Bispecific antibodies and constructs for targeted degradation of lysosomes and methods of use thereof

Abstract

Provided herein are antigen binding proteins (ABPs) that selectively bind to M6PR, and isoforms and homologues thereof, and compositions comprising the ABPs. Also provided herein are bispecific antigen binding proteins (ABPs) that selectively bind to internalizing receptors, as well as soluble or cell surface target molecules, and isoforms and homologues thereof, and compositions comprising the ABPs.

Description

[0001] 1. Cross-reference to related applications

[0002] This application claims the benefit of U.S. Provisional Application No. 63 / 397,751, filed on August 12, 2022, which is hereby incorporated by reference in its entirety.

[0003] 2. Sequence Listing

[0004] This application contains a sequence listing that has been filed with the Patent Center and is hereby incorporated by reference in its entirety. The XML copy was created at XXXX, is named XXXX, and is XXX bytes in size. 3. Background technology

[0005] Internalized receptors are receptors that move from the plasma membrane to the interior of the cell when cell surface receptors become monoubiquitinated following ligand-induced activation. These receptors are subsequently taken up into endocytic vesicles, from where they are targeted to lysosomes or vacuoles for degradation or are recycled back to the plasma membrane.

[0006] An example of an internalizing receptor includes the mannose-6-phosphate receptor (M6PR), which is a transmembrane glycoprotein that targets enzymes to lysosomes. These receptors play an important role in transporting proteins from the Golgi complex and the cell surface to lysosomes. Cation-independent (CI) M6PRs are multifunctional because these receptors bind to two different types of ligands at the cell surface: the M6P carrier protein and insulin-like growth factor 2 (IGF2). Therefore, CI-M6PRs are synonymously referred to as "M6PR" and "IGF2-R".

[0007] IGF2 plays a key role in metabolic regulation through three types of receptors, two tyrosine kinase receptors (IGF1 receptor and insulin receptor isoform A) and CI-M6PR. Stimulation of type 1 receptor tyrosine kinase induces a protein phosphorylation cascade leading to biological effects, particularly insulin-mediated growth and an increase in CI-M6PR expression, which mediates endocytosis and clearance of IGF2. Lysosomal degradation of IGF2 is critical because elevated IGF2 levels induce overgrowth. When CI-M6PR binds to IGF2, it blocks IGF2-stimulated DNA synthesis in hepatocytes and fibroblasts, and at physiological concentrations, this receptor can block tumor growth mediated by IGF2.

[0008] Therefore, there is a need to develop internalizing receptor antigen binding proteins (ABPs) that can be used to treat, diagnose, and study a variety of diseases, including cancer, lysosomal disorders, and autoimmune diseases.

[0009] Additionally, targeting soluble target molecules or cell surface target molecules for internalization may be used to treat additional diseases.

[0010] For example, it is known that IgE (a soluble molecule) interacts with two major receptors (FcεRI and CD23 / FcεRII4), which are involved in different immunological processes. The binding of allergen-specific IgE to FcεRI expressed on immunological effector cells (including basophils and mast cells) occurs with high affinity. This interaction occurs via two asymmetric binding sites on the receptor and is stabilized by inducing conformational changes in IgE. Exposure to allergens induces cross-linking of FcεRI bound by IgE, resulting in immediate activation of allergic effector cells, which ultimately leads to cell degranulation and the release of vasoactive and proinflammatory mediators. Omalizumab is an anti-IgE antibody that binds to free IgE and prevents IgE from binding to FcεRI on mast cells and basophils. Omalizumab has been approved for severe persistent allergic asthma and chronic spontaneous urticaria. The therapeutic efficacy of anti-IgE has also been reported for allergic rhinitis, allergic bronchopulmonary aspergillosis, latex allergic reactions, atopic dermatitis, allergic urticaria, anaphylaxis, etc.

[0011] In another example, a cell surface target molecule, the epidermal growth factor receptor (EGFR), regulates normal growth and differentiation, but dysregulation of this receptor or one of the EGFR ligands is implicated in the pathogenesis of many cancers. EGFR and insulin-like growth factor receptor (IFGR) play key roles in the tumorigenesis of several types of human cancers.

[0012] Inhibition of receptor tyrosine kinases (such as EGFR) has become an effective treatment strategy for certain human malignancies (for review, see Roussidis AE, In Vivo. 2002 16 (6): 459-69). Although targeted monotherapy may initially be effective in treating cancer, therapeutic resistance often follows, which may be due to the upregulation of other signaling cascades (see, for example, Nahta R et al., Breast Cancer Res. 2006 8 (6): 215 and Horn L et al., Clin Lung Cancer. 2007 8: S68-73). Matuzumab is a humanized immunoglobulin G (1) (IgG (1)) anti-EGFR monoclonal antibody that blocks the activation of EGFR. Therefore, there is a need to develop improved therapeutic agents related to targeting of cell surface molecules (such as EGFR). 4. Summary of the invention

[0013] Provided herein are novel antibody binding proteins (ABPs) and methods of using such ABPs, the novel ABPs comprising a first antigen binding portion having binding specificity to the internalization domain of the cation-independent mannose 6-phosphate receptor (CI-M6PR). In some embodiments, the internalization domain is a fragment of human CI-M6PR.

[0014] Also provided herein are novel ABPs and methods of using such ABPs, the novel ABPs comprising a first binding moiety having binding specificity to human CI-M6PR and a target binding moiety having specificity to a soluble extracellular target molecule or a cell surface target molecule. In some embodiments, the target binding moiety is a second antigen binding moiety. In some embodiments, the target binding moiety is optionally conjugated to the first antigen binding moiety via a linker.

[0015] Also provided herein are ABPs comprising a cargo portion. In some embodiments, the cargo portion is a polypeptide fused to a first antigen binding portion or a second antigen binding portion. In some embodiments, the cargo portion is conjugated to the first antigen binding portion or the second antigen binding portion, optionally via a linker.

[0016] In some embodiments, the first antigen binding moiety specifically binds to a domain of human CI-M6PR selected from the group consisting of: domain 1, domain 4, domain 5, domain 6, domain 7, and domain 8.

[0017] In some embodiments, the first antigen binding portion binds to human CI-M6PR with high affinity. In some embodiments, the first antigen binding portion binds to human CI-M6PR with a dissociation equilibrium constant (K) of about 10 nM or less. D ) binds to human CI-M6PR. In some embodiments, the first antigen binding portion has a K between about 1 nM and about 500 nM D In some embodiments, the first antigen binding moiety binds CI-M6PR with a K between about 10 nM and about 100 nM. D In some embodiments, the first antigen binding portion binds CI-M6PR with a K between about 100 nM and about 200 nM. D In some embodiments, the first antigen binding portion binds to CI-M6PR with a K between about 200 nM and about 300 nM. D In some embodiments, the first antigen binding portion binds to CI-M6PR with a K between about 300 nM and about 400 nM. D In some embodiments, the first antigen binding portion binds CI-M6PR with a K between about 400 nM and about 500 nM. D Combined with CI-M6PR.

[0018] In some embodiments, the dissociation rate (K) of the first antigen binding moiety against CI-M6PR off ) in 1x 10 -8 s -1 With 0.1s -1 In some embodiments, the first antigen binding moiety is directed against the K of CI-M6PR. off In 1x 10 -6 s -1 With 1x10 -2 s -1 In some embodiments, the first antigen binding moiety is directed against the K of CI-M6PR. off About 1x 10 -5 s -1 Or slower.

[0019] In some embodiments, the binding of the first antigen binding moiety to human CI-M6PR is pH dependent. In some embodiments, the first antigen binding moiety is released from CI-M6PR at a pH of 7.4 or less. In some embodiments, the first antigen binding moiety is released from CI-M6PR at a pH of 6.0 or less. In some embodiments, the first antigen binding moiety is released from CI-M6PR at a pH of 5.5 or less. In some embodiments, the first antigen binding moiety is released from CI-M6PR at a pH of 5.0 or less.

[0020] In some embodiments, the first antigen binding portion further comprises (a) a light chain CDR3 (LCDR3) having a sequence of SEQ ID NO: 76 and a heavy chain CDR3 (HCDR3) having a sequence of SEQ ID NO: 35; or (b) a light chain CDR3 having a sequence of SEQ ID NO: 78 and a heavy chain CDR3 having an amino acid sequence of SEQ ID NO: 38; or (c) a light chain CDR3 having a sequence of SEQ ID NO: 81 and a heavy chain CDR3 having a sequence of SEQ ID NO: 41; or (d) a light chain CDR3 having a sequence of SEQ ID NO: 84 and a heavy chain CDR3 having a sequence of SEQ ID NO: 44; or (e) a light chain CDR3 having a sequence of SEQ ID NO: 85 and a heavy chain CDR3 having a sequence of SEQ ID NO: 46; or (f) a light chain CDR3 having a sequence of SEQ ID NO: 88 and a heavy chain CDR3 having a sequence of SEQ ID NO: 49; or (g) a light chain CDR3 having a sequence of SEQ ID NO: 81 and a heavy chain CDR3 having a sequence of SEQ ID NO: 41; NO:91 and a heavy chain CDR3 having a sequence of SEQ ID NO:52; or (h) a light chain CDR3 having a sequence of SEQ ID NO:94 and a heavy chain CDR3 having a sequence of SEQ ID NO:54; or (i) a light chain CDR3 having a sequence of SEQ ID NO:96 and a heavy chain CDR3 having a sequence of SEQ ID NO:55; or (j) a light chain CDR3 having a sequence of SEQ ID NO:99 and a heavy chain CDR3 having a sequence of SEQ ID NO:58; or (k) a light chain CDR3 having a sequence of SEQ ID NO:101 and a heavy chain CDR3 having a sequence of SEQ ID NO:60; or (l) a light chain CDR3 having a sequence of SEQ ID NO:104 and a heavy chain CDR3 having a sequence of SEQ ID NO:63; or (m) a light chain CDR3 having a sequence of SEQ ID NO:107 and a heavy chain CDR3 having a sequence of SEQ ID NO: NO:66; or (n) a light chain CDR3 having a sequence of SEQ ID NO:110 and a heavy chain CDR3 having a sequence of SEQ ID NO:69; or (o) a light chain CDR3 having a sequence of SEQ ID NO:113 and a heavy chain CDR3 having a sequence of SEQ ID NO:72; or (p) a light chain CDR3 having a sequence of SEQ ID NO:116 and a heavy chain CDR3 having a sequence of SEQ ID NO:73.

