region and heavy chain variable region is grafted onto the framework of another species.For example, mouse CDR can be grafted onto human FR.In some embodiments, the CDR of light chain variable region and heavy chain variable region of antibody is grafted onto human FR or consensus human FR.To create consensus human FR, FR from several human heavy chain or light chain amino acid sequences is aligned, and consensus amino acid sequence is identified. CDR porting is applicable to U.S. Patent Nos. 7,022,500 (Queen); 6,982,321 (Winter); 6,180,370 (Queen); 6,054,297 (Carter); ; 5,859,205 (Adair); 5,693,761 (Queen); 5,565,332 (Hoogenboom); 5,585,089 (Queen); 5,530,101 (Queen); Jones et al.(1986)Nature 321:522-525;Riechmann et al.(1988)Nature 332:323-327;Verhoeyen et al.(1988)Science 239:1534-1536; and Winter (1998) Febs Lett 430:92-94.

[0154] In an approach called "SUPERHUMANIZATION™," human CDR sequences are selected from human germline genes based on the structural similarity of the human CDRs to the CDRs of the mouse antibody being humanized. See, e.g., U.S. Patent No. 6,881,557 (Foote) and Tan et al., 2002, J. Immunol. 169:1119-1125.

[0155] Other methods for reducing immunogenicity include "reshaping," "high chimerization," and "veneering / resurfacing." See, for example, Vaswami et al., 1998, Annals of Allergy, Asthma, & Immunol. 81:105; Roguska et al., 1996, Prot. Engineer 9:895-904 and U.S. Patent No. 6,072,035 (Hardman). In the veneering / resurfacing approach, surface-accessible amino acid residues in mouse antibodies are replaced with amino acid residues that are more frequently found at the same positions in human antibodies. This type of antibody resurfacing is described, for example, in U.S. Patent No. 5,639,641 (Pedersen).

[0156] Another approach to converting mouse antibodies into a form suitable for human medical use is known as ACTIVMAB™ technology (Vaccinex, Inc., Rochester, NY), which involves a vaccinia virus-based vector for expressing antibodies in mammalian cells. High levels of combinatorial diversity of IgG heavy and light chains can be generated. See, for example, U.S. Patent Nos. 6,706,477 (Zauderer); 6,800,442 (Zauderer); and 6,872,518 (Zauderer). Another approach to converting mouse antibodies into a form suitable for human use is the technology commercially implemented by KaloBios Pharmaceuticals, Inc. (Palo Alto, CA). This technology involves the use of a proprietary human "acceptor" library to create an "epitope-focused" library for antibody selection. Another approach to modifying mouse antibodies into a form suitable for human medical use is the HUMAN ENGINEERING™ technology, which is commercially practiced by XOMA (US) LLC. See, for example, WO 93 / 11794 and U.S. Patent Nos. 5,766,886 (Studnicka); 5,770,196 (Studnicka); 5,821,123 (Studnicka); and 5,869,619 (Studnicka).

[0157] Any suitable approach can be used to reduce or eliminate human immunogenicity of antibodies, including any of the approaches described above.

[0158] G. Antibody Characterization Antibodies according to the present disclosure can be defined, in a first example, by their binding specificity. By evaluating the binding specificity / affinity of a given antibody using techniques well known to those skilled in the art, those skilled in the art can determine whether such an antibody falls within the scope of the present claims. For example, the epitope to which a given antibody binds may consist of a single contiguous sequence of three or more (e.g., 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20) amino acids located within an antigen molecule (e.g., a linear epitope in a domain). Alternatively, the epitope may consist of multiple non-contiguous amino acids (or amino acid sequences) located within an antigen molecule (e.g., a conformational epitope).

[0159] Various techniques known to those skilled in the art can be used to determine whether an antibody "interacts with one or more amino acids" within a polypeptide or protein. Exemplary techniques include routine cross-blocking assays, such as those described in Antibodies, Harlow and Lane (Cold Spring Harbor Press, Cold Spring Harbor, NY). Cross-blocking can be measured by various binding assays, such as ELISA, biolayer interferometry, or surface plasmon resonance. Other methods include alanine scanning mutation analysis, peptide blot analysis (Reineke (2004) Methods Mol. Biol. 248:443-63), peptide truncation analysis, high-resolution electron microscopy techniques using single-particle reconstruction, cryoEM, or tomography, crystallographic studies, and NMR analysis. Additionally, methods such as epitope excision, epitope extraction, and chemical modification of antigens can be used (Tomer (2000) Prot. Sci. 9:487-496). Another method that can be used to identify the amino acids in a polypeptide with which an antibody interacts is hydrogen / deuterium exchange, detected by mass spectrometry. Generally speaking, the hydrogen / deuterium exchange method involves deuterium-labeling a protein of interest and then binding an antibody to the deuterium-labeled protein. The protein / antibody complex is then transferred to water, and exchangeable protons in amino acids protected by the antibody complex undergo back-exchange from deuterium to hydrogen at a slower rate than exchangeable protons in amino acids that are not part of the interface. As a result, amino acids that form part of the protein / antibody interface may retain deuterium and therefore exhibit a relatively higher mass than amino acids that are not included in the interface. After dissociation of the antibody, the target protein is subjected to protease cleavage and mass spectrometry, thereby revealing deuterium-labeled residues corresponding to the specific amino acids with which the antibody interacts. See, for example, Ehring (1999) Analytical Biochemistry 267:252-259; Engen and Smith (2001) Anal. Chem. 73:256A-265A.

[0160] The term "epitope" refers to a site on an antigen to which B cells and / or T cells respond. B cell epitopes can be formed both from contiguous amino acids or from noncontiguous amino acids juxtaposed by tertiary folding of a protein. Epitopes formed from adjacent amino acids are typically retained upon exposure to denaturing solvents, whereas epitopes formed by tertiary folding are typically lost upon treatment with denaturing solvents. Epitopes typically contain at least three, more commonly at least five or 8-10 amino acids in a unique spatial conformation.

[0161] Modification-assisted profiling (MAP), also known as antigen structure-based antibody profiling (ASAP), is a method for classifying multiple monoclonal antibodies raised against the same antigen according to the similarity of each antibody's binding profile to chemically or enzymatically modified antigen surfaces (see U.S. Patent Application Publication No. 2004 / 0101920, specifically incorporated herein by reference in its entirety). Each category may reflect a unique epitope that is distinct from or partially overlaps with the epitopes represented by another category. This technique allows for the rapid filtering of genetically identical antibodies so that characterization can be focused on genetically distinct antibodies. When applied to hybridoma screening, MAP can facilitate the identification of rare hybridoma clones producing monoclonal antibodies with desired characteristics. MAP can be used to sort the antibodies of the present disclosure into groups of antibodies that bind to different epitopes.

[0162] The present disclosure includes antibodies that can bind to the same epitope or a part of the same epitope.By using routine methods known in the art, it can be easily determined whether an antibody binds to the same epitope as a reference antibody or competes with a reference antibody for binding.For example, to determine whether a test antibody binds to the same epitope as a reference antibody, the reference antibody is bound to a target molecule under saturating conditions.Then, the ability of the test antibody to bind to the target molecule is evaluated.If the test antibody can bind to the target molecule after saturating binding with the reference antibody, it can be concluded that the test antibody binds to a different epitope from the reference antibody.On the other hand, if the test antibody cannot bind to the target molecule after saturating binding with the reference antibody, the test antibody may bind to the same epitope as the epitope bound by the reference antibody.

[0163] To determine whether an antibody competes for binding with, for example, the 77A antibody, the above-described binding methodology is carried out in two ways: in the first way, the 77A antibody is allowed to bind to the HSP70 protein under saturating conditions, and then the binding of the test antibody to the HSP70 protein is evaluated. In the second way, the test antibody is allowed to bind to the HSP70 protein under saturating conditions, and then the binding of the 77A antibody to the HSP70 protein is evaluated. In both ways, if only the first (saturating) antibody can bind to the HSP70 molecule, it is concluded that the test antibody and the 77A antibody compete for binding to HSP70. As will be understood by those skilled in the art, an antibody that competes for binding with a reference antibody does not necessarily bind to the same epitope as the reference antibody, but may sterically block the binding of the reference antibody by binding to an overlapping or adjacent epitope.