[0021] In some embodiments, the first antigen binding portion further comprises (a) a light chain CDR1 (LCDR1) having a sequence of SEQ ID NO: 74 and a heavy chain CDR1 (HCDR1) having a sequence of SEQ ID NO: 33; and a light chain CDR2 (LCDR2) having a sequence of SEQ ID NO: 75 and a heavy chain CDR2 (HCDR2) having a sequence of SEQ ID NO: 34; or (b) a LCDR1 having a sequence of SEQ ID NO: 77 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 36; and a LCDR2 having a sequence of SEQ ID NO: 75 and a HCDR2 having a sequence of SEQ ID NO: 37; or (c) a LCDR1 having a sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 39; and a LCDR2 having a sequence of SEQ ID NO: 80 and a HCDR2 having a sequence of SEQ ID NO: 40; or (d) a LCDR1 having a sequence of SEQ ID NO: 82 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 83. or (g) a LCDR1 having a sequence of SEQ ID NO:89 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:90 and a HCDR2 having a sequence of SEQ ID NO:53; or (h) a LCDR1 having a sequence of SEQ ID NO:79 and a heavy chain HCDR1 having a sequence of SEQ ID NO:45; and a LCDR2 having a sequence of SEQ ID NO:80 and a HCDR2 having a sequence of SEQ ID NO:34; or (f) a LCDR1 having a sequence of SEQ ID NO:86 and a heavy chain HCDR1 having a sequence of SEQ ID NO:47; and a LCDR2 having a sequence of SEQ ID NO:87 and a HCDR2 having a sequence of SEQ ID NO:48; or (g) a LCDR1 having a sequence of SEQ ID NO:89 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:90 and a HCDR2 having a sequence of SEQ ID NO:53; or (h) a LCDR1 having a sequence of SEQ ID NO:79 and a heavy chain HCDR1 having a sequence of SEQ ID NO:45; and a LCDR2 having a sequence of SEQ ID NO:80 and a HCDR2 having a sequence of SEQ ID NO:34 NO:92 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:93 and a HCDR2 having the sequence of SEQ ID NO:53; or (i) a LCDR1 having the sequence of SEQ ID NO:95 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:93 and a HCDR2 having the sequence of SEQ ID NO:51;or (j) a LCDR1 having a sequence of SEQ ID NO:97 and a heavy chain HCDR1 having a sequence of SEQ ID NO:56; and a LCDR2 having a sequence of SEQ ID NO:98 and a HCDR2 having a sequence of SEQ ID NO:57; or (k) a LCDR1 having a sequence of SEQ ID NO:100 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:59; or (l) a LCDR1 having a sequence of SEQ ID NO:102 and a heavy chain HCDR1 having a sequence of SEQ ID NO:61; and a LCDR2 having a sequence of SEQ ID NO:103 and a HCDR2 having a sequence of SEQ ID NO:62; or (m) a LCDR1 having a sequence of SEQ ID NO:105 and a heavy chain HCDR1 having a sequence of SEQ ID NO:64; and a LCDR2 having a sequence of SEQ ID NO:106 and a HCDR2 having a sequence of SEQ ID NO: or (n) a LCDR1 having a sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 67; and a LCDR2 having a sequence of SEQ ID NO: 109 and a HCDR2 having a sequence of SEQ ID NO: 68; or (o) a LCDR1 having a sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 112 and a HCDR2 having a sequence of SEQ ID NO: 71; or (p) a LCDR1 having a sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 115 and a HCDR2 having a sequence of SEQ ID NO: 71. ;

[0022] In some embodiments, the first antigen binding moiety comprises a variable light chain (VL) having an amino acid sequence at least 95% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

[0023] In some embodiments, the first antigen binding moiety comprises a variable heavy chain (VH) having an amino acid sequence at least 95% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

[0024] In some embodiments, the first antigen binding moiety comprises a variable light chain (VL) having an amino acid sequence at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

[0025] In some embodiments, the first antigen binding moiety comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

[0026] In some embodiments, the first antigen binding moiety is an antibody fragment (eg, a single chain variable fragment (scFv) or an antigen binding fragment (Fab)).

[0027] In some embodiments, the ABP disclosed herein is transported to lysosomes. In some embodiments, the ABP is transported back to the cell surface after internalization. In some embodiments, the ABP is degraded in the cell.

[0028] In some embodiments, the ABP comprises a multispecific (e.g., bispecific) antibody (see, e.g., Figure 1 ). In some embodiments, the multispecific antibody comprises a first antigen binding portion and a second antigen binding portion. In certain embodiments, the first antigen binding portion is an antigen binding fragment selected from Fab, F(ab')2, a single-chain antibody (e.g., scFv), a diabody, a triabody, a tetrabody, and a domain antibody. In some embodiments, the antibody is selected from a human antibody, a humanized antibody, or a chimeric antibody. In certain embodiments, the second antigen binding portion is an antigen binding fragment selected from Fab, F(ab')2, a single-chain antibody (e.g., scFv), a diabody, a triabody, a tetrabody, and a domain antibody. In some embodiments, the antibody is selected from a human antibody, a humanized antibody, or a chimeric antibody.

[0029] Also provided herein is a method for degrading a soluble target molecule or a cell surface target molecule, the method comprising (a) contacting the target molecule with a multispecific antigen binding protein (ABP), wherein the ABP comprises a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR) on the surface of the cell; and (b) transporting the ABP, the target molecule and the CI-M6PR to a lysosome in the cell, wherein the target molecule is degraded in the lysosome. In some embodiments, the method further comprises (c) transporting the ABP to the cell surface.

[0030] Also provided herein is a method of internalizing a target molecule, the method comprising (a) contacting the target molecule with an antigen binding protein (ABP) as disclosed herein, and (b) internalizing the ABP into a cell. 5. Description of the drawings

[0031] Figure 1A schematic diagram of a bispecific antigen binding protein (ABP) is provided.

[0032] Figure 2A Non-limiting examples of multispecific antigen-binding proteins having a first antigen-binding portion having a full-length antibody that targets M6PR and a second antigen-binding portion having a binding site to a target molecule (such as IgE ( Figure 2A ) or EGFR( Figure 2B ))'s scFv.

[0033] Figures 3A to 3B Non-limiting examples of multispecific antigen-binding proteins having a first antigen-binding portion having an scFv targeting M6PR and a second antigen-binding portion comprising a scFv that binds to a target molecule (such as IgE ( Figure 3A ) or EGFR( Figure 3B ))'s full-length antibody.

[0034] Figure 4 Shown are cell surface and intracellular staining of human M6PR in WT and M6PR-KO K562 cells.

[0035] Figure 5 Shown are titer validation data for mice using recombinant antigens confirmed via flow-based assays using human cell lines (K562 WT, K562 M6PR-KO, Hela WT, and Hela M6PR-KO) and cross-screened against two murine cell lines (Dipak MC38 and 4T1).

[0036] Figure 6 A schematic diagram of the biosensor preparation and binding events is provided for the association / dissociation rates and K of human M6PR via Octet. D The first 192 clones were evaluated for affinity.

[0037] Figure 7 Baseline measurements of each biosensor loaded with a representative mIgG clone (and poly-mIgG control) from the supernatant are provided. Additional biosensors were regenerated to measure additional analyte interactions (mouse M6PR-His6 and off-target binding to His). Binding curve data for representative clones are shown in the top graph.

[0038] Figure 8 An overview of how to screen M6PR-targeting monoclonal antibodies for uptake activity is provided.

[0039] Fig. 9AFigures 4 to 5 show response curves from binding assays of various anti-M6PR antibodies.

[0040] Fig.10 The affinities (K values) of 15 anti-M6PR antibodies are shown. D ) and dynamics (K on , K off )surface.

[0041] Fig.11 is a schematic diagram showing domains D1 to D9 of human CI-M6PR (left panel) and an antibody that binds to D2; the antibody does not bind to a chimeric CI-M6PR in which D2 of human CI-M6PR has been replaced with the corresponding domain from mouse CI-M6PR. Ligands for CI-M6PR are listed adjacent to the domains to which they bind.

[0042] Fig. 12A An illustration of an assay for epitope binning is shown. The conjugated antibody (conjugated Ab) is an anti-M6PR antibody labeled with a probe.

[0043] Fig. 12B Binding curves are shown, with each line indicating a different bin based on the kinetics of the curve.

[0044] Fig.13 A network analysis of epitope binning is shown. The left panel shows the bin assignments for each of the top 12 anti-M6PR antibodies. The right panel shows the positions of bin 7, bin 17, and bin 3 within the entire network analysis.

[0045] Fig.14 Shown is an expression gel of each of the HIS-tagged D1 to D9 CI-M6PR domains.

[0046] Figures 15A to 15H Binding curves of different anti-M6PR antibodies binding to mouse CI-M6PR D1 to D9 domains (LT011-muD1 - LT011muD9) are shown. Human wild-type CI-M6PR was used as a positive control (huLT011).

[0047] Fig.16 It is a summary from Figures 15A to 15H Schematic diagram of the domain mapping results.

[0048] Fig.17A The experimental workflow for evaluating the pH association and dissociation of anti-M6PR antibodies to CI-M6PR is shown.

[0049] Figures 17B to 17D Clone 17F11 ( Fig. 17B )、18G4( Fig. 17C ) and 21D5( Fig.17D) Binding curves for association at pH 7.4 and dissociation at pH 5.0, pH 6.0 and pH 7.4.

[0050] Fig.18 Figure 2 is a histogram quantifying the pH-dependent dissociation of nine different anti-M6PR antibody clones. The pH-dependent dissociation was determined by comparing the K of each antibody at pH 5.0 (pH 5.0 / 7.4) or pH 6.0 (pH 6.0 / pH7.4). off Divided by the K at pH 7.4 off Dissociation was quantified as the fold change in binding displacement after 300 s at the indicated pH.

[0051] Fig.19A is a schematic diagram illustrating the steps and biology of the pH rodo uptake assay.

[0052] Fig.19B Shown is a histogram of the mean fluorescence intensity of the pH rodo uptake assay for 30 anti-M6PR antibodies. The dashed line represents the uptake of the KLH-IGF2 positive control. * indicates antibodies used at less than 25 nM due to low expression during production.

[0053] Figures 20A to 20F Anti-M6PR-omalizumab bispecific antibody ( Fig. 20A and Fig. 20B ) and anti-M6PR antibody ( Figures 20C to 20F ). The uptake of IgE was measured by the mean fluorescence intensity (MFI) of Alexa Fluor 488 conjugated to IgE. Uptake was measured at various concentrations of the bispecific antibody or antibody as indicated on the x-axis.

[0054] Fig.21 Schematic diagrams of the surface HiBit assay and the total HiBit assay are provided along with example readouts for each assay.

[0055] Fig.22A is a histogram showing the span (ie, surface EGFR depletion) of 16 Matuzumab-anti-M6PR bispecific antibodies.

[0056] Fig. 22B is a histogram showing the span (ie, total EGFR depletion) of 16 Matuzumab-anti-M6PR bispecific antibodies.

[0057] Fig. 22C The percentage of surface EGFR activity of exemplary matuzumab-anti-M6PR bispecific antibodies (3C7, 53B11) is shown.

[0058] Fig.22DThe percentage of total EGFR activity of exemplary matuzumab-anti-M6PR bispecific antibodies (3C7, 53B11) is shown.

[0059] Fig.23 is a schematic diagram of the experimental workflow of the hIgE uptake assay using the omalizumab-anti-M6PR (MPR6-Oma) bispecific antibody.

[0060] Fig.24A Shown is the internalization and recycling of the M6PR6-Oma bispecific antibody by HepG2 cells (as indicated by mean fluorescence intensity (MFI))

[0061] Fig. 24B Shows Fig.24A Quantification of internalization and recycling of the M6PR-Oma bispecific antibody shown in FIG. 0' condition was used as an internal baseline and set to 100%. Quantification was measured as hIgE surface labeling / uptake.

[0062] Fig.25A Shown is the internalization and recycling of the MPR6-Oma bispecific antibody by K562 cells (as indicated by mean fluorescence intensity (MFI)).

[0063] Fig.25B Shows Fig.25A Quantification of internalization and recycling of the M6PR-Oma bispecific antibody shown in FIG. 0' condition was used as an internal baseline and set to 100%. Quantification was measured as hIgE surface labeling / uptake.

[0064] Figures 26A to 26B Shown using pH rodo ( Fig.26A ) or AF647( Fig.26B ) Fluorescently labeled C5 uptake by C5 of anti-M6PR / anti-C5 bispecific antibody.