[0164] Two antibodies bind to the same or overlapping epitopes if each competitively inhibits (blocks) the binding of the other to the antigen. That is, a 1-fold, 5-fold, 10-fold, 20-fold, or 100-fold excess of one antibody inhibits the binding of the other by at least 50%, but preferably 75%, 90%, or even 99%, as measured in a competitive binding assay (see, e.g., Junghans et al., Cancer Res. 1990 50:1495-1502). Alternatively, two antibodies have the same epitope if essentially all amino acid mutations in the antigen that reduce or eliminate binding of one antibody also reduce or eliminate binding of the other. Two antibodies have overlapping epitopes if some amino acid mutations that reduce or eliminate binding of one antibody also reduce or eliminate binding of the other.

[0165] Further routine experiments (e.g., peptide mutations and binding analysis) can then be performed to confirm whether the observed lack of binding of the test antibody is indeed due to binding to the same epitope as the reference antibody, or whether steric blocking (or another phenomenon) is responsible for the observed lack of binding. These types of experiments can be performed using ELISA, RIA, surface plasmon resonance, flow cytometry, or any other quantitative or qualitative antibody binding assay available in the art.

[0166] In another aspect, antibodies can be defined by their variable sequences, including additional "framework" regions. These are provided in Tables 2, 3, 6, 9, and 10, which represent complete variable regions. Additionally, antibody sequences can vary from these sequences, optionally using methods discussed in more detail below. For example, nucleic acid sequences can be modified in a variety of ways, including: (a) the variable regions can be separated from the light and heavy chain constant domains; (b) the nucleic acid can vary from the nucleic acid described above without affecting the residues encoded thereby; (c) the nucleic acid can vary from the nucleic acid described above by a given percentage, e.g., 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% homology; and (d) the nucleic acid can be modified by precipitation in about 0.02 M to about 0.15 M NaCl at a temperature of about 50° C. to about 70° C. (e) amino acids may vary from the above nucleic acid sequences at a given percentage, e.g., 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% homology; or (f) amino acids may vary from the above nucleic acid sequences in that they may differ from the above amino acids by allowing for conservative substitutions.

[0167] When comparing polynucleotide and polypeptide sequences, if the nucleotide or amino acid sequences in the two sequences are the same when aligned for maximum matching, as described below, the two sequences are said to be "identical." Comparison between two sequences is typically performed by comparing the sequences over a comparison window to identify and compare local regions of sequence similarity. As used herein, a "comparison window" refers to a segment of at least about 20 consecutive positions, usually 30 to about 75, 40 to about 50, within which the sequence can be compared to a reference sequence of the same number of consecutive positions after the two sequences are optimally aligned.

[0168] Optimal alignment of sequences for comparison can be performed using the Megalign program in the Lasergene suite of bioinformatics software (DNASTAR, Inc., Madison, Wis.) using default parameters. Alternatively, optimal alignment of sequences for comparison can be performed by the local identity algorithm of Smith and Waterman (1981) Add. APL. Math 2:482, by the identity alignment algorithm of Needleman and Wunsch (1970) J. Mol. Biol. 48:443, by the similarity search method of Pearson and Lipman (1988) Proc. Natl. Acad. Sci. USA 85:2444, by computerized implementations of these algorithms (GAP, BESTFIT, BLAST, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group (GCG), 575 Science Dr., Madison, Wis.), or by inspection.

[0169] Specific examples of algorithms suitable for determining percent sequence identity and sequence similarity are the BLAST and BLAST 2.0 algorithms, described in Altschul et al. (1977) Nucl. Acids Res. 25:3389-3402 and Altschul et al. (1990) J. Mol. Biol. 215:403-410, respectively. BLAST and BLAST 2.0 can be used, for example, with the parameters described herein, to determine percent sequence identity for the polynucleotides and polypeptides of the present disclosure. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information. Due to the rearranged nature of antibody sequences and the variable length of each gene, multiple BLAST searches are required for a single antibody sequence. Furthermore, manual assembly of the different genes is difficult and prone to error. The sequence analysis tool IgBLAST (available on the world wide web at ncbi.nlm.nih.gov / igblast / ) identifies matches to germline V, D, and J genes, detailing rearrangement junctions and delineating IgV domain framework regions and complementarity-determining regions. IgBLAST can analyze either nucleotide or protein sequences, can process sequences in batches, and allows simultaneous searches against germline gene databases and other sequence databases, minimizing the chance of missing potentially best-matching germline V genes.

[0170] In one approach, "percent sequence identity" is determined by comparing two optimally aligned sequences over a comparison window of at least 20 positions, where the portion of the polynucleotide or polypeptide sequence within the comparison window may contain 20% or fewer, typically 5-15%, or 10-12% additions or deletions (i.e., gaps) compared to the reference sequence (which does not contain additions or deletions) for optimal alignment of the two sequences. The percentage is calculated by determining the number of positions in both sequences where identical nucleic acid bases or amino acid residues occur to obtain the number of matched positions, dividing the number of matched positions by the total number of positions in the reference sequence (i.e., the window size), and multiplying the result by 100 to obtain the percentage sequence identity.

[0171] Yet another way to define an antibody is as a "derivative" of any of the antibodies and antigen-binding fragments thereof provided herein. Derivative antibodies or antibody fragments can be modified by chemical modification using techniques known to those skilled in the art, including, but not limited to, specific chemical cleavage, acetylation, formulation, metabolic synthesis of tunicamycin, and the like. In one embodiment, an antibody derivative has similar or identical function as the parent antibody. In another embodiment, an antibody derivative exhibits altered activity compared to the parent antibody. For example, a derivative antibody (or fragment thereof) can bind to its epitope more tightly than the parent antibody or be more resistant to proteolysis.

[0172] The term "derivative" refers to an antibody or antigen-binding fragment thereof that immunospecifically binds to an antigen but contains one, two, three, four, five, or more amino acid substitutions, additions, deletions, or modifications compared to the "parent" (or wild-type) molecule. Such amino acid substitutions or additions may introduce naturally occurring (i.e., DNA-encoded) or non-naturally occurring amino acid residues. The term "derivative" encompasses variants with altered CH1, hinge, CH2, CH3, or CH4 regions, e.g., to form antibodies with variant Fc regions that exhibit enhanced or impaired effector or binding properties. The term "derivative" further encompasses non-amino acid modifications, e.g., glycosylation (e.g., having altered mannose, 2-N-acetylglucosamine, galactose, fucose, glucose, sialic acid, 5-N-acetylneuraminic acid, 5-glycolneuraminic acid, etc. content), acetylation, pegylation, phosphorylation, amidation, amino acids that can be derivatized with known protecting / blocking groups, proteolytic cleavage, linked to cellular ligands or other proteins, etc. In some embodiments, the altered carbohydrate modification modulates one or more of the following: antibody solubilization, promoting intracellular antibody trafficking and secretion, promoting antibody assembly, conformational integrity, and antibody-mediated effector function. In certain embodiments, the altered carbohydrate modification enhances antibody-mediated effector function compared to an antibody lacking the carbohydrate modification. Carbohydrate modifications that result in altered antibody-mediated effector function are well known in the art.

[0173] The biophysical properties of antibodies can be determined. High temperatures can be used to unfold the antibody and determine its relative stability using the average apparent melting temperature. Differential scanning calorimetry (DSC) measures the heat capacity, C, of ​​the molecule as a function of temperature. p DSC measures the heat required to warm a molecule per degree. DSC can be used to study the thermal stability of antibodies. DSC data for mAbs resolves the unfolding of individual domains within the mAb structure, and is shown in the thermogram (Fab, C). H2 and C HThis is particularly interesting because it can produce up to three peaks (from the unfolding of three domains). Typically, unfolding of the Fab domain results in the most intense peak. The DSC profile and relative stability of the Fc portion show characteristic differences for human IgG1, IgG2, IgG3, and IgG4 subclasses (Garber and Demarest, Biochem. Biophys. Res. Commun. 355, 751-757, 2007). Circular dichroism (CD) performed on a CD spectrometer can also be used to determine the average apparent melting temperature. Far-UV CD spectra are measured for antibodies in the range of 200 nm to 260 nm with 0.5 nm increments. The final spectrum can be determined as the average of 20 accumulations. Residual ellipticity values ​​can be calculated after background subtraction. Thermal unfolding of an antibody (0.1 mg / mL) can be monitored at 235 nm from 25 to 95 °C and a heating rate of 1 °C / min. Dynamic light scattering (DLS) can be used to assess aggregation propensity. DLS is used to characterize the size of various particles, including proteins. If the system is not size-dispersed, the average effective diameter of the particles can be determined. This measurement depends on the size of the particle core, the size of the surface structure, and the particle concentration. DLS essentially measures the fluctuations in scattered light intensity by the particles, allowing the particle's diffusion coefficient to be determined. DLS software in commercially available DLS instruments indicates particle populations of different diameters. Stability studies can be conveniently performed using DLS. DLS measurements of a sample can indicate whether particles aggregate over time or with temperature fluctuations by determining whether the hydrodynamic radius of the particles increases. If particles aggregate, a larger population of particles with a larger radius can be observed. Temperature-dependent stability can be analyzed by controlling the temperature in situ. Capillary electrophoresis (CE) techniques include a proven methodology for characterizing antibody stability.The iCE approach can be used to separate antibody protein charge variants resulting from deamidation, C-terminal lysine, sialylation, oxidation, glycosylation, and any other alterations to the protein that may result in a change in the protein's pI. Each expressed antibody protein can be evaluated by high-throughput free-solution isoelectric focusing (IEF) (cIEF) in a capillary column using the Protein Simple Maurice instrument. Whole-column UV absorbance detection can be performed every 30 seconds for real-time monitoring of molecules focusing at their isoelectric point (pI). This approach combines the high resolution of traditional gel IEF with the quantification and automation benefits found in column-based separations while eliminating the need for a transfer step. This technique provides reproducible quantitative analysis of the identity, purity, and heterogeneity profile of expressed antibodies. Results identify charge heterogeneity and molecular size on the antibody in both absorbance and intrinsic fluorescence detection modes, with a detection sensitivity down to 0.7 μg / mL.

[0174] The intrinsic solubility score of an antibody sequence can be determined. This can be calculated using CamSol Intrinsic (Sormanni et al., J Mol Biol 427, 478-490, 2015). The amino acid sequence of residues 95-102 (Kabat numbering) in the HCDR3 of each antibody fragment, such as an scFv, can be evaluated via an online program to calculate the solubility score. Laboratory techniques can also be used to determine solubility. Various techniques exist, including adding lyophilized protein to a solution until the solution is saturated and the solubility limit is reached, or concentrating by ultrafiltration in a microconcentrator with an appropriate molecular weight cutoff. The simplest method is the induction of an amorphous precipitate, which involves protein precipitation using ammonium sulfate (Trevino et al., J Mol Biol 366:449-460, 2007). Ammonium sulfate precipitation provides rapid and accurate information on relative solubility values. Ammonium sulfate precipitation produces a precipitation solution with clearly separated aqueous and solid phases and requires a relatively small amount of protein. Solubility measurements performed using ammonium sulfate induction of amorphous precipitates can also be easily performed at different pH values. Protein solubility is highly pH-dependent, and pH is considered the most important extrinsic factor affecting solubility.

[0175] H. Specific Aspects In one embodiment, a heavy chain variable region (VH) comprising: a VHCDR1 amino acid sequence of GYX1FTX2YG (SEQ ID NO: 214), wherein X1 is T, S, or I, and X2 is N or K; a VHCDR2 amino acid sequence of INTYTGEX1 (SEQ ID NO: 215), wherein X1 is P, S, T, or A; and a VHCDR3 amino acid sequence of X1RYDHX2MDY (SEQ ID NO: 216), wherein X1 is A, T, V, or G, and X2 is A, R, F, T, P, V, S, D, N, H, L, Y, or G; and / or a VLCDR1 amino acid sequence of QSLX1NSGTRKNY (SEQ ID NO: 212), wherein X1 is L, F, or V; and a VLCDR1 amino acid sequence of SEQ ID NO: 213. Provided herein is a monoclonal antibody or antibody fragment comprising a light chain variable region (VL) comprising the VLCDR2 amino acid sequence of NO:5 and the VLCDR3 amino acid sequence of KQSYX1LYT (SEQ ID NO:213), wherein X1 is T, N, or S.

[0176] In one aspect, provided herein is an antibody or antibody fragment comprising a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence selected from the group consisting of SEQ ID NO:1 and 164-166, a VHCDR2 amino acid sequence selected from the group consisting of SEQ ID NO:2 and 167-169, and a VHCDR3 amino acid sequence selected from the group consisting of SEQ ID NO:3 and 170-185; and / or a light chain variable region (VL) comprising a VLCDR1 amino acid sequence selected from the group consisting of SEQ ID NO:4 and 159-161, a VLCDR2 amino acid sequence of SEQ ID NO:5, and a VLCDR3 amino acid sequence selected from the group consisting of SEQ ID NO:6, 162, and 163.

[0177] In one aspect, provided herein is an antibody or antibody fragment, wherein said antibody or antibody fragment comprises: (i) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (ii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:164, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (iii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (iv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:162; (v) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:171; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (vi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:172; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (vii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:159, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (viii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:173; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (ix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (x) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 164, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 175; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:163; (xii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:176; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (xiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 171; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 159, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 177; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 164, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 178; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 179; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 179; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 159, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 164, the VHCDR2 amino acid sequence of SEQ ID NO: 167, and the VHCDR3 amino acid sequence of SEQ ID NO: 174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 180; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xx) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 181; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (xxi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 182; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 181; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 168, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 165, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 185; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 166, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 164, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 165, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 169, and the VHCDR3 amino acid sequence of SEQ ID NO: 174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 160, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxx) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (xxxi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:175; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (xxxii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 167, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 169, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 166, the VHCDR2 amino acid sequence of SEQ ID NO: 167, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 168, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 166, the VHCDR2 amino acid sequence of SEQ ID NO: 168, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 161, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 1, the VHCDR2 amino acid sequence of SEQ ID NO: 168, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 161, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 166, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xl) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:2, and the VHCDR3 amino acid sequence of SEQ ID NO:178; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:163; (xli) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 164, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (xlii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 166, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 161, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xliii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 165, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xliv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 166, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 4, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 165, the VHCDR2 amino acid sequence of SEQ ID NO: 2, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 161, the VLCDR2 amino acid sequence of SEQ ID NO: 5, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; or (xlvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:1, the VHCDR2 amino acid sequence of SEQ ID NO:168, and the VHCDR3 amino acid sequence of SEQ ID NO:183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:4, the VLCDR2 amino acid sequence of SEQ ID NO:5, and the VLCDR3 amino acid sequence of SEQ ID NO:6.

[0178] In one aspect, provided herein is an antibody or antibody fragment comprising a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence selected from the group consisting of SEQ ID NOs: 192-195, a VHCDR2 amino acid sequence selected from the group consisting of SEQ ID NOs: 196-211, and a VHCDR3 amino acid sequence selected from the group consisting of SEQ ID NOs: 3 and 170-185, and / or a light chain variable region (VL) comprising a VLCDR1 amino acid sequence selected from the group consisting of SEQ ID NOs: 186-190, a VLCDR2 amino acid sequence of SEQ ID NO: 191, and a VLCDR3 amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 162, and 163.