[0065] Figures 27A to 27B LUM-anti-M6PR bispecific antibody mediated HER3 internalization and impaired cell viability in trastuzumab-resistant cells. Fig.27A Surface depletion (i.e., span) and total depletion (i.e., span) of HER3 for a subset of LUM-anti-M6PR bispecific antibodies are shown. Lumretuzumab (LUM), humanized anti-human epidermal growth factor receptor 3 (HER3) monoclonal antibody, and NRG1 (Neuregulin 1) antibody were used as positive controls. Non-binding antibody (KLH) fused to LUM was used as a negative control for uptake activity. Fig.27B Cell viability after treatment of a subset of LUM-anti-M6PR bispecific antibodies is shown. 6. Specific implementation methods

[0066] 6.1. Definitions

[0067] Unless otherwise defined herein, the scientific and technical terms used in conjunction with the present invention should have the meanings commonly understood by those of ordinary skill in the art. In addition, unless the context otherwise requires, singular terms should include plural meanings, and plural terms should include singular meanings. Typically, the terms and techniques used in conjunction with cell and tissue culture, molecular biology, immunology, microbiology, genetics, protein and nucleic acid chemistry, and hybridization described herein are those terms and techniques well known and commonly used in the art. Unless otherwise indicated, the methods and techniques of the present invention are typically performed according to conventional methods well known in the art and as described in each general and more specific reference cited and discussed throughout this specification. See, e.g., Sambrook et al. Molecular Cloning: A Laboratory Manual, 2nd edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1989) and Ausubel et al., Current Protocols in Molecular Biology, Greene Publishing Associates (1992), and Harlow and Lane Antibodies: A Laboratory Manual Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1990), which are incorporated herein by reference. Enzymatic reactions and purification techniques are performed according to the manufacturer's instructions, as is commonly accomplished in the art or as described herein. Terms and laboratory procedures and techniques used in conjunction with analytical chemistry, synthetic organic chemistry, and medicinal and pharmaceutical chemistry described herein are those well known and commonly used in the art. Standard techniques can be used for chemical synthesis, chemical analysis, drug preparation, formulation and delivery, and patient treatment.

[0068] Unless otherwise indicated, the following terms shall be understood to have the following meanings:

[0069] The terms "M6PR", "M6PR protein" and "M6PR antigen" are used interchangeably herein and refer to human M6PR, human CI-M6PR, or any variants (e.g., splice variants and allelic variants) of M6PR expressed naturally by cells or by cells transfected with the M6PR gene, isoforms and species homologs. In some aspects, the M6PR protein is an M6PR protein naturally expressed by primates (e.g., monkeys or humans), rodents (e.g., mice or rats), dogs, camels, cats, cattle, goats, horses or sheep. In some aspects, the M6PR protein is human cation-independent M6PR (NCBI Accession No. NP_000867; SEQ ID NO: 276).

[0070] The term "internalizing receptor" refers to a receptor that can move from an area outside the cell (e.g., including the cell surface) to the interior of the cell. Internalizing receptors include, but are not limited to, endocytic receptors and receptors transported to lysosomes. Non-limiting examples of internalizing receptors include mannose-6-phosphate receptor (CD-M6PR), cation-independent mannose-6-phosphate receptor (CI-M6PR).

[0071] The term "immunoglobulin" refers to a class of structurally related proteins that typically contain two pairs of polypeptide chains: a pair of light (L) chains and a pair of heavy (H) chains. In a "complete immunoglobulin", all four chains are interconnected by disulfide bonds. The structure of immunoglobulins has been well characterized. See, e.g., Paul, Fundamental Immunology, 7th Edition, Chapter 5 (2013) Lippincott Williams & Wilkins, Philadelphia, PA. In short, each heavy chain typically contains a heavy chain variable region (V H ) and the heavy chain constant region (C H The heavy chain constant region usually contains three domains, abbreviated as C H1 , C H2 and C H3 Each light chain generally comprises a light chain variable region (V L ) and the light chain constant region. The light chain constant region usually contains a domain, abbreviated as C L .

[0072] The term "antigen binding protein" (ABP) refers to a protein comprising one or more antigen binding domains that specifically bind to an antigen or epitope. In some embodiments, the antigen binding domain binds to an antigen or epitope with a specificity and affinity similar to that of a naturally occurring antibody. In some embodiments, the ABP comprises an antibody. In some embodiments, the ABP consists of an antibody. In some embodiments, the ABP consists essentially of an antibody. In some embodiments, the ABP comprises an alternative scaffold. In some embodiments, the ABP consists of an alternative scaffold. In some embodiments, the ABP consists essentially of an alternative scaffold. In some embodiments, the ABP comprises an antibody fragment. In some embodiments, the ABP consists of an antibody fragment. In some embodiments, the ABP consists essentially of an antibody fragment. "Internalizing receptor ABP", "anti-internalizing receptor ABP" or "internalizing receptor specific ABP" is an ABP that specifically binds to an antigen of an internalizing receptor as provided herein. In some embodiments, the ABP binds to the extracellular domain of an internalizing receptor. In certain embodiments, the internalizing receptor ABP provided herein binds to an epitope of an internalizing receptor that is conserved between or among internalizing receptor proteins from different species. "Soluble target molecule ABP", "anti-soluble target molecule ABP" or "soluble target molecule specific ABP" is an ABP that specifically binds to an antigen of a soluble target molecule as provided herein. In some embodiments, the soluble target molecule extracellular domain that ABP binds to. In certain embodiments, the soluble target molecule ABP provided herein binds to an epitope of a soluble target molecule that is conserved between or among soluble target molecule proteins from different species. "Cell surface target molecule ABP", "anti-cell surface target molecule ABP" or "cell surface target molecule specific ABP" is an ABP that specifically binds to an antigen of a cell surface target molecule as provided herein. In some embodiments, ABP binds to the extracellular domain of a cell surface target molecule. In certain embodiments, the cell surface target molecule ABP provided herein binds to an epitope of a cell surface target molecule that is conserved between or among cell surface target molecule proteins from different species.

[0073] The term "antibody" is used herein in its broadest sense and includes certain types of immunoglobulin molecules that contain one or more antigen binding domains that specifically bind to an antigen or epitope. Antibodies specifically include complete antibodies (e.g., complete immunoglobulins), antibody fragments, and multispecific antibodies. An example of an antigen binding domain is a V-binding domain. H -V L The antigen binding domain formed by dimerization. Antibodies are a type of ABP.

[0074] The term "bispecific ABP" or "multispecific" refers to an ABP, e.g., an antibody, having at least two different antigen-binding moieties (e.g., antigen-binding domains), each of which specifically binds to a different antigen, e.g., an internalizing receptor and a soluble target molecule or a cell surface target molecule. A bispecific ABP can have any number of valencies, e.g., divalent, trivalent, tetravalent, etc.

[0075] The term "multispecific ABP" refers to an ABP, e.g., an antibody, having at least two (e.g., at least three, at least four, at least five, at least six, etc.) different antigen binding moieties (e.g., antigen binding domains), each of which specifically binds to a different antigen, e.g., an internalizing receptor and a soluble target molecule or a cell surface target molecule. A bispecific ABP can have any number of valencies, e.g., divalent, trivalent, tetravalent, etc. The term "antigen binding moiety" or "antigen binding domain" means the portion of an ABP that is capable of specifically binding to an antigen or epitope.

[0076] The terms "full length antibody," "intact antibody," and "whole antibody" are used interchangeably herein to refer to an antibody that has a structure substantially similar to a naturally occurring antibody structure, or to an antibody that has heavy chains that include an Fc region.

[0077] The term "Fc region" means the C-terminal region of an immunoglobulin heavy chain, which in naturally occurring antibodies interacts with Fc receptors and certain proteins of the complement system. The structures of the Fc regions of various immunoglobulins and the glycosylation sites contained therein are known in the art. See Schroeder and Cavacini, J. Allergy Clin. Immunol., 2010, 125: S41-52, which is incorporated by reference in its entirety. The Fc region may be a naturally occurring Fc region, or a modified Fc region as described elsewhere in the present disclosure.

[0078] V H and V L The V regions can be further subdivided into regions of hypervariability ("hypervariable regions (HVRs)"; also called "complementarity determining regions" (CDRs)), interspersed with more conserved regions. The more conserved regions are called framework regions (FRs). Each V H and V LTypically, it contains three CDRs and four FRs, which are arranged in the following order (from N-terminus to C-terminus): FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. CDRs are involved in antigen binding and affect the antigen specificity and binding affinity of antibodies. See Kabat et al., Sequences of Proteins of Immunological Interest, 5th Edition (1991) Public Health Service, National Institutes of Health, Bethesda, MD, which is incorporated by reference in its entirety.

[0079] The light chain of any vertebrate species can be assigned to one of two types, called kappa (κ) and lambda (λ), based on the sequence of its constant domain.

[0080] The heavy chains of any vertebrate species can be assigned to one of five different classes (or isotypes): IgA, IgD, IgE, IgG, and IgM. These classes are also designated α, δ, ε, γ, and μ, respectively. The IgG and IgA classes are further divided into subclasses based on differences in sequence and function. Humans express the following subclasses: IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2.

[0081] The amino acid sequence boundaries of a CDR can be determined by one of skill in the art using any of a variety of known numbering schemes, including those described by Kabat et al., supra ("Kabat" numbering scheme); Al-Lazikani et al., 1997, J. Mol. Biol., 273:927-948 ("Chothia" numbering scheme); MacCallum et al., 1996, J. Mol. Biol. 262:732-745 ("Contact" numbering scheme); Lefranc et al., Dev. Comp. Immunol., 2003, 27:55-77 ("IMGT" numbering scheme); and Honegge and Plückthun, J. Mol. Biol., 2001, 309:657-70 ("AHo" numbering scheme); each of which is incorporated by reference in its entirety.

[0082] Table 1 provides CDR1-L (V L CDR1), CDR2-L (V L CDR2), CDR3-L (V L CDR3), CDR1-H (V HCDR1), CDR2-H (V H CDR2) and CDR3-H (V H For CDR1-H, residue numbers are provided using both the Kabat and Chothia numbering schemes.

[0083] For example, CDRs can be assigned using antibody numbering software such as Abnum, available at www.bioinf.org.uk / abs / abnum / and described in Abhinandan and Martin, Immunology, 2008, 45:3832-3839, which is incorporated by reference in its entirety.

[0084]

[0085] *When numbering using the Kabat numbering convention, the C-terminus of CDR1-H varies between 32 and 34, depending on the length of the CDR.

[0086] When referring to residues in the antibody heavy chain constant region, the "EU numbering scheme" (eg, as reported in Kabat et al., supra) is generally used.

[0087] "Antibody fragments" include a portion of an intact antibody, such as the antigen binding region or variable region of an intact antibody. Antibody fragments include, for example, Fv fragments, Fab fragments, F(ab')2 fragments, Fab' fragments, scFv (sFv) fragments, and scFv-Fc fragments.

[0088] An "Fv" fragment comprises a non-covalently linked dimer of one heavy- and one light-chain variable domain.

[0089] The "Fab" fragment contains, in addition to the heavy and light chain variable domains, the constant domain of the light chain and the first constant domain of the heavy chain (C H1 ). Fab fragments can be generated, for example, by recombinant methods or by papain digestion of full-length antibodies.

[0090] "F(ab')2" fragments contain two Fab' fragments joined by a disulfide bond near the hinge region. F(ab')2 fragments can be generated, for example, by recombinant methods or by pepsin digestion of intact antibodies. F(ab') fragments can be dissociated, for example, by treatment with β-mercaptoethanol.

[0091] "Single-chain Fv" or "sFv" or "scFv" antibody fragments comprise V H Domain and VL Domain. V H and V L Typically linked via a peptide linker. See Plückthun A. (1994). In some embodiments, the linker is (GGGGS) n In some embodiments, n=1, 2, 3, 4, 5 or 6. See antibodies from Escherichia coli. The antibodies are described in Rosenberg M. & Moore GP (eds.), The Pharmacology of Monoclonal Antibodies, Vol. 113 (pp. 269-315). Springer-Verlag, New York, which is incorporated by reference in its entirety.

[0092] "scFv-Fc" fragments comprise an scFv attached to an Fc domain. For example, the Fc domain can be attached to the C-terminus of the scFv. The Fc domain can be located at the V H or V L This then depends on the orientation of the variable domains in the scFv (i.e., V H -V L or V L -V H ). Any suitable Fc domain known in the art or described herein can be used. In some cases, the Fc domain comprises an IgG4 Fc domain.