[0179] In one aspect, provided herein is an antibody or antibody fragment, wherein said antibody or antibody fragment comprises: (i) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:192, the VHCDR2 amino acid sequence of SEQ ID NO:196, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:186, the VLCDR2 amino acid sequence of SEQ ID NO:191, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (ii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:193, the VHCDR2 amino acid sequence of SEQ ID NO:197, and the VHCDR3 amino acid sequence of SEQ ID NO:3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:186, the VLCDR2 amino acid sequence of SEQ ID NO:191, and the VLCDR3 amino acid sequence of SEQ ID NO:6; (iii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (iv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 198, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (v) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (vi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (vii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 171; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (viii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 172; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (ix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (x) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 173; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 199, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 192, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 175; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 200, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 201, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 201, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 188, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 163; (xviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 189, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 176; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xx) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 171; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 173; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 197, and the VHCDR3 amino acid sequence of SEQ ID NO: 177; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 192, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 178; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 179; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 179; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 192, the VHCDR2 amino acid sequence of SEQ ID NO: 202, and the VHCDR3 amino acid sequence of SEQ ID NO: 174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 180; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 201, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (xxix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 181; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (xxx) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 182; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 163; (xxxiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 181; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 203, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 194, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 185; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 189, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 197, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 204, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 192, the VHCDR2 amino acid sequence of SEQ ID NO: 205, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xxxix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 206, and the VHCDR3 amino acid sequence of SEQ ID NO: 171; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xl) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 194, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xli) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 207, and the VHCDR3 amino acid sequence of SEQ ID NO: 174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 190, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xliii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 206, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xliv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 175; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 189, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 202, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 207, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 202, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (xlix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 208, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (l) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 209, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (li) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 209, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 206, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (liii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 210, and the VHCDR3 amino acid sequence of SEQ ID NO: 178; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 163; (liv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 192, the VHCDR2 amino acid sequence of SEQ ID NO: 197, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (lv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 210, and the VHCDR3 amino acid sequence of SEQ ID NO: 174; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 188, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 209, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lviii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 188, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lix) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 194, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lx) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 188, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lxi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 192, the VHCDR2 amino acid sequence of SEQ ID NO: 211, and the VHCDR3 amino acid sequence of SEQ ID NO: 170; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lxii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 210, and the VHCDR3 amino acid sequence of SEQ ID NO: 172; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 189, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lxiii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 195, the VHCDR2 amino acid sequence of SEQ ID NO: 196, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lxiv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 194, the VHCDR2 amino acid sequence of SEQ ID NO: 206, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 162; (lxv) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 208, and the VHCDR3 amino acid sequence of SEQ ID NO: 183; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 186, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; (lxvi) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO: 193, the VHCDR2 amino acid sequence of SEQ ID NO: 199, and the VHCDR3 amino acid sequence of SEQ ID NO: 3; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO: 187, the VLCDR2 amino acid sequence of SEQ ID NO: 191, and the VLCDR3 amino acid sequence of SEQ ID NO: 6; or (lxvii) a heavy chain variable region (VH) comprising the VHCDR1 amino acid sequence of SEQ ID NO:193, the VHCDR2 amino acid sequence of SEQ ID NO:199, and the VHCDR3 amino acid sequence of SEQ ID NO:184; and / or a light chain variable region (VL) comprising the VLCDR1 amino acid sequence of SEQ ID NO:186, the VLCDR2 amino acid sequence of SEQ ID NO:191, and the VLCDR3 amino acid sequence of SEQ ID NO:6.

[0180] In some aspects, the antibody or antibody fragment comprises: 1. A heavy chain variable sequence having the sequence of TIFF2024536133000002.tif19159, wherein X1 is Q or E, X2 is I or V, X3 is V or Q, X4 is Q or E, X5 is A, P or G, X6 is E or G, X7 is V or L, X8 is V or K, and X9 is A, E, G or S; 10 is V or L, and X 11 is K or R, and X 12 is V, L or I, and X 13 is A or T, and X14 is K or Q, and X 15 is E or K, and X 16 is M or V, and X 17 is F or V, and X 18 is F, M or I, and X 19 is T or S, and X 20 is T, R or A, and X 21 is T, D or E, and X 22 is T, A or K, and X 23 is S or N, and X 24 is L or A, and X 25 is M or L, and X 26 is E or Q, and X 27 is L or M, and X 28 is R, S, T or N, and X 29 is S or G, and X 30 is R, K or M, and X 31 is S or T, and X 32 is D or E, and X 33 is L, S, or T; and / or TIFF2024536133000003.tif19159, wherein X1 is E or D, X2 is I or V, X3 is V or Q, X4 is L or M, X5 is D or S, X6 is A or S, X7 is V or A, X8 is L or V, X9 is E or D, and X 10 is A or V, and X 11 is N or T, and X 12 is A or P, and X 13 is Q or K, and X 14 is S, V or P, and X 15 is K or R, and X 16 is D or S, and X 17 is S, D or N, and X 18 is S or T, and X 19 is A or P, and X 20 is V or T, and X21 is Q or G, and X 22 is L or V, Includes.

[0181] In some aspects, the antibody or antibody fragment comprises: 1. A heavy chain variable sequence having the sequence of TIFF2024536133000004.tif28159, wherein X1 is Q or H, X2 is A, D, T, V, S or P, X3 is T, S or I, X4 is N or K, X5 is P, S, T or A, X6 is T, R, K or I, X7 is A, T, V, S or G, X8 is S, R or T, X9 is A, V or G, and X 10 is E or D, and X 11 is L or V, and X 12 is A, T, V or G, and X 13 is A, R, F, T, P, V, S, D, N, H, L, Y or G, and X 14 is T or S; and / or 1. A light chain variable sequence having the sequence of TIFF2024536133000005.tif21159, wherein X1 is A, T, or S, X2 is N or K, X3 is L, F, or V, X4 is A, S, or T, X5 is Q or K, X6 is A, P, or S, X7 is K or N, X8 is L, V, or I, X9 is G or A, and X 10 is T or S, and X 11 is S or R, and X 12 is A or T, and X 13 is V, I or L, and X 14 is T, N or S, Includes.

[0182] In some aspects, the antibody or antibody fragment comprises a heavy chain variable sequence having a sequence selected from the group consisting of SEQ ID NOs: 7, 12-17, 26-103, and 225-229, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to any one of SEQ ID NOs: 7, 12-17, 26-103, and 225-229; and / or a light chain variable sequence having a sequence selected from the group consisting of SEQ ID NOs: 8, 19-24, 105-157, and 230-234, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to any one of SEQ ID NOs: 7, 12-17, 26-103, and 225-229; and a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to any one of NOs:8, 19-24, 105-157 and 230-234.

[0183] In some aspects, the antibody or antibody fragment comprises: (i) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:7, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:8, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:8; (ii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:12, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:19, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:19; (iii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:12, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:20, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:20; (iv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:12, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:21, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:21; (v) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:12, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:22, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:22; (vi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:12, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:23, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:23; (vii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:12, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:24, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:24; (viii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:13, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:13; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:19, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:19; (ix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:13, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:13; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:20, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:20; (x) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:13, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:13; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:21, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:21; (xi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:13, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:13; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:22, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:22; (xii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:13, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:13; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:23, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:23; (xiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:13, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:13; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:24, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:24; (xiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:14, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:14; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:19, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:19; (xv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:14, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:14; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:20, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:20; (xvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:14, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:14; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:21, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:21; (xvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:14, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:14; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:22, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:22; (xviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:14, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:14; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:23, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:23; (xix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:14, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:14; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:24, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:24; (xx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:15, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:15; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:19, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:19; (xxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:15, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:15; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:20, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:20; (xxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 15, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 15; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 21, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 21; (xxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 15, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 15; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 22, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 22; (xxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 15, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 15; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 23, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 23; (xxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:15, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:15; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:24, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:24; (xxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 16, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 16; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 19, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 19; (xxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 16, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 16; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 20, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 20; (xxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 16, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 16; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 21, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 21; (xxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 16, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 16; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 22, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 22; (xxx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:16, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:16; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:23, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:23; (xxxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:16, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:16; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:24, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:24; (xxxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:17, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:17; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:19, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:19; (xxxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:17, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:17; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:20, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:20; (xxxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:17, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:17; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:21, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:21; (xxxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:17, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:17; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:22, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:22; (xxxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO: 17, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 17; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO: 23, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO: 23; (xxxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:17, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:17; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:24, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:24; (xxxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:26, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:26; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xxxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:27, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:27; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xl) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:28, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:28; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xli) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:29, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:29; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xlii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:30, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:30; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:106, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:106; (xliii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:31, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:31; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xliv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:32, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:32; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xlv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:30, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:30; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:107, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:107; (xlvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:33, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:33; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xlvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:34, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:34; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xlviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:30, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:30; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:108, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:108; (xlix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:30, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:30; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:109, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:109; (l) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:35, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:35; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (li) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:36, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:36; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:37, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:37; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (liii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:26, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:26; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:107, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:107; (liv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:38, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:38; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:31, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:31; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:110, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:110; (lvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:39, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:39; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:40, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:40; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:34, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:34; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:111, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:111; (lix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:41, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:41; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:109, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:109; (lx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:30, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:30; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:112, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:112; (lxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:28, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:28; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:113, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:113; or (lxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:32, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:32; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:114, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:114; (lxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:42, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:42; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:36, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:36; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:115, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:115; (lxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:43, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:43; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:32, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:32; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:109, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:109; (lxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:44, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:44; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:116, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:116; (lxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:35, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:35; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:117, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:117; (lxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:45, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:45; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:46, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:46; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:36, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:36; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:118, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:118; (lxxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:47, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:47; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:115, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:115; (lxxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:48, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:48; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:109, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:109; (lxxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:49, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:49; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:50, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:50; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:51, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:51; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:106, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:106; (lxxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:52, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:52; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:119, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:119; (lxxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:53, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:53; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:108, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:108; (lxxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:54, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:54; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (lxxx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:55, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:55; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:116, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:116; (lxxxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:56, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:56; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:116, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:116; (lxxxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:57, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:57; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:120, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:120; (lxxxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:58, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:58; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:121, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:121; (lxxxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:59, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:59; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:122, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:122; (lxxxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:60, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:60; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:108, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:108; (lxxxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:61, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:61; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:123, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:123; (lxxxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:62, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:62; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:114, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:114; (lxxxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:63, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:63; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:124, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:124; (lxxxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:64, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:64; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xc) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:65, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:65; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:125, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:125; (xci) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:66, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:66; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:105; (xcii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:67, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:67; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:125, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:125; (xciii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:68, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:68; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:126, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:126; (xciv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:69, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:69; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:127, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:127; (xcv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:70, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:70; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:128, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:128; (xcvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:71, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:71; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:117, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:117; (xcvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:72, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:72; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:129, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:129; (xcviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:73, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:73; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:130, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:130; (xcix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:74, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:74; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:131, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:131; (c) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:73, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:73; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:132, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:132; (ci) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:75, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:75; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:133, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:133; (cii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:76, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:76; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:134, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:134; (ciii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:77, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:77; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:107, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:107; (civ) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:78, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:78; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:135, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:135; (cv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:79, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:79; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:136, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:136; (cvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:80, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:80; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:137, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:137; (cvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:41, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:41; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:138, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:138; (cviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:81, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:31; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:139, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:139; (cix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:82, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:82; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:105, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:105; (cx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:83, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:83; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:126, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:126; (cxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:84, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:84; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:140, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:140; (cxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:85, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:85; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:141, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:141; (cxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:86, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:86; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:141, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:141; (cxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:87, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:87; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:117, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:117; (cxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:88, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:88; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:142, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:142; (cxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:89, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:89; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:143, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:143; (cxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:90, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:90; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:144, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:144; (cxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:91, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:91; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:109, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:109; (cxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:92, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:92; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:145, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:145; (cxx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:93, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:93; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:146, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:146; (cxxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:94, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:94; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:147, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:147; (cxxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:95, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:95; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:148, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:148; (cxxiii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:96, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:96; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:149, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:149; (cxxiv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:97, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:97; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:150, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:150; (cxxv) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:98, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:98; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:151, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:151; (cxxvi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:99, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:99; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:152, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:152; (cxxvii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:100, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:100; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:136, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:136; (cxxviii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:91, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:91; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:153, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:153; (cxxix) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:101, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:101; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:154, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:154; (cxxx) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:102, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:102; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:155, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:155; (cxxxi) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:36, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:36; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:156, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:156; or (cxxxii) a heavy chain variable sequence having the sequence set forth in SEQ ID NO:103, or a heavy chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:103; and / or a light chain variable sequence having the sequence set forth in SEQ ID NO:157, or a light chain variable sequence having at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity to SEQ ID NO:157.

[0184] In some aspects, the antibody or antibody fragment comprises: (a) an immunoglobulin heavy chain variable region comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO:1, a VHCDR2 comprising the amino acid sequence of SEQ ID NO:2, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO:3, wherein the VHCDR1 has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:12; (b) an immunoglobulin light chain variable region comprising a VLCDR1 comprising the amino acid sequence of SEQ ID NO:4, a VLCDR2 comprising the amino acid sequence of SEQ ID NO:5, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO:6, wherein the VLCDR1 has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:19; Includes.

[0185] In one embodiment, provided herein is an antibody or antibody fragment that competes for binding to the same epitope on HSP70 as an antibody or antibody fragment according to any one of the present embodiments. In one embodiment, provided herein is an antibody or antibody fragment that binds to, or is capable of binding to, the epitope on HSP70 recognized by the antibody or antibody fragment of any one of the present embodiments.

[0186] In one embodiment, provided herein is an antibody or antibody fragment, wherein the antibody or antibody fragment binds to an epitope of HSP70 defined by a peptide corresponding to K573 to Q601 of SEQ ID NO:11. In some aspects, when bound to HSP70, the antibody or antibody fragment binds to one or two of the following residues: H594, K595, and Q601 of SEQ ID NO:11. In some aspects, when bound to HSP70, the antibody or antibody fragment binds to all of the following residues: H594, K595, and Q601 of SEQ ID NO:11. In some aspects, when bound to HSP70, the antibody or antibody fragment further binds to at least one of the following residues: K573, E576, W580, R596, and E598 of SEQ ID NO:11. In some aspects, when bound to HSP70, the antibody or antibody fragment further binds to at least two, at least three, at least four, or at least five of the following residues: K573, E576, W580, R596, and E598 of SEQ ID NO: 11. In some aspects, when bound to HSP70, the antibody or antibody fragment binds to all of the following residues: K573, E576, W580, H594, K595, R596, E598, and Q601 of SEQ ID NO: 11. In some aspects, when bound to HSP70, the antibody or antibody fragment binds to an epitope of HSP70 comprising KEWHKREQ (SEQ ID NO:250).

[0187] In some aspects of any of the present embodiments, the antibody binds or is capable of binding to HSP70. In some aspects of any of the present embodiments, the antibody binds or is capable of binding to HSP70 in an ADP-bound form. In some aspects of any of the present embodiments, the antibody binds or is capable of binding to HSP70 in a peptide-bound form. In some aspects of any of the present embodiments, the antibody binds or is capable of binding to HSP70 in an ADP-bound and peptide-bound form. In some aspects of any of the present embodiments, the antibody exhibits altered, e.g., enhanced, antibody-dependent cellular cytotoxicity. In some aspects of any of the present embodiments, the antibody exhibits altered, e.g., enhanced, complement-dependent cytotoxicity. In some aspects of any of the present embodiments, the antibody enhances HSP70 uptake by immune effector cells, such as monocytes / macrophages and dendritic cells. In some aspects, uptake is mediated by human FcγR1, human FcγR2a, human FcγR2b, human FcγR2c, human FcγR3a and / or human FcγR3b. In some aspects, the antibody binds or is capable of binding to HSP70. In some aspects, the antibody has a denaturing activity of less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 9 nM, less than about 8 nM, less than about 7 nM, less than about 6 nM, less than about 5 nM, less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM, less than about 0.9 nM, less than about 0.85 nM, less than about 0.8 nM, less than about 0.75 nM, less than about 0.7 nM, or less than about 0.8 nM, as determined by surface plasmon resonance or Octet Biolayer Interferometry (BLI) analysis. K of less than about nM, less than about 0.6 nM, less than about 0.5 nM, less than about 0.1 nM, less than about 0.05 nM, or less than about 20 pM, less than about 10 pM, less than about 9 pM, less than about 8 pM, less than about 7 pM, less than about 6 pM, less than about 5 pM, less than about 4 pM, less than about 3 pM, less than about 2 pM, less than about 1 pM, less than about 0.9 pM, less than about 0.85 pM, less than about 0.8 pM, less than about 0.75 pM, less than about 0.7 pM, less than about 0.6 pM, less than about 0.5 pM, less than about 0.1 pM, or less than about 0.05 pM Dand binds to human HSP70 (e.g., HSP70 in an ADP-bound and / or peptide-bound form). In some aspects, the antibody has a ribonucleotide binding activity of about 50 nM to about 0.05 nM, about 50 nM to about 0.075 nM, about 50 nM to about 0.1 nM, about 50 nM to about 0.5 nM, about 50 nM to about 1 nM, about 40 nM to about 0.05 nM, about 40 nM to about 0.075 nM, about 40 nM to about 0.1 nM, or about 40 nM to about 0.5 nM, as determined by surface plasmon resonance or Octet biolayer interferometry (BLI) analysis. , about 40nM to about 1nM, about 30nM to about 0.05nM, about 30nM to about 0.075nM, about 30nM to about 0.1nM, about 30nM to about 0.5nM, about 30nM to about 1nM, about 20nM to about 0.05nM , about 20nM to about 0.075nM, about 20nM to about 0.1nM, about 20nM to about 0.5nM, about 20nM to about 1nM, about 10nM to about 0.05nM, about 10nM to about 0.075nM, about 10nM to about 0.1 nM, about 10nM to about 0.5nM, about 10nM to about 1nM, about 5nM to about 0.05nM, about 5nM to about 0.075nM, about 5nM to about 0.1nM, about 5nM to about 0.5nM, about 5nM to about 1nM, about 3n M ~ about 0.05nM, about 3nM to about 0.075nM, about 3nM to about 0.1nM, about 3nM to about 0.5nM, about 3nM to about 1nM, about 3nM to about 2nM, about 2nM to about 0.05nM, about 2nM to about 0.075 nM, about 2nM to about 0.1nM, about 2nM to about 0.5nM, about 2nM to about 1nM, about 1nM to about 0.05nM, about 1nM to about 0.075nM, about 1nM to about 0.1nM, about 1nM to about 0.5nM, about 0.5 K of nM to about 0.05nM, about 0.5nM to about 0.075nM, about 0.5nM to about 0.1nM, about 0.1nM to about 0.05nM, about 0.1nM to about 0.075nM or about 0.075nM to about 0.05nM Dand binds to human HSP70 (e.g., ADP-bound and / or peptide-bound forms of HSP70). In some aspects, the antibody has a saturation level of about 20 pM to about 0.05 pM, about 20 pM to about 0.075 pM, about 20 pM to about 0.1 pM, about 20 pM to about 0.5 pM, about 20 pM to about 1 pM, about 10 pM to about 0.05 pM, about 10 pM to about 0.075 pM, about 10 pM to about 0.1 pM, about 10 pM to about 0.5 pM, about 10 pM to about 1 pM, about 5 pM to about 0.05 pM, about 5 pM to about 0.075 pM, about 5 pM to about 0.1 pM, about 5 pM to about 0.5 pM, about 5 pM to about 1 pM, about 3 pM to about 0.05 pM, as determined by surface plasmon resonance or Octet Biolayer Interferometry (BLI) analysis. M, about 3pM to about 0.075pM, about 3pM to about 0.1pM, about 3pM to about 0.5pM, about 3pM to about 1pM, about 3pM to about 2pM, about 2pM to about 0 .05pM, about 2pM to about 0.075pM, about 2pM to about 0.1pM, about 2pM to about 0.5pM, about 2pM to about 1pM, about 1pM to about 0.05pM, Approximately 1pM to approximately 0.075pM, approximately 1pM to approximately 0.1pM, approximately 1pM to approximately 0.5pM, approximately 0.5pM to approximately 0.05pM, approximately 0.5pM to approximately 0.075p M, K of about 0.5pM to about 0.1pM, about 0.1pM to about 0.05pM, about 0.1pM to about 0.075pM or about 0.075pM to about 0.05pM D and binds to human HSP70 (e.g., ADP-bound and / or peptide-bound forms of HSP70).