[0093] The term "single domain antibody" refers to a molecule in which one variable domain of an antibody specifically binds to an antigen without the presence of another variable domain. Single domain antibodies and fragments thereof are described in Arabi Ghahroudi et al., FEBS Letters, 1998, 414: 521-526 and Muyldermans et al., Trends in Biochem. Sci., 2001, 26: 230-245, each of which is incorporated by reference in its entirety.

[0094] A "monospecific ABP" is an ABP that contains a binding site that specifically binds to a single epitope. An example of a monospecific ABP is the naturally occurring IgG molecule, which, although bivalent, recognizes the same epitope at each antigen binding domain. The binding specificity may be present in any suitable valency.

[0095] The term "monoclonal antibody" refers to an antibody from a population of substantially homogeneous antibodies. A population of substantially homogeneous antibodies comprises antibodies that are substantially similar and that bind to the same epitope, except for variants that may generally occur during the production of monoclonal antibodies. Such variants generally exist only in small amounts. Monoclonal antibodies are generally obtained by including a process for selecting a single antibody from a variety of antibodies. For example, a selection process may be to select a unique clone from a plurality of clones (such as a hybridoma clone library, a phage clone, a yeast clone, a bacterial clone or other recombinant DNA clones). The selected antibody may be further altered, for example, to improve affinity to a target ("affinity maturation"), to humanize the antibody, to improve its production in cell culture and / or to reduce its immunogenicity in a subject.

[0096] The term "chimeric" antibody refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while the remainder of the heavy and / or light chain is derived from a different source or species.

[0097] The "humanized" form of non-human antibodies is a chimeric antibody containing the minimum sequence derived from non-human antibodies. Humanized antibodies are usually human antibodies (receptor antibodies), wherein the residues from one or more CDRs are replaced by the residues from one or more CDRs of non-human antibodies (donor antibodies). Donor antibodies can be any suitable non-human antibodies, such as mice, rats, rabbits, chickens or non-human primate antibodies with desired specificity, affinity or biological effects. In some cases, the framework region residues of the selection of the receptor antibody are replaced by the corresponding framework region residues from the donor antibody. Humanized antibodies can also be included in residues not found in the receptor antibody or the donor antibody. Such modifications can be carried out to further improve the antibody function. For further details, see Jones et al., Nature, 1986, 321:522-525; Riechmann et al., Nature, 1988, 332:323-329; and Presta, Curr. Op. Struct. Biol., 1992, 2:593-596, each of which is incorporated herein by reference in its entirety.

[0098] "Human antibody" refers to an antibody having an amino acid sequence corresponding to the amino acid sequence of an antibody produced by a human or human cell or derived from a non-human source utilizing a human antibody library or human antibody encoding sequence (e.g., an antibody obtained or redesigned from a human source). Human antibodies specifically exclude humanized antibodies.

[0099] An "isolated ABP" or "isolated nucleic acid" is an ABP or nucleic acid that has been separated and / or recovered from a component of its natural environment. Components of the natural environment may include enzymes, hormones, and other proteinaceous or non-proteinaceous materials. In some embodiments, the isolated ABP is purified to a degree sufficient to obtain at least 15 residues of the N-terminal or internal amino acid sequence, for example, by using a spinning cup sequencer. In some embodiments, the isolated ABP is purified to homogeneity by gel electrophoresis (e.g., SDS-PAGE) under reducing or non-reducing conditions and detected by Coomassie blue or silver staining. The isolated ABP includes the ABP in situ within a recombinant cell, because at least one component of the natural environment of the ABP is absent. In some embodiments, the isolated ABP or isolated nucleic acid is prepared by at least one purification step. In some embodiments, the isolated ABP or isolated nucleic acid is purified to at least 80%, 85%, 90%, 95%, or 99% by weight. In some embodiments, the isolated ABP or isolated nucleic acid is purified to at least 80%, 85%, 90%, 95%, or 99% by volume. In some embodiments, the isolated ABP or isolated nucleic acid is provided as a solution comprising at least 85%, 90%, 95%, 98%, 99% to 100% ABP or nucleic acid by weight. In some embodiments, the isolated ABP or isolated nucleic acid is provided as a solution comprising at least 85%, 90%, 95%, 98%, 99% to 100% ABP or nucleic acid by volume.

[0100] "Affinity" refers to the overall strength of non-covalent interactions between a single binding site of a molecule (e.g., ABP) and its binding partner (e.g., antigen or epitope). Unless otherwise indicated, as used herein, "affinity" refers to intrinsic binding affinity, which reflects a 1:1 interaction between members of a binding pair (e.g., ABP and antigen or epitope). The affinity of a molecule X for its partner Y can be expressed by the dissociation equilibrium constant (K D ). The kinetic components that contribute to the dissociation equilibrium constant are described in more detail below. Affinity can be measured by common methods known in the art, including those described herein. For example, affinity can be measured using surface plasmon resonance (SPR) technology (e.g., ) or biolayer interferometry (e.g. ) to determine.

[0101] With respect to the binding of an ABP to a target molecule, the terms "bind," "specifically binds," "binds specifically to," "is specific for," "selectively binds," and "is selective for" a particular antigen (e.g., a polypeptide target) or an epitope on a particular antigen mean binding that is distinct from non-specific or non-selective interactions (e.g., with non-target molecules). For example, specific binding can be measured by measuring binding to a target molecule and comparing it to binding to a non-target molecule. Specific binding can also be determined by competition with a control molecule that mimics the epitope recognized on the target molecule. In this case, if the binding of the ABP to the target molecule is competitively inhibited by the control molecule, specific binding is indicated. For example, in some embodiments, the affinity of an internalizing receptor ABP for a non-target molecule is less than about 50% of the affinity for the internalizing receptor. In some embodiments, the affinity of an internalizing receptor ABP for a non-target molecule is less than about 40% of the affinity for the internalizing receptor. In some embodiments, the affinity of an internalizing receptor ABP for a non-target molecule is less than about 30% of the affinity for the internalizing receptor. In some embodiments, the internalizing receptor ABP has an affinity for a non-target molecule that is less than about 20% of the affinity for the internalizing receptor. In some embodiments, the internalizing receptor ABP has an affinity for a non-target molecule that is less than about 10% of the affinity for the internalizing receptor. In some embodiments, the internalizing receptor ABP has an affinity for a non-target molecule that is less than about 10% of the affinity for the internalizing receptor. In some embodiments, the internalizing receptor ABP has an affinity for a non-target molecule that is less than about 1% of the affinity for the internalizing receptor. In some embodiments, the internalizing receptor ABP has an affinity for a non-target molecule that is less than about 0.1% of the affinity for the internalizing receptor.

[0102] As used herein, the term “k d "(Second -1 ) refers to the dissociation rate constant for a specific ABP-antigen interaction. This value is also called K off value.

[0103] As used herein, the term “k a ”(M -1 ×Seconds -1 ) refers to the association rate constant for a specific ABP-antigen interaction. This value is also called K on value.

[0104] As used herein, the term "K D “(M)” refers to the dissociation equilibrium constant for a specific ABP-antigen interaction. K D =k d / k a .

[0105] As used herein, the term "K A ”(M -1 ) refers to the association equilibrium constant for a specific ABP-antigen interaction. A =k a / k d .

[0106] An "affinity matured" ABP is an ABP that has one or more changes (e.g., in one or more CDRs or FRs) that result in an increased affinity of the ABP for its antigen compared to a parent ABP that does not have the changes. In one embodiment, the affinity matured ABP has nanomolar or picomolar affinity for the target antigen. A variety of methods known in the art can be used to produce affinity matured ABPs. For example, Marks et al. (Bio / Technology, 1992, 10:779-783, which is incorporated by reference in its entirety) describe affinity matured ABPs by V H and V L Affinity maturation of domain shuffling. For example, random mutagenesis of CDR and / or framework residues is described by Barbas et al. (Proc. Nat. Acad. Sci. USA, 1994, 91: 3809-3813); Schier et al., Gene, 1995, 169: 147-155; Yelton et al., J. Immunol., 1995, 155: 1994-2004; Jackson et al., J. Immunol., 1995, 154: 3310-33199; and Hawkins et al., J. Mol. Biol., 1992, 226: 889-896; each of these documents is incorporated by reference in its entirety.

[0107] An "immunoconjugate" is an ABP conjugated to one or more heterologous molecules.

[0108] "Effector functions" refer to those biological activities mediated by the Fc region of an antibody, which may vary depending on the antibody isotype. Examples of antibody effector functions include C1q binding to activate complement-dependent cytotoxicity (CDC), Fc receptor binding to activate antibody-dependent cellular cytotoxicity (ADCC), and antibody-dependent cellular phagocytosis (ADCP).

[0109] When used herein in the context of two or more ABPs, the term "competes with" or "cross-competes with" indicates that two or more ABPs compete for binding to an antigen (e.g., an internalizing receptor). In an exemplary assay, an internalizing receptor is coated on a surface and contacted with a first internalizing receptor ABP, after which a second internalizing receptor ABP is added. In another exemplary assay, a first internalizing receptor ABP is coated on a surface and contacted with an internalizing receptor, and then a second internalizing receptor ABP is added. If the presence of a first internalizing receptor ABP reduces the binding of a second internalizing receptor ABP in either assay, the ABP competes. The term "competes with" also includes a combination of ABPs in which one ABP reduces the binding of another ABP, but in which no competition is observed when the ABPs are added in the opposite order. However, in some embodiments, the first ABP and the second ABP inhibit the binding of each other, regardless of the order in which they are added. In some embodiments, one ABP reduces the binding of another ABP to its antigen by at least 25%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, or at least 95%. The technician can select the concentration of the antibody for competitive determination based on the affinity of the ABP to the internalized receptor and the valency of the ABP. The determination described in this definition is illustrative, and the technician can use any suitable determination to determine whether the antibodies compete with each other. Suitable determinations are described in, for example, the following documents: Cox et al., "Immunoassay Methods," in Assay Guidance Manual [Internet], updated on December 24, 2014 (www.ncbi.nlm.nih.gov / books / NBK92434 / ; Access date: September 29, 2015); Silman et al., Cytometry, 2001, 44: 30-37; and Finco et al., J. Pharm. Biomed. Anal., 2011, 54: 351-358; Each of these documents is incorporated by reference in its entirety.

[0110] The term "epitope" means a part of an antigen that specifically binds to an ABP. An epitope is usually composed of surface accessible amino acid residues and / or sugar side chains, and may have specific three-dimensional structural features and specific charge characteristics. The difference between a conformational epitope and a non-conformational epitope is that, in the presence of a denaturing solvent, the binding to the former rather than the latter may be lost. An epitope may include amino acid residues that are directly involved in binding and other amino acid residues that are not directly involved in binding. The epitope that is bound to an ABP can be determined using known techniques for epitope determination, such as, for example, testing the binding of an ABP to an internalizing receptor, a soluble target molecule, and / or a cell surface target molecule variant with different point mutations, or testing the binding to a chimeric internalizing receptor, a soluble target molecule, and / or a cell surface target molecule variant.

[0111] The percentage "identity" between a polypeptide sequence and a reference sequence is defined as the percentage of amino acid residues in the polypeptide sequence that are identical to the amino acid residues in the reference sequence, after aligning the sequences and introducing gaps (if necessary) to achieve the maximum percentage of sequence identity. Alignment for determining percentage of amino acid sequence identity can be achieved in various ways known to those skilled in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, MEGALIGN (DNASTAR), CLUSTALW, CLUSTAL OMEGA or MUSCLE software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithm required for achieving maximum alignment over the full length of the compared sequences.

[0112] "Conservative substitution" or "conservative amino acid substitution" refers to the replacement of an amino acid with a chemically or functionally similar amino acid. Conservative substitution tables providing similar amino acids are well known in the art. For example, in some embodiments, the amino acid groups provided in Tables 2 to 4 are considered conservative substitutions of each other.

[0113]

[0114]

[0115]

[0116]

[0117] Additional conservative substitutions can be found, for example, in Creighton, Proteins: Structures and Molecular Properties, 2nd ed., (1993) WH Freeman & Co., New York, NY. ABPs generated by making one or more conservative substitutions of amino acid residues in a parent ABP are termed "conservatively modified variants."

[0118] The term "treating" (and variants thereof, such as "treat" or "treatment") refers to a clinical intervention intended to alter the natural course of a disease or condition in a subject in need thereof. Treatment may be used for prevention as well as during clinical pathology. Desirable therapeutic effects of treatment include preventing the occurrence or recurrence of the disease, alleviating symptoms, reducing any direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, ameliorating or relieving the disease state, and alleviating or improving prognosis.

[0119] As used herein, the term "therapeutically effective amount" or "effective amount" refers to an amount of an ABP or pharmaceutical composition provided herein that is effective in treating a disease or condition when administered to a subject.

[0120] As used herein, the term "subject" means a mammalian subject. Exemplary subjects include humans, monkeys, dogs, cats, mice, rats, cows, horses, camels, goats, rabbits, and sheep. In certain embodiments, the subject is a human. In some embodiments, the subject suffers from a disease or condition that can be treated with the ABP provided herein. In some aspects, the disease or condition is cancer. In some aspects, the disease or condition is a viral infection.

[0121] The term "package insert" is used to refer to instructions customarily included in commercial packages of therapeutic or diagnostic products (e.g., kits), that contain information about the indications, usage, dosage, administration, combination therapy, contraindications and / or warnings concerning the use of such therapeutic or diagnostic products.

[0122] As used herein, the term "cytotoxic agent" refers to a substance that inhibits or prevents the function of cells and / or causes cell death or destruction.

[0123] "Chemotherapeutic agents" refer to chemical compounds that can be used to treat cancer. Chemotherapeutic agents include "anti-hormonal agents" or "endocrine therapeutic agents," which are used to modulate, reduce, block or inhibit the effects of hormones that can promote cancer growth.

[0124] The term "tumor" refers to all neoplastic cell growth and proliferation, whether malignant or benign, and all precancerous and cancerous cells and tissues. As referred to herein, the terms "cancer," "cancerous," "cell proliferative disorders," "proliferative disorders," and "tumor" are not mutually exclusive. The terms "cell proliferative disorders" and "proliferative disorders" refer to disorders associated with some degree of abnormal cell proliferation. In some embodiments, the cell proliferative disease is cancer.

[0125] The term "pharmaceutical composition" refers to a preparation which is in a form that permits the biological activity of the active ingredients contained therein to be effective in treating a subject, and which contains no additional components which are unacceptably toxic to the subject.

[0126] The terms "modulate" and "modulation" refer to the decrease or inhibition or alternatively the activation or increase of the recited variable.

[0127] The terms "increase" and "activate" refer to increasing the recited variable by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 50-fold, 100-fold or more.

[0128] The terms "reduce" and "inhibit" refer to a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 50-fold, 100-fold or greater decrease in the recited variable.

[0129] The term "agonism" refers to activation of receptor signaling to induce a biological response associated with activated receptors. An "agonist" is an entity that binds to a receptor and agonizes the receptor.

[0130] The term "antagonize" refers to the inhibition of receptor signaling to suppress the biological response associated with activated receptors. An "antagonist" is an entity that binds to a receptor and antagonizes the receptor.

[0131] A "variant" of a polypeptide (e.g., an antibody) comprises an amino acid sequence in which one or more amino acid residues are inserted into, deleted from, and / or substituted into the amino acid sequence relative to the native polypeptide sequence, and retains substantially the same biological activity as the native polypeptide. Standard techniques in the art can be used to measure the biological activity of the polypeptide (e.g., if the variant is an antibody, its activity can be tested by binding assays, as described herein). Variants of the present invention include fragments, analogs, recombinant polypeptides, synthetic polypeptides, and / or fusion proteins.

[0132] A "derivative" of a polypeptide is a polypeptide (e.g., an antibody) that has been chemically modified, for example, via conjugation to another chemical moiety such as polyethylene glycol, albumin (e.g., human serum albumin), phosphorylation, and glycosylation. Unless otherwise indicated, the term "antibody" includes derivatives, variants, fragments, and muteins thereof other than antibodies comprising two full-length heavy chains and two full-length light chains, examples of which are described below.

[0133] A nucleotide sequence is "operably linked" to a regulatory sequence if the regulatory sequence affects the expression of the nucleotide sequence (e.g., the level, timing, or position of expression). A "regulatory sequence" is a nucleic acid that affects the expression of a nucleic acid to which it is operably linked (e.g., the level, timing, or position of expression). For example, a regulatory sequence can exert its effect, for example, directly on the regulated nucleic acid, or through one or more other molecules (e.g., a polypeptide that binds to the regulatory sequence and / or nucleic acid). Examples of regulatory sequences include promoters, enhancers, and other expression control elements (e.g., polyadenylation signals). Further examples of regulatory sequences are described, for example, in the following literature: Goeddel, 1990, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, CA, and Baron et al., 1995, Nucleic Acids Res. 23: 3605–06.

[0134] A "host cell" is a cell that can be used to express a nucleic acid (e.g., a nucleic acid of the invention). The host cell can be a prokaryotic organism, such as E. coli, or it can be a eukaryotic organism, such as a unicellular eukaryotic organism (e.g., a yeast or other fungi), a plant cell (e.g., a tobacco or tomato plant cell), an animal cell (e.g., a human cell, a monkey cell, a hamster cell, a rat cell, a mouse cell, or an insect cell), or a hybridoma. Examples of host cells include CS-9 cells, the COS-7 line of monkey kidney cells (ATCC CRL 1651) (see Gluzman et al., 1981, Cell 23:175), L cells, C127 cells, 3T3 cells (ATCC CCL 163), Chinese hamster ovary (CHO) cells or their derivatives such as Veggie CHO and related cell lines grown in serum-free medium (see Rasmussen et al., 1998, Cytotechnology 28:31), HeLa cells, BHK (ATCC CRL 10) cell line, CV1 / EBNA cell line derived from the African green monkey kidney cell line CV1 (ATCC CCL 70) (see McMahan et al., 1991, EMBO J. 10:2821), human embryonic kidney cells (such as 293, 293EBNA or MSR 293), human epidermal A431 cells, human Colo205 cells, other transformed primate cell lines, normal diploid cells, cell lines derived from primary tissue cultured in vitro, primary explants, HL-60, U937, HaK or Jurkat cells. Generally, the host cell is a cultured cell that can be transformed or transfected with a polypeptide encoding nucleic acid, which can then be expressed in the host cell.

[0135] The phrase "recombinant host cell" can be used to refer to a host cell that has been transformed or transfected with a nucleic acid to be expressed. A host cell can also be a cell that contains a nucleic acid but does not express it at a desired level unless a regulatory sequence is introduced into the host cell so that it is operably linked to the nucleic acid. It should be understood that the term host cell refers not only to a specific subject cell, but also to the progeny or potential progeny of this cell. Because certain modifications may occur in subsequent generations due to, for example, mutations or environmental influences, such progeny may not actually be identical to the parental cell, but are still included within the scope of the term as used herein.

[0136] 6.2. Other interpretation rules

[0137] The ranges recited herein are to be understood as shorthand for all values ​​within the range, including the recited endpoints. For example, a range of 1 to 50 is to be understood to include any number, combination of numbers, or subrange from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, and 50.

[0138] Unless otherwise stated, reference to a compound having one or more stereocenters is intended to refer to each and every stereoisomer thereof and to all combinations of stereoisomers thereof.

[0139] 6.3. Nucleic Acids

[0140] In one aspect, the invention provides isolated nucleic acid molecules. Nucleic acids include, for example, polynucleotides encoding all or part of an antigen binding protein (e.g., one or two chains of an antibody, or a fragment, derivative, mutant protein, or variant thereof of the invention); polynucleotides sufficient for use as hybridization probes; PCR primers or sequencing primers for identifying, analyzing, mutating, or amplifying polynucleotides encoding polypeptides; antisense nucleic acids for inhibiting expression of polynucleotides; and the aforementioned complementary sequences. Nucleic acids can be of any length. They can have, for example, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75, 100, 125, 150, 175, 200, 250, 300, 350, 400, 450, 500, 750, 1,000, 1,500, 3,000, 5,000 or more nucleotides in length, and / or can include one or more additional sequences, such as regulatory sequences, and / or be part of a larger nucleic acid (e.g., a vector). Nucleic acids can be single-stranded or double-stranded and can comprise RNA and / or DNA nucleotides, as well as artificial variants thereof (eg, peptide nucleic acids).

[0141] Nucleic acids encoding antibody polypeptides (e.g., heavy chain or light chain, variable domains only or full length) can be isolated from cells of mice immunized with internalized receptors, soluble target molecules and / or cell surface target molecules. Nucleic acids can be isolated by conventional procedures such as polymerase chain reaction (PCR).

[0142] Nucleic acid sequences encoding heavy chain variable regions and light chain variable regions are shown herein. It will be appreciated by a skilled artisan that due to the degeneracy of the genetic code, each of the polypeptide sequences disclosed herein is encoded by a large number of other nucleic acid sequences. The present invention provides each degenerate nucleotide sequence encoding each antigen-binding protein of the present invention.

[0143] The present invention further provides nucleic acids that hybridize with other nucleic acids (e.g., nucleic acids containing nucleotide sequences of any one of internalized receptors, soluble target molecules and / or cell surface target molecule genes) under specific hybridization conditions. Methods for making nucleic acid hybridization are well known in the art. See, for example, Curr. Prot. Mol. Biol., John Wiley & Sons, NY (1989), 6.3.1-6.3.6. As defined herein, moderately stringent hybridization conditions use a hybridization buffer containing 5 times sodium chloride / sodium citrate (SSC), 0.5% SDS, 1.0 mM EDTA (pH 8.0), about 50% formamide, 6 times SSC and a pre-wash solution of a hybridization temperature of 55°C (or other similar hybridization solutions, such as a hybridization solution containing about 50% formamide, whose hybridization temperature is 42°C), and washing conditions are 60°C, in 0.5 times SSC, 0.1% SDS. Stringent hybridization conditions are hybridization in 6x SSC at 45° C., followed by one or more washes in 0.1x SSC, 0.2% SDS at 68° C. In addition, those skilled in the art can manipulate hybridization and / or wash conditions to increase or decrease the stringency of hybridization so that nucleic acids containing nucleotide sequences that are at least 65, 70, 75, 80, 85, 90, 95, 98, or 99% identical to each other typically remain hybridized to each other. Basic parameters that influence the choice of hybridization conditions and guidance for designing appropriate conditions are described, for example, by Sambrook, Fritsch, and Maniatis (1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, Chapters 9 and 11; and 1995 in Curr. Prot. Mol. Biol., Ausubel et al., eds., John Wiley & Sons, Inc., Sections 2.10 and 6.3-6.4) and can be readily determined by one of ordinary skill in the art based on, for example, the length and / or base composition of the DNA.

[0144] Can be introduced into nucleic acid by sudden change, thereby cause the amino acid sequence of its encoded polypeptide (for example, antigen-binding proteins) to change.Can use any technology known in the art to introduce sudden change.In one embodiment, use for example site-directed mutagenesis scheme to change one or more specific amino acid residues.In another embodiment, use for example random mutagenesis scheme to change one or more randomly selected residues.No matter how it is made, mutant polypeptide can be expressed and for expected attribute (for example, combined with internalization receptor, soluble target molecule and / or cell surface target molecule) be screened.

[0145] 6.4. Antibodies

[0146] Internalizing receptor antibodies can be purified from host cells that have been transfected with a gene encoding the antibody by eluting the filtered supernatant of the host cell culture fluid using a heparin HP column using a salt gradient.