[0188] In some aspects, an antibody forms or is capable of forming a higher order complex upon binding to HSP70. As used herein, the term "higher order complex" refers to any complex other than a 1:1 antibody:HSP70 complex (e.g., a 1:2, 2:1, 2:2, 2:4, 5:10, or 6:12 complex). The antibody:HSP70 complex may be, for example, at least about 250 kDa (or greater than about 250 kDa), at least about 290 kDa (or greater than about 290 kDa), at least about 300 kDa (or greater than about 300 kDa), at least about 400 kDa (or greater than about 400 kDa), at least about 500 kDa (or greater than about 500 kDa), at least about 580 kDa (or greater than about 580 kDa). a), at least about 600 kDa (or more than about 600 kDa), at least about 700 kDa (or more than about 700 kDa), at least about 800 kDa (or more than about 800 kDa), at least about 900 kDa (or more than about 900 kDa), at least about 1,000 kDa (or more than about 1,000 kDa), at least about 1,100 kDa (or more than about 1,100 kDa), at least at least about 1,200 kDa (or greater than about 1,200 kDa), at least about 1,300 kDa (or greater than about 1,300 kDa), at least about 1,400 kDa (or greater than about 1,400 kDa), at least about 1,450 kDa (or greater than about 1,450 kDa), at least about 1,500 kDa (or greater than about 1,500 kDa), at least about 1,600 kDa (or greater than about 1,600 kDa), 1,000 kDa), at least about 1,700 kDa (or greater than about 1,700 kDa), at least about 1,750 kDa (or greater than about 1,750 kDa), at least about 1,800 kDa (or greater than about 1,800 kDa), at least about 1,900 kDa (or greater than about 1,900 kDa), or at least about 2,000 kDa (or greater than about 2,000 kDa).The antibody:HSP70 complex can have a molecular weight of, for example, about 250 kDa, about 290 kDa, about 300 kDa, about 400 kDa, about 500 kDa, about 580 kDa, about 600 kDa, about 700 kDa, about 800 kDa, about 900 kDa, about 1,000 kDa, about 1,100 kDa, about 1,200 kDa, about 1,300 kDa, about 1,400 kDa, about 1,450 kDa, about 1,500 kDa, about 1,600 kDa, about 1,700 kDa, about 1,750 kDa, about 1,800 kDa, about 1,900 kDa, or about 2,000 kDa. In one aspect, the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:2. For example, the antibody:HSP70 complex may contain about 1 antibody molecule and about 2 HSP70 molecules; about 2 antibody molecules and about 4 HSP70 molecules; about 3 antibody molecules and about 6 HSP70 molecules; about 4 antibody molecules and about 8 HSP70 molecules; about 5 antibody molecules and about 10 HSP70 molecules; or about 6 antibody molecules and about 12 HSP70 molecules. The formation of higher order complexes can be measured by any method known in the art, including, for example, size exclusion chromatography, as described in Example 18 herein.

[0189] In some aspects, the antibody:HSP70 complexes provided herein (e.g., complexes comprising 77A or an Fc variant thereof) activate immune effector cells, e.g., human immune effector cells. In some embodiments, the immune effector cells, e.g., human immune effector cells, include CD8+ T cells, CD4+ T cells, NK cells, and dendritic cells, e.g., immature dendritic cells.

[0190] In some embodiments, an antibody:HSP70 complex provided herein (e.g., a complex comprising 77A or an Fc variant thereof) activates CD8+ T cells. In certain embodiments, such activation of CD8+ T cells increases cytokine synthesis and secretion of activated Th-1 biased cytokines, e.g., cytokines in Table 18, in CD8+ T cells treated with the complex compared to control expression (e.g., untreated T cells). In certain embodiments, such activation of CD8+ T cells increases cytokine synthesis and secretion of activated Th-1 biased cytokines, e.g., IL-12, IFB-γ, IFN-α, IL-8, G-CSF, GM-CSF, IFN-α2, and TNF-α, in CD8+ T cells treated with the complex compared to control expression (e.g., untreated T cells). In some embodiments, cytokine secretion is increased by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%, at least 400%, or at least 500% compared to a control (e.g., untreated cells). Methods for quantifying cytokine secretion are known in the art and include, but are not limited to, cytokine arrays.

[0191] In some embodiments, an antibody:HSP70 complex provided herein (e.g., a complex comprising 77A or an Fc variant thereof) activates CD4+ T cells. In certain embodiments, such activation of CD4+ T cells increases cytokine synthesis and secretion of activation cytokines, e.g., the cytokines in Table 20, in CD4+ T cells treated with the complex compared to control expression (e.g., untreated T cells). In certain embodiments, such activation of CD4+ T cells increases cytokine synthesis and secretion of activation cytokines, e.g., IL-12, IL-10, IL-17A, IL-2, IL-8, IFN-γ, and IL-1b, in CD4+ T cells compared to control expression (e.g., untreated T cells). In some embodiments, cytokine secretion is increased by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%, at least 400%, or at least 500% compared to a control (e.g., untreated cells). Methods for quantifying cytokine secretion are known in the art and include, but are not limited to, cytokine arrays.

[0192] In some embodiments, an antibody:HSP70 complex provided herein (e.g., a complex comprising 77A or an Fc variant thereof) activates cytokine release in NK cells. In certain embodiments, such activation of NK cells increases cytokine synthesis and secretion of activating cytokines, e.g., the cytokines in Table 19, in NK cells treated with the complex compared to control expression (e.g., untreated NK cells). In certain embodiments, such activation of NK cells increases cytokine synthesis and secretion of activating cytokines, e.g., IL-2, IL-2Ra, and M-CSF, in NK cells compared to control expression (e.g., untreated NK cells). In certain embodiments, cytokine secretion is increased by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%, at least 400%, or at least 500% compared to a control (e.g., untreated cells). Methods for quantifying cytokine secretion are known in the art and include, for example, but are not limited to, cytokine arrays.