[0147] Fab fragments have V L 、V H , C L and C H1 The F(ab')2 fragment is a bivalent fragment with two Fab fragments connected by a disulfide bridge at the hinge region; the Fd fragment has a V H and C H1 Domain; Fv fragment has the V L and V H domain; and the dAb fragment has a V H Domain, V L domain, or V H or V L Antigen-binding fragments of the cytokine domain (U.S. Pat. Nos. 6,846,634, 6,696,245, U.S. Application Publication Nos. 05 / 0202512, 04 / 0202995, 04 / 0038291, 04 / 0009507, 03 / 0039958, Ward et al., Nature 341:544-546, 1989).

[0148] The polynucleotide and polypeptide sequences of specific light chain variable domains and heavy chain variable domains are described below. Antibodies comprising light chains and heavy chains are named by combining the name of the light chain and the name of the heavy chain variable domain. In some embodiments, light chain variable sequences are provided in SEQ ID NOs: 32-63, and heavy chain variable sequences are provided in SEQ ID NOs: 1-31.

[0149] In other embodiments, the antibody may include a specific heavy chain or light chain, while the complementary light chain variable domain or heavy chain variable domain remains non-specific. In particular, certain embodiments herein include antibodies that bind to specific antigens (such as M6PR) by specific light chains or heavy chains, so that the complementary heavy chains or light chains can be promiscuous, or even unrelated, but can be determined by, for example, screening combinatorial libraries. Portolano et al., Immunol. Vol. 150 (3), pp. 880-887, (1993); Clackson et al., Nature, Vol. 352, pp. 624 to 628 (1991); Adler et al., Anatively paired antibody library yields drug leads with higher sensitivity and specificity than a randomly paired antibody library, MAbs (2018)); Adler et al., Rare, high-affinity mouse anti-PD-1 antibodies that function in checkpoint blockade, discovered using microfluidics and molecular genomics, MAbs (2017).

[0150] Naturally occurring immunoglobulin chains exhibit the same general structure of relatively conserved framework regions (FRs) joined by three hypervariable regions (also referred to as complementary determining regions or CDRs). From N-terminus to C-terminus, both light and heavy chains contain domains FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The amino acid distribution of each domain is consistent with the definition of Kabat et al. in Sequences of Proteins of Immunological Intere, 5th edition, USDept. of Health and Human Services, PHS, NIH, NIH Publication no. 91-3242, 1991.

[0151] The term "human antibody" is also referred to as a "fully human antibody" and includes all antibodies having one or more variable regions and constant regions derived from human immunoglobulin sequences. In one embodiment, all variable domains and constant domains are derived from human immunoglobulin sequences (fully human antibodies). These antibodies can be prepared in a variety of ways, examples of which are described below, including by immunizing with an antigen of interest in a mouse that is genetically modified to express antibodies derived from human heavy chain encoding genes and / or light chain encoding genes.

[0152] Humanized antibodies have sequences with one or more amino acid substitutions, deletions and / or additions of differences with sequences of antibodies derived from non-human species, so that when the humanized antibodies are applied to human subjects, compared with non-human species antibodies, the humanized antibodies are less likely to induce immune responses, and / or induce less severe immune responses. In one embodiment, certain amino acids in the framework and constant domains of the heavy chain and / or light chain of non-human species antibodies are mutated to produce humanized antibodies. In another embodiment, the constant domains from human antibodies are fused with the variable domains of non-human species. In another embodiment, one or more amino acid residues in one or more CDR sequences of non-human antibodies are changed to reduce the possible immunogenicity of non-human antibodies when applied to human subjects, wherein the amino acid residues changed are not critical for the immunospecific binding of antibodies to their antigens, or the changes made to the amino acid sequence are conservative changes, so that the combination of humanized antibodies and antigens will not be significantly worse than the combination of non-human antibodies and antigens. Examples of how to make humanized antibodies can be found in US Pat. Nos. 6,054,297, 5,886,152, and 5,877,293.

[0153] The term "chimeric antibody" refers to an antibody containing one or more regions from one antibody and one or more regions from one or more other antibodies. In one embodiment, one or more of the CDRs are derived from a human anti-internalizing receptor antibody. In another embodiment, all CDRs are derived from a human anti-internalizing receptor antibody. In another embodiment, CDRs from more than one human anti-internalizing receptor antibody are mixed and matched into a chimeric antibody. For example, a chimeric antibody may include CDR1 from a light chain of a first human anti-internalizing receptor antibody, CDR2 and CDR3 from a light chain of a second human anti-internalizing receptor antibody, and CDRs from a heavy chain of a third human anti-internalizing receptor antibody. Further, the framework region may be derived from one of the same anti-internalizing receptor antibodies, derived from one or more different antibodies (such as human antibodies), or derived from a humanized antibody. In one example of a chimeric antibody, a portion of the heavy and / or light chain is identical, homologous, or derived from an antibody from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain is identical, homologous, or derived from an antibody from another species or belonging to another antibody class or subclass. Fragments of such antibodies that exhibit the desired biological activity (i.e., the ability to specifically bind to an internalizing receptor) are also included.

[0154] Those of ordinary skill in the art can easily prepare fragments or analogs of antibodies according to the teachings of this specification and using techniques well known in the art. Preferred amino-terminal and carboxyl-terminal ends of fragments or analogs appear near the boundaries of functional domains. Structural domains and functional domains can be identified by comparing nucleotide and / or amino acid sequence data with public or proprietary sequence databases. Computerized comparison methods can be used to identify sequence motifs or predicted protein conformation domains present in other proteins of known structure and / or function. Methods for identifying protein sequences that fold into known three-dimensional structures are known. See, for example, Bowie et al., 1991, Science 253:164.

[0155] Antigen-binding fragments derived from antibodies can be obtained, for example, by proteolysis of antibodies, for example, by digesting whole antibodies with pepsin or papain according to conventional methods. For example, antibody fragments can be produced by enzymatic cleavage of antibodies with pepsin to provide a 5S fragment known as F(ab')2. The fragment can be further cleaved using a thiol reducing agent to produce a 3.5S Fab' monovalent fragment. Optionally, the cleavage reaction can be carried out using a capping group of a thiol group produced by the cleavage of a disulfide bond. As an alternative, enzymatic cleavage using papain can directly produce two monovalent Fab fragments and an Fc fragment. These methods are described, for example, by Goldenberg, U.S. Pat. No. 4,331,647, Nisonoff et al., Arch. Biochem. Biophys. 89:230, 1960; Porter, Biochem. J. 73:119, 1959; Edelman et al., in Methods in Enzymology 1:422 (Academic Press 1967); and Andrews, SM and Titus, JA, in Current Protocols in Immunology (Coligan JE et al., eds.), John Wiley and Sons, New York (2003), pp. 2.8.1 to 2.8.10 and 2.10A.1 to 2.10A.5. Other methods for cleaving antibodies (such as separation of heavy chains to form monovalent light and heavy chain fragments (Fd), further cleavage of fragments, or other enzymatic, chemical or genetic techniques) may also be used, as long as the fragments bind to the antigen recognized by the intact antibody.

[0156] Antibody fragments can also be any synthetic or genetically engineered proteins. For example, antibody fragments include isolated fragments consisting of the variable region of the light chain, "Fv" fragments consisting of the variable regions of the heavy and light chains, and recombinant single-chain polypeptide molecules in which the variable regions of the light chain and the heavy chain are connected by a peptide linker (scFv protein).

[0157] Another form of antibody fragment is a peptide comprising one or more complementary determining regions (CDRs) of an antibody. CDRs (also referred to as "minimal recognition units" or "hypervariable regions") can be covalently or non-covalently incorporated into a molecule to make it an antigen binding protein. CDRs can be obtained by constructing a polynucleotide encoding the CDRs of interest. Such polynucleotides are prepared, for example, by synthesizing the variable region using the polymerase chain reaction using mRNA from antibody-producing cells as a template (see, e.g., Larrick et al., Methods: A Companion to Methods in Enzymology 2:106, 1991; Courtenay Luck, "Genetic Manipulation of Monoclonal Antibodies", In Monoclonal Antibodies: Production, Engineering and Clinical Application, Ritter et al. (eds.), p. 166 (Cambridge University Press 1995); and Ward et al., "Genetic Manipulation and Expression of Antibodies", In Monoclonal Antibodies: Principles and Applications, Birch et al., (eds.), p. 137 (Wiley Liss, Inc. 1995).

[0158] Therefore, in one embodiment, the binding agent comprises at least one CDR as described herein. The binding agent may comprise at least two, three, four, five or six CDRs as described herein. The binding agent may further comprise at least one variable region domain of an antibody described herein. The variable region domain may have any size or amino acid composition, and generally comprises at least one CDR sequence responsible for binding to human M6PR, such as HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3, which are specifically described herein and are adjacent to or in frame with one or more framework sequences. In general, the variable (V) region domain may be an immunoglobulin heavy (V H ) chain variable domain and / or light (V L ) chain variable domains. Thus, for example, the V region domain may be a monomer and be V H Domain or V Ldomains that are capable of independently binding to human M6PR with an affinity ranging from 1 nM to 1 pM. Alternatively, the V region domains may be dimers and contain V H V H 、V H V L or V L V L Dimer. A V region dimer contains at least one V H and at least one non-covalently associated V L Chain (hereinafter referred to as F V ). If desired, these chains can be covalently coupled directly, for example via a disulfide bond between the two variable domains or through a linker (eg, a peptide linker), to form a single-chain Fv (scFv).

[0159] The variable region domain can be any naturally occurring variable domain or an engineered version thereof. The so-called engineered version means a variable region domain created using recombinant DNA engineering techniques. Such engineered versions include, for example, those created from a specific antibody variable region by insertion, deletion or change in or to the amino acid sequence of a specific antibody. Specific examples include engineered variable region domains containing at least one CDR from a first antibody and optionally one or more framework amino acids and the remainder of the variable region domain from a second antibody.

[0160] The variable region domain may be covalently attached to at least one other antibody domain or fragment thereof at the C-terminal amino acid. Thus, for example, a V present in a variable region domain H The V domain can be linked to an immunoglobulin CH1 domain or a fragment thereof. L The V domain may be linked to a CK domain or a fragment thereof. In this manner, for example, the antibody may be a Fab fragment in which the antigen binding domain contains an associated V domain. H Domain and V L The CH1 domain may be extended with further amino acids, for example to provide a hinge region or a portion of a hinge region domain as found in a Fab' fragment, or to provide further domains, such as antibody CH2 and CH3 domains.

[0161] As described herein, the antibody comprises at least one of these CDRs. For example, one or more CDRs can be incorporated into known antibody framework regions (IgG1, IgG2, etc.), or conjugated to a suitable carrier to enhance its half-life. Suitable carriers include, but are not limited to, Fc, polyethylene glycol (PEG), albumin, transferrin, etc. These carriers and other suitable carriers are known in the art. Such conjugated CDR peptides can be monomers, dimers, tetramers or other forms. In one embodiment, one or more water-soluble polymers are bonded to one or more specific positions of the binding agent, for example, bonded to the amino terminus.

[0162] 6.5. Antigen Binding Proteins

[0163] In one aspect, the invention provides antigen binding proteins (eg, antibodies, antibody fragments, antibody derivatives, antibody muteins, and antibody variants) that bind to internalized receptors, soluble target molecules, and / or cell surface target molecules.

[0164] In some embodiments, the ABP comprises an antigen binding portion (e.g., at least one antigen binding portion) that specifically binds to an antigen of an internalizing receptor. In certain embodiments, binding of the antigen promotes intracellular transport of the ABP by the internalizing receptor.

[0165] In certain embodiments, the first antigen binding moiety specifically binds to an epitope on the antigen that promotes intracellular transport of the ABP through an internalizing receptor. In certain embodiments, the internalizing receptor is a cell surface internalizing receptor. In certain embodiments, the internalizing receptor is an endocytic receptor. In certain embodiments, the internalizing receptor is capable of targeting lysosomes. In certain embodiments, the internalizing receptor is an oncogenic receptor.

[0166] In certain embodiments, the internalizing receptor is CI-M6PR.

[0167] In some embodiments, the antigen binding portion comprises an antigen binding fragment selected from the group consisting of: Fab, F(ab')2, single chain antibody (scFv), (scFv)2, diabodies, triabodies, tetrabodies, and domain antibodies.

[0168] In some embodiments, the antigen binding portion comprises a Fab fragment and an Fc domain. In some embodiments, a "Fab-Fc" fragment comprises a Fab fragment attached to an Fc domain. For example, the Fc domain can be attached to the C-terminus of the Fab fragment. Any suitable Fc domain known in the art or described herein can be used. In some cases, the Fc domain comprises an IgG4 Fc domain.

[0169] In another aspect of the disclosure, the ABP comprises a multispecific ABP.In some embodiments, the multispecific ABP of the disclosure comprises a first antigen binding portion and a second antigen binding portion.

[0170] In some embodiments, the multispecific ABP comprises a first antigen binding moiety that specifically binds to an antigen of an internalizing receptor. In certain embodiments, binding of the antigen promotes intracellular transport of the multispecific ABP through the internalizing receptor.

[0171] In certain embodiments, the first antigen binding moiety specifically binds to an epitope on an antigen that promotes intracellular transport of the multispecific ABP through an internalizing receptor. In certain embodiments, a specific epitope provides a favorable internalization function. In certain embodiments, the internalizing receptor is a cell surface internalizing receptor. In certain embodiments, the internalizing receptor is an endocytic receptor. In certain embodiments, the internalizing receptor is capable of targeting lysosomes. In certain embodiments, the internalizing receptor is an oncogenic receptor.

[0172] In some embodiments, the multispecific ABP binds to an antigen of an internalizing receptor, wherein the internalizing receptor is the cation-dependent mannose-6-phosphate receptor (CD-M6PR).

[0173] In certain embodiments, the internalizing receptor is CD-M6PR. In certain embodiments, the internalizing receptor is CI-M6PR.

[0174] In certain embodiments, the internalizing receptor is CI-M6PR. In some embodiments, the binding of the antigen binding portion that binds to CI-M6PR is an allosteric binder and / or a direct confirmatory agonist that rapidly promotes the internalization of M6PR. In some embodiments, the binding of the antigen binding portion to CI-M6PR promotes the binding of M6PR to M6P (e.g., competitive binding and / or non-competitive binding).

[0175] In some embodiments, the first antigen binding moiety comprises an antigen binding fragment selected from the group consisting of: Fab, F(ab')2, single chain antibody (scFv), (scFv)2, diabodies, triabodies, tetrabodies, and domain antibodies.

[0176] In some embodiments, the antigen binding portion comprises a Fab fragment and an Fc domain. In some embodiments, a "Fab-Fc" fragment comprises a Fab fragment attached to an Fc domain. For example, the Fc domain can be attached to the C-terminus of the Fab fragment. Any suitable Fc domain known in the art or described herein can be used. In some cases, the Fc domain comprises an IgG4 Fc domain.

[0177] In certain embodiments, the multispecific ABP comprises a second antigen binding moiety that specifically binds to an antigen of a soluble target molecule or a cell surface target molecule. In some embodiments, internalization of a soluble target molecule or a cell surface target molecule provides a therapeutic benefit.

[0178] In certain embodiments, the second antigen binding portion specifically binds to a soluble target portion. In some embodiments, the soluble target molecule is immunoglobulin E (IgE). Non-limiting examples of antigen binding portions that bind to IgE can be found in U.S. Patent Nos.: 8,071,097 and 7,867,494, and PCT Patent Publication No.: WO2017211928A1, all of which are incorporated herein by reference in their entirety.

[0179] In some embodiments, the second antigen binding portion comprises: a second variable light chain region (V L ) CDR1, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.: 138. In some embodiments, the second antigen binding portion comprises: a second variable light chain region (V L ) CDR2, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.: 139. In certain embodiments, the second antigen binding portion comprises: a second variable light chain region (V L )CDR3 having an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.: 140.

[0180] In some embodiments, the second antigen binding portion comprises: a second variable heavy chain region (V H ) CDR1, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.: 135. In some embodiments, the second antigen binding portion comprises: a second variable heavy chain region (V H) CDR2, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.: 136. In some embodiments, the second antigen binding portion comprises: a second variable heavy chain region (V H )CDR3 having an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.:137.

[0181] In some embodiments, the second antigen binding moiety comprises: a variable light chain (V L ), which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO: 134. In some embodiments, the second antigen binding portion comprises: a variable heavy chain region (V H ), which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the amino acid sequence of SEQ ID NO.: 133.

[0182] In certain embodiments, the second antigen binding moiety specifically binds to a cell surface target molecule. In a specific embodiment, the cell surface target molecule is selected from: ERBB receptors (e.g., EGFR, ERBB2, ERBB3, ERBB4), erythropoietin-producing hepatocytes (EPH) receptors, fibroblast growth factor (FGF) receptors (e.g., FGFR1, FGFR2, FGFR3, FGFR4, FGFR5), platelet-derived growth factor (PDGF) receptors (e.g., PDGFR-A, PDGFR-B), vascular endothelial growth factor (VEGF) receptors (e.g., VEGFR1 / FLT1, VEGFR2 / FLK1, VEGF3), tyrosine kinase with immunoglobulin-like and EGF-like domains (TIE) receptors, insulin-like growth factor (IGF) receptors (e.g., INS-R, IGFIR, IR-R), discoidin domain (DD) receptors, c-Met receptor (MET), Nantes proto-oncogene receptor (RON); also known as macrophage stimulating 1 receptor, Flt3 fin-related tyrosine kinase 3 (Flt3), colony stimulating factor 1 (CSF1) receptor, adhesion-associated kinase receptors (e.g., Axl), c-kit receptor (KIT) and insulin receptor-related (IRR) receptor.

[0183] In certain embodiments, the cell surface target molecule is epidermal growth factor receptor (EGFR).Non-limiting examples of antigen binding moieties that bind to EGFR are found in U.S. Pat. Nos. 10,954,286, 10,844,127, and 11,040,111, each of which is hereby incorporated by reference in its entirety.

[0184] In some embodiments, the second antigen binding portion comprises: a second variable light chain region (V L ) CDR1, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 122 and SEQ ID NO: 130. In some embodiments, the second antigen binding portion comprises: a second V L CDR2, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 123 and SEQ ID NO: 131. In some embodiments, the second antigen binding portion comprises: a second V LCDR3, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 124 and SEQ ID NO: 132. In certain embodiments, the second antigen binding portion comprises: a second variable heavy chain region (V H )CDR1 having an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 119 and SEQ ID NO: 127.

[0185] In certain embodiments, the second antigen binding moiety comprises: a second V H CDR2, which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 120 and SEQ ID NO: 128. In certain embodiments, the second antigen binding portion comprises: a second V H A CDR3 having an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 121 and SEQ ID NO: 129.

[0186] In some embodiments, the second antigen binding moiety comprises: a variable light chain (V L ), which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 118 and SEQ ID NO: 126. In some embodiments, the second antigen binding portion comprises: a second variable heavy chain (V H ), which has an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to an amino acid sequence selected from SEQ ID NO.: 117 and SEQ ID NO: 125.

[0187] In some embodiments, the second antigen binding moiety comprises an antigen binding fragment selected from the group consisting of: Fab, F(ab')2, single chain antibody (scFv), (scFv)2, diabodies, triabodies, tetrabodies, and domain antibodies.

[0188] Antigen binding proteins can have the structure of, for example, naturally occurring immunoglobulins. "Immunoglobulins" are tetrameric molecules. In naturally occurring immunoglobulins, each tetramer is composed of two pairs of identical polypeptide chains, each pair having a "light" chain (about 25 kDa) and a "heavy" chain (about 50 to 70 kDa). The amino terminal portion of each chain includes a variable region of about 100 to 110 or more amino acids that are primarily responsible for antigen recognition. The carboxyl terminal portion of each chain defines a constant region that is primarily responsible for effector function. Human light chains are classified as kappa light chains and lambda light chains. Heavy chains are classified as μ, δ, γ, α or ε, and the isotypes of antibodies are defined as IgM, IgD, IgG, IgA and IgE, respectively. In light and heavy chains, the variable and constant regions are joined by a "J" region of about 12 or more amino acids, wherein the heavy chain also includes a "D" region of about 10 more amino acids. See generally, Fundamental Immunology, Chapter 7 (Paul, W., ed., 2nd ed., Raven Press, NY (1989)) (which is incorporated by reference in its entirety for all purposes). The variable regions of each light / heavy chain pair form the antibody combining site, such that an intact immunoglobulin has two combining sites.

[0189] Antigen binding proteins according to the present invention include antigen binding proteins that promote intracellular transport of ABP through internalization receptors. Antigen binding proteins according to the present invention also include multispecific ABPs that include a second antigen binding protein that internalizes a soluble target molecule or a cell surface target molecule to provide a therapeutic benefit.

[0190] Different antigen binding proteins can bind to different domains of internalized receptors, soluble target molecules or cell surface target molecules through different mechanisms of action. As indicated in particular herein, unless otherwise indicated, domain regions are specified as such as including groups. For example, amino acids 4 to 12 refer to nine amino acids: the amino acids at positions 4 and 12, and seven intervening amino acids in the sequence.

[0191] Derivatives of ABP within the scope of the present invention include covalent conjugates or aggregated conjugates of ABP or its fragments with other proteins or polypeptides, such as by expressing a recombinant fusion protein containing a heterologous polypeptide fused to the N-terminus or C-terminus of the ABP polypeptide. For example, the conjugated peptide can be a heterologous signal (or leader) polypeptide, for example, a yeast α-factor leader or a peptide such as an epitope tag. Fusion proteins containing antigen-binding proteins can include peptides (e.g., polyhistidine) added to facilitate purification or identification of antigen-binding proteins. Antigen-binding proteins can also be connected to the FLAG peptide Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Asp-Lys (DYKDDDDK) as described in the following documents: Hopp et al., Bio / Technology 6: 1204, 1988, and U.S. Patent No. 5,011,912. The FLAG peptide is highly antigenic and provides an epitope that is reversibly bound by a specific monoclonal antibody (mAb), thereby enabling rapid determination and easy purification of expressed recombinant proteins. Reagents useful for preparing fusion proteins in which the FLAG peptide is fused to a given polypeptide are commercially available (Sigma, St. Louis, MO).

[0192] One suitable Fc polypeptide described in PCT application WO 93 / 10151 (incorporated herein by reference) is a single-chain polypeptide extending from the N-terminal hinge region of the Fc region of a human IgG1 antibody to the native C-terminus. Another useful Fc polypeptide is the Fc mutant protein described in U.S. Pat. No. 5,457,035 and Baum et al., 1994, EMBO J. 13: 3992-4001. The amino acid sequence of the mutant protein is identical to the native Fc sequence presented in WO 93 / 10151, except that amino acid 19 has been changed from Leu to Ala, amino acid 20 has been changed from Leu to Glu, and amino acid 22 has been changed from Gly to Ala. The mutant protein exhibits reduced affinity for Fc receptors.

[0193] In some embodiments, the Fc region comprises human IgG1, human IgG2, human IgG4 or variants thereof. In some embodiments, the Fc region comprises modifications that affect effector function. In some embodiments, the Fc region comprises LALAPG, N297A, DANA, LALA or N297Q mutations. The LALAPG mutation may refer to L234A, L235A and P329G mutations in the Fc region. The DANA mutation may refer to D265A and N297A mutations in the Fc region. The LALA mutation may refer to L234A and L235A mutations in the Fc region.