[0193] In some embodiments, the antibody:HSP70 complexes provided herein (e.g., complexes comprising 77A or an Fc variant thereof) activate dendritic cells, e.g., immature and / or mature dendritic cells. In certain embodiments, such activation of dendritic cells, e.g., immature dendritic cells, increases the expression of activation markers, e.g., CD83 on CD11c dendritic cells, compared to control expression (e.g., untreated dendritic cells, e.g., immature dendritic cells). In certain embodiments, the expression of CD83 is increased by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%, at least 400%, or at least 500% compared to a control (e.g., untreated cells).

[0194] In some aspects, the antibody (alone or in complex with HSP70) binds or is capable of binding to a human FcγR (e.g., FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b). In some aspects, the antibody (alone or in complex with HSP70, e.g., HSP70 in its ADP-bound and / or peptide-bound form) has a binding affinity of less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 9 nM, less than about 8 nM, less than about 7 nM, less than about 6 nM, less than about 5 nM, less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM, less than about 0.9 nM, less than about 0.85 nM, as determined by surface plasmon resonance or Octet biolayer interferometry (BLI) analysis. K of less than about 0.8 nM, less than about 0.75 nM, less than about 0.7 nM, less than about 0.6 nM, less than about 0.5 nM, less than about 0.1 nM, less than about 0.05 nM or less than about 20 pM, less than about 10 pM, less than about 9 pM, less than about 8 pM, less than about 7 pM, less than about 6 pM, less than about 5 pM, less than about 4 pM, less than about 3 pM, less than about 2 pM, less than about 1 pM, less than about 0.9 pM, less than about 0.85 pM, less than about 0.8 pM, less than about 0.75 pM, less than about 0.7 pM, less than about 0.6 pM, less than about 0.5 pM, less than about 0.1 pM or less than about 0.05 pM Dand binds to human FcγR (e.g., FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a, and / or FcγR3b) at a specific cleavage site. In some aspects, the antibody (alone or in complex with HSP70, e.g., HSP70 in ADP-bound and / or peptide-bound form) has a cleavage site of about 50 nM to about 0.05 nM, about 50 nM to about 0.075 nM, about 50 nM to about 0.1 nM, about 50 nM to about 0.5 nM, about 50 nM to about 1 nM, or about 40 nM to about 0.05 nM, as determined by surface plasmon resonance or Octet biolayer interferometry (BLI) analysis. M, about 40nM to about 0.075nM, about 40nM to about 0.1nM, about 40nM to about 0.5nM, about 40nM to about 1nM, about 30nM to about 0.05nM, about 30nM to about 0.075nM, about 30nM to about 0.1nM, about 3 0nM to about 0.5nM, about 30nM to about 1nM, 20nM to about 0.05nM, about 20nM to about 0.075nM, about 20nM to about 0.1nM, about 20nM to about 0.5nM, about 20nM to about 1nM, about 10nM to about 0.05n M, about 10nM to about 0.075nM, about 10nM to about 0.1nM, about 10nM to about 0.5nM, about 10nM to about 1nM, about 5nM to about 0.05nM, about 5nM to about 0.075nM, about 5nM to about 0.1nM, about 5nM Approximately 0.5nM, approximately 5nM to approximately 1nM, approximately 3nM to approximately 0.05nM, approximately 3nM to approximately 0.075nM, approximately 3nM to approximately 0.1nM, approximately 3nM to approximately 0.5nM, approximately 3nM to approximately 1nM, approximately 3nM to approximately 2nM, approximately 2nM to approximately 0.05n M, about 2nM to about 0.075nM, about 2nM to about 0.1nM, about 2nM to about 0.5nM, about 2nM to about 1nM, about 1nM to about 0.05nM, about 1nM to about 0.075nM, about 1nM to about 0.1nM, about 1nM to about 0.5 nM, about 0.5nM to about 0.05nM, about 0.5nM to about 0.075nM, about 0.5nM to about 0.1nM, about 0.1nM to about 0.05nM, about 0.1nM to about 0.075nM or about 0.075nM to about 0.05nM K D and binds to human FcγR (FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b).

[0195] In some aspects, it is contemplated that a heavy chain variable region sequence, e.g., the VH sequence of any one of SEQ ID NOs:7, 12-17, and 26-1...

Claims

1. (i) an immunoglobulin heavy chain (hVH-1-G1m3) comprising the amino acid sequence of SEQ ID NO: 242 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (ii) an immunoglobulin heavy chain (hVH-1-G1m3-GA) comprising the amino acid sequence of SEQ ID NO:243 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO:249; (iii) an immunoglobulin heavy chain (hVH-1-G1m3-GAALIE) comprising the amino acid sequence of SEQ ID NO: 244 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (iv) an immunoglobulin heavy chain (hVH-1-G1m3-YTE) comprising the amino acid sequence of SEQ ID NO: 245 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (v) an immunoglobulin heavy chain (hVH-1-G1m3-LS) comprising the amino acid sequence of SEQ ID NO: 246 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (vi) an immunoglobulin heavy chain (hVH-1-G1m3-DF215) comprising the amino acid sequence of SEQ ID NO: 247 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; or (vii) an immunoglobulin heavy chain (hVH-1-G1m3-DF228) comprising the amino acid sequence of SEQ ID NO: 248 and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; An antibody that binds to human HSP70, comprising:

2. K of 50 nM or less, 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM or less, or 0.05 nM or less as measured by surface plasmon resonance or biolayer interferometry D The antibody of claim 1, which binds to human HSP70 at

3. 3. The antibody of claim 1 or 2, which is capable of forming a higher order antibody:HSP70 complex.

4. 4. The antibody of any one of claims 1 to 3, wherein the antibody:HSP70 complex has a molecular weight of at least about 290 kDa, at least about 300 kDa, at least about 580 kDa, at least about 1,450 kDa, or at least about 1,750 kDa.

5. The antibody of claim 4, wherein the antibody:HSP70 complex has a molecular weight of at least about 300 kDa.

6. The antibody of any one of claims 1 to 4, wherein the antibody:HSP70 complex has a molecular weight of about 290 kDa, about 580 kDa, about 1,450 kDa or about 1,750 kDa.

7. The antibody of any one of claims 3 to 6, wherein the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:

2.

8. The antibody of any one of claims 3 to 7, wherein the antibody:HSP70 complex comprises: (i) about 1 antibody molecule and about 2 HSP70 molecules; (ii) about 2 antibody molecules and about 4 HSP70 molecules; (iii) about 5 antibody molecules and about 10 HSP70 molecules; or (iv) about 6 antibody molecules and about 12 HSP70 molecules.

9. The antibody:HSP70 complex has a K of 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM, or 0.05 nM or less, as measured by surface plasmon resonance or biolayer interferometry. D The antibody of any one of claims 1 to 8, which binds to human FcγR at

10. The antibody of claim 9, wherein the FcγR is FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b.

11. The antibody of any one of claims 1 to 10, which enhances the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells.

12. The antibody of claim 11, wherein the uptake is mediated by human FcγR1, human FcγR2a, human FcγR2b, human FcγR2c, human FcγR3a and / or human FcγR3b.

13. The antibody of any one of claims 3 to 12, wherein the antibody:HSP70 complex activates immune effector cells.

14. The antibody of claim 13, wherein the immune effector cells include CD8+ T cells, CD4+ T cells, NK cells and dendritic cells.

15. Higher order antibody: An antibody capable of forming an HSP70 complex.

16. 16. The antibody of claim 15, wherein the antibody:HSP70 complex has a molecular weight of at least about 290 kDa, at least about 300 kDa, at least about 580 kDa, at least about 1,450 kDa, or at least about 1,750 kDa.

17. 17. The antibody of claim 16, wherein the antibody:HSP70 complex has a molecular weight of at least about 300 kDa.

18. 17. The antibody of claim 15 or 16, wherein the antibody:HSP70 complex has a molecular weight of about 290 kDa, about 580 kDa, about 1,450 kDa or about 1,750 kDa.

19. 19. The antibody of any one of claims 15 to 18, wherein the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:

2.