[0194] In some embodiments, the first antigen binding portion of the multispecific ABP is in Fab format and the second antigen binding portion of the multispecific ABP is in scFv format, or the first antigen binding portion is in scFv format and the second antigen binding portion is in Fab format (see, e.g., Figure 1 and Figure 2A 3A to 3B and 3A to 3B show non-limiting examples of anti-M6PR / IgE and anti-M6PR / EGFR multispecific antibodies. In some embodiments, the first antigen binding portion of the multispecific ABP is in Fab format and the second antigen binding portion of the multispecific ABP is in Fab format. In some embodiments, the first antigen binding portion of the multispecific ABP is in scFv format and the second antigen binding portion of the multispecific ABP is in scFv format.

[0195] In some embodiments, the multispecific ABP comprises a full-length antibody sequence that binds to an internalizing receptor, comprising a variable light chain amino acid sequence of an internalizing receptor antibody, a variable heavy chain amino acid sequence of an internalizing receptor antibody, an FC region of an internalizing receptor antibody; and an antigen binding portion that binds to a soluble target molecule or a cell surface target molecule, such as an scFv or Fab region of an antibody that binds to a soluble target molecule or a cell surface target molecule, such as Figures 2A to 2B shown.

[0196] In some embodiments, the multispecific ABP comprises a full-length antibody sequence targeting a soluble target molecule or a cell surface target molecule, which comprises a variable light chain amino acid sequence of an antibody against a soluble target molecule or a cell surface target molecule, a variable heavy chain amino acid sequence of an antibody against a soluble target molecule or a cell surface target molecule, an FC region of an antibody against a soluble target molecule or a cell surface target molecule; and an antigen binding portion that binds to an internalizing receptor, such as an scFv or Fab region of an antibody that binds to an internalizing receptor, such as Figures 3A to 3B shown.

[0197] The antigen-binding proteins (e.g., antibodies, antibody fragments, multispecific antibodies, and antibody derivatives) of the present invention may include any constant region known in the art. The light chain constant region may be, for example, a κ- or λ-type light chain constant region, for example, a human κ- or λ-type light chain constant region. The heavy chain constant region may be, for example, an α-, δ-, ε-, γ-, or μ-type heavy chain constant region, for example, a human α-, δ-, ε-, γ-, or μ-type heavy chain constant region. In one embodiment, the light chain constant region or the heavy chain constant region is a fragment, derivative, variant, or mutant protein of a naturally occurring constant region.

[0198] Techniques for deriving antibodies of different subclasses or isotypes from the antibody of interest (i.e., subclass switching) are known. Thus, for example, an IgG antibody can be derived from an IgM antibody, and vice versa. Such techniques allow the preparation of new antibodies that have the antigen binding properties of a given antibody (parent antibody), but also exhibit biological properties associated with an antibody isotype or subclass different from that of the parent antibody. Recombinant DNA techniques can be used. Cloned DNA encoding specific antibody polypeptides can be used for such procedures, for example, DNA encoding the constant domains of an antibody of a desired isotype. See also Lantto et al., 2002, Methods Mol. Biol. 178: 303-16.

[0199] In one aspect, the present disclosure provides an antigen binding portion comprising an antigen binding fragment of an internalizing receptor, a soluble target molecule and / or a cell surface target molecule of the present invention. Such fragments may consist entirely of antibody-derived sequences, or may contain additional sequences. Examples of antigen binding fragments include Fab, F(ab')2, single-chain antibodies (scFv), (scFv)2, diabodies, triabodies, tetrabodies, and domain antibodies. Other examples are provided in the following literature: Lunde et al., 2002, Biochem.Soc.Trans.30:500-06.

[0200] In some embodiments, the present disclosure provides a first antigen binding moiety that is a first antigen binding fragment and a second antigen binding moiety that is a second antigen binding fragment.

[0201] Single-chain antibodies (scFv) can be formed by connecting a heavy chain variable domain fragment and a light chain variable domain (Fv region) fragment via an amino acid bridge (a short peptide linker, e.g., a synthetic sequence of amino acid residues), thereby producing a single polypeptide chain. L and V H ) DNA to prepare such single-chain Fv (scFv). The resulting polypeptides can fold back on themselves to form antigen-binding monomers, or they can form multimers (e.g., dimers, trimers, or tetramers), depending on the length of the flexible linker between the two variable domains (Kortt et al., 1997, Prot. Eng. 10:423; Kortt et al., 2001, Biomol. Eng. 18:95-108, Bird et al., 1988, Science 242:423-26, and Huston et al., 1988, Proc. Natl. Acad. Sci. USA 85:5879-83). By combining different V L and VH Polypeptides can form multimeric scFvs that bind to different epitopes (Kriangkum et al., 2001, Biomol. Eng. 18: 31-40). Technologies developed for producing single-chain antibodies include those described in the following literature: U.S. Pat. No. 4,946,778; Bird, 1988, Science 242: 423; Huston et al., 1988, Proc. Natl. Acad. Sci. USA 85: 5879; Ward et al., 1989, Nature 334: 544, de Graaf et al., 2002, Methods Mol Biol. 178: 379-87. The invention encompasses ScFvs comprising the variable domain combinations 45A7, 45A12, 43C8, 12D5, 53B11, 35E2, 06D12, 03C12, 18G4, 02C7, 14A3, 10H5, 06G11, 06H10, 01E5, 21E12, 06F1, 20E12, 21D5, 12A1, 06D1, 07B2, 07E11, 56A2, 01A7, 07A4, 16H4, 08D8, 06C1, 35B2, and 17F11.

[0202] Antigen binding portion that binds to M6PR

[0203] In one aspect of the disclosure, the ABP comprises an antigen binding protein comprising at least a first antigen binding portion that binds to M6PR. In some embodiments, the disclosure provides an antigen binding portion comprising a light chain variable region selected from SEQ ID NO: 17-32 of Table 6 or a heavy chain variable region selected from SEQ ID NO: 1-16 of Table 5, as well as fragments, derivatives, muteins and variants thereof.

[0204] The position of the CDR (underline) that creates a part of antigen binding site is also shown below, and framework region (FR) is the intervening fragment of these variable domain sequences.There are three CDRs (CDR1 to 3) and four FRs (FR 1 to 4) in both light chain variable region and heavy chain variable region.The CDR district of each light chain and heavy chain is also grouped by antibody type.Antigen-binding proteins of the present invention include, for example, antigen-binding proteins with a combination of light chain variable domains and heavy chain variable domains, and the light chain variable domains and heavy chain variable domains are selected from the group of combinations of antibody clones composed of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Tables 5 to 8.

[0205] In some embodiments, the first antigen binding portion comprises all six CDR sequences identical to one of the clones in the library of internal receptor binding clones of Tables 5 to 8 (three CDRs of light chain "LCDR1 to LCDR3" and three CDRs of heavy chain "HCDR1 to HCDR3"). In some embodiments, the first antigen binding portion comprises three of the six CDR sequences identical to one of the clones in the library of M6PR binding clones of Tables 5 to 8 (three CDRs of light chain or three CDRs of heavy chain). In some embodiments, the first antigen binding portion comprises one, two, three, four or five of the six CDR sequences identical to one of the clones in the library of M6PR binding clones of Tables 5 to 8.

[0206] In one embodiment, the invention provides a first antigen binding portion comprising a light chain variable domain, which light chain variable domain comprises an amino acid sequence that differs from the sequence of a light chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 6 by one or more residues, wherein each such sequence difference is independently a deletion, insertion or substitution of an amino acid residue. In another embodiment, the light chain variable domain comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 97% or 99% identical to the sequence of a light chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 6. In another embodiment, the light chain variable domain comprises an amino acid sequence encoded by a nucleotide sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 97% or 99% identical to a nucleotide sequence encoding a light chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 6. In another embodiment, the light chain variable domain comprises an amino acid sequence encoded by a polynucleotide that hybridizes under moderately stringent conditions with the complement of a polynucleotide encoding a light chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 6. In another embodiment, the light chain variable domain comprises an amino acid sequence encoded by a polynucleotide that hybridizes under moderately stringent conditions with the complement of a polynucleotide encoding a light chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 6. In another embodiment, the light chain variable domain comprises an amino acid sequence encoded by a polynucleotide that hybridizes under moderately stringent conditions to the complement of the light chain polynucleotides of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 6.

[0207] In another embodiment, the invention provides a first antigen binding portion comprising a heavy chain variable domain, which heavy chain variable domain comprises an amino acid sequence that differs from the sequence of a heavy chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 5 by one or more residues, wherein each such sequence difference is independently a deletion, insertion or substitution of an amino acid residue. In another embodiment, the heavy chain variable domain comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to the sequence of a heavy chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 5. In another embodiment, the heavy chain variable domain comprises an amino acid sequence encoded by a nucleotide sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to a nucleotide sequence encoding a heavy chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 5. In another embodiment, the heavy chain variable domain comprises an amino acid sequence encoded by a polynucleotide that hybridizes under moderately stringent conditions with the complement of a polynucleotide encoding a heavy chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Table 5. In another embodiment, the heavy chain variable domain comprises an amino acid sequence encoded by a polynucleotide that hybridizes under moderately stringent conditions to the complement of a polynucleotide encoding a heavy chain variable domain selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 of Table 5. In another embodiment, the heavy chain variable domain comprises an amino acid sequence encoded by a polynucleotide that hybridizes under moderately stringent conditions to the complement of a heavy chain polynucleotide described herein.

[0208] In some embodiments, the heavy chain variable domain comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to an amino acid sequence selected from SEQ ID NO.: 1-16. In some embodiments, the light chain variable domain comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to an amino acid sequence selected from SEQ ID NO: 17-32.

[0209] Specific embodiments of the antigen binding portion of the present disclosure include one or more amino acid sequences identical to the amino acid sequences of one or more of the CDRs and / or FRs mentioned herein. In one embodiment, the antigen binding portion includes the light chain CDR1 (LCDR1) sequence exemplified above. In another embodiment, the antigen binding portion includes the light chain CDR2 (LCDR2) sequence exemplified above. In another embodiment, the antigen binding portion includes the light chain CDR3 (LCDR3) sequence exemplified above. In another embodiment, the antigen binding portion includes the heavy chain CDR1 (HCDR1) sequence exemplified above. In another embodiment, the antigen binding portion includes the heavy chain CDR2 (HCDR2) sequence exemplified above. In another embodiment, the antigen binding portion includes the heavy chain CDR3 (HCDR3) sequence exemplified above.

[0210] In one embodiment, the invention provides an antigen binding portion comprising one or more CDR sequences that differ from the CDR sequences set forth above by no more than 5, 4, 3, 2 or 1 amino acid residues.

[0211] In some embodiments, at least one of the HCDR1 sequences of the antigen binding portion is a HCDR1 sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 from Table 7. In some embodiments, at least one of the HCDR2 sequences of the antigen binding portion is a HCDR2 sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 from Table 7. In some embodiments, at least one of the HCDR3 sequences of the antigen binding portion is a HCDR3 sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 from Table 7.

[0212] In some embodiments, the antigen binding portion comprises: a variable heavy chain region (V H ) CDR1 having an amino acid sequence that is at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 96%, at least 98%, at least 99% or at least 100% identical to an amino acid sequence selected from SEQ ID NO.: 33, 36, 39, 42, 45, 47, 50, 56, 61, 64, 67 and 70. In some embodiments, the antigen binding portion comprises: V H CDR2, which has an amino acid sequence that is at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 96%, at least 98%, at least 99% or at least 100% identical to an amino acid sequence selected from SEQ ID NO.: 34, 37, 40, 43, 48, 51, 53, 57, 59, 62, 65, 68 and 71. In some embodiments, the antigen binding portion comprises: V H A CDR3 having an amino acid sequence that is at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 96%, at least 98%, at least 99% or at least 100% identical to an amino acid sequence selected from SEQ ID NO.: 35, 38, 41, 44, 46, 49, 52, 54, 55, 58, 60, 63, 66, 69, 72 and 73.

[0213] In some embodiments, the antigen binding portion comprises: a variable heavy chain region (V H ) CDR1 having an amino acid