20. The antibody of any one of claims 15 to 19, wherein the antibody:HSP70 complex comprises: (i) about 1 antibody molecule and about 2 HSP70 molecules; (ii) about 2 antibody molecules and about 4 HSP70 molecules; (iii) about 5 antibody molecules and about 10 HSP70 molecules; or (iv) about 6 antibody molecules and about 12 HSP70 molecules.

21. The antibody:HSP70 complex has a K of 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM, or 0.05 nM or less, as measured by surface plasmon resonance or biolayer interferometry. D The antibody of any one of claims 15 to 20, which binds to human FcγR at

22. The antibody of claim 21, wherein the FcγR is FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b.

23. The antibody of any one of claims 15 to 22, which binds to an epitope of HSP70 corresponding to K573 to Q601 of SEQ ID NO:

11.

24. 24. The antibody of any one of claims 15 to 23, which binds to one, two or three of the following residues of SEQ ID NO:11: H594, K595 and Q601.

25. 25. The antibody of any one of claims 24, which further binds to one, two, three, four or five of the following residues of SEQ ID NO:11: K573, E576, W580, R596 and E598.

26. 26. The antibody of any one of claims 15 to 25, which binds to an epitope of HSP70 comprising KEWHKREQ (SEQ ID NO:250).

27. (i) an immunoglobulin heavy chain variable region comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO:1, a VHCDR2 comprising the amino acid sequence of SEQ ID NO:2, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO:3, and an immunoglobulin light chain variable region comprising a VLCDR1 comprising the amino acid sequence of SEQ ID NO:4, a VLCDR2 comprising the amino acid sequence of SEQ ID NO:5, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO:6; or (ii) an immunoglobulin heavy chain variable region (hVH-1) comprising the amino acid sequence of SEQ ID NO: 12, and an immunoglobulin light chain variable region (hVL-1) comprising the amino acid sequence of SEQ ID NO: 19; The antibody of any one of claims 15 to 26, comprising:

28. 28. The antibody of any one of claims 15 to 27, which enhances the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells.

29. The antibody of claim 28, wherein the uptake is mediated by human FcγR1, human FcγR2a, human FcγR2b, human FcγR2c, human FcγR3a and / or human FcγR3b.

30. (i) an antibody; and (ii) a higher order antibody:HSP70 complex, including HSP70.

31. 31. The antibody:HSP70 complex of claim 30, having a molecular weight of at least about 290 kDa, at least about 300 kDa, at least about 580 kDa, at least about 1,450 kDa or at least about 1,750 kDa.

32. 32. The antibody:HSP70 complex of claim 31, having a molecular weight of at least about 300 kDa.

33. 32. The antibody:HSP70 complex of claim 30 or 31, having a molecular weight of about 290 kDa, about 580 kDa, about 1,450 kDa or about 1,750 kDa.

34. 34. The antibody:HSP70 complex of any one of claims 30 to 33, wherein the ratio of antibody to HSP70 in the antibody:HSP70 complex is about 1:

2.

35. The antibody:HSP70 complex of any one of claims 30-34, comprising: (i) about 1 antibody molecule and about 2 HSP70 molecules; (ii) about 2 antibody molecules and about 4 HSP70 molecules; (iii) about 5 antibody molecules and about 10 HSP70 molecules; or (iv) about 6 antibody molecules and about 12 HSP70 molecules.

36. K of 10 nM or less, 5 nM or less, 1 nM or less, 0.75 nM or less, 0.5 nM or less, 0.1 nM, 0.075 nM or less, or 0.05 nM or less as measured by surface plasmon resonance or biolayer interferometry D The antibody:HSP70 complex of any one of claims 30 to 35, which binds to human FcγR at

37. The antibody:HSP70 complex of claim 36, wherein the FcγR is FcγR1, FcγR2a, FcγR2b, FcγR2c, FcγR3a and / or FcγR3b.

38. 38. The antibody:HSP70 complex of any one of claims 30 to 37, wherein the antibody binds to an epitope of HSP70 corresponding to K573 to Q601 of SEQ ID NO:

11.

39. The antibody:HSP70 complex of any one of claims 30 to 38, wherein the antibody binds to one, two or three of the following residues of SEQ ID NO:11: H594, K595 and Q601.

40. The antibody:HSP70 complex of claim 39, which further binds to one, two, three, four or five of the following residues of SEQ ID NO:11: K573, E576, W580, R596 and E598.

41. 41. The antibody:HSP70 complex of any one of claims 30 to 40, wherein the antibody binds to an epitope of HSP70 comprising KEWHKREQ (SEQ ID NO:250).

42. The antibody (i) an immunoglobulin heavy chain variable region comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO:1, a VHCDR2 comprising the amino acid sequence of SEQ ID NO:2, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO:3, and an immunoglobulin light chain variable region comprising a VLCDR1 comprising the amino acid sequence of SEQ ID NO:4, a VLCDR2 comprising the amino acid sequence of SEQ ID NO:5, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO:6; (ii) an immunoglobulin heavy chain variable region (hVH-1) comprising the amino acid sequence of SEQ ID NO:12, and an immunoglobulin light chain variable region (hVL-1) comprising the amino acid sequence of SEQ ID NO:19; (iii) an immunoglobulin heavy chain (hVH-1-G1m3) comprising the amino acid sequence of SEQ ID NO: 242, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (iv) an immunoglobulin heavy chain (hVH-1-G1m3-GA) comprising the amino acid sequence of SEQ ID NO: 243, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (v) an immunoglobulin heavy chain (hVH-1-G1m3-GAALIE) comprising the amino acid sequence of SEQ ID NO: 244, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (vi) an immunoglobulin heavy chain (hVH-1-G1m3-YTE) comprising the amino acid sequence of SEQ ID NO: 245, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (vii) an immunoglobulin heavy chain (hVH-1-G1m3-LS) comprising the amino acid sequence of SEQ ID NO: 246, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; (viii) an immunoglobulin heavy chain (hVH-1-G1m3-DF215) comprising the amino acid sequence of SEQ ID NO: 247, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; or (ix) an immunoglobulin heavy chain (hVH-1-G1m3-DF228) comprising the amino acid sequence of SEQ ID NO: 248, and an immunoglobulin light chain (hVL-1-Km3) comprising the amino acid sequence of SEQ ID NO: 249; 42. The antibody:HSP70 complex of any one of claims 30 to 41, comprising:

43. 15. An isolated nucleic acid comprising a nucleotide sequence encoding an immunoglobulin heavy chain of an antibody according to any one of claims 1 to 14, and / or a nucleotide sequence encoding an immunoglobulin light chain of an antibody according to any one of claims 1 to 14.

44. 15. An expression vector comprising: (i) a nucleic acid comprising a nucleotide sequence encoding an immunoglobulin heavy chain of the antibody of any one of claims 1 to 14; and / or (ii) a nucleic acid comprising a nucleotide sequence encoding an immunoglobulin light chain of the antibody of any one of claims 1 to 14.

45. 45. A host cell comprising the expression vector of claim 44.

46. A pharmaceutical composition comprising the antibody of any one of claims 1 to 29.

47. 47. The pharmaceutical composition of claim 46 for use in a method of treating cancer in a subject in need thereof, said method comprising administering to said subject an effective amount of the antibody of any one of claims 1 to 42 or the pharmaceutical composition of claim 46.

48. 48. The pharmaceutical composition of claim 47, wherein the cancer is multiple myeloma, breast cancer, melanoma, colon cancer, pancreatic cancer, or prostate cancer.

49. 49. The pharmaceutical composition of claim 48, wherein the cancer is multiple myeloma, breast cancer, melanoma, pancreatic cancer, or colon cancer.

50. 50. The pharmaceutical composition of any one of claims 47 to 49, wherein the subject has a serum HSP70 level greater than 20 ng / mL.

51. 47. The pharmaceutical composition of claim 46, used in combination with chemotherapy or radiation therapy.

52. 52. The pharmaceutical composition of claim 51, wherein the radiation therapy is selected from gamma irradiation, X-ray irradiation, and radioisotope therapy.

53. 47. The antibody of any one of claims 1 to 29 or the pharmaceutical composition of claim 46 for use in a method for enhancing the uptake of tumor-derived ADP-HSP70-peptide antigen complexes by immune effector cells, said method comprising contacting said cells with an effective amount of said antibody.

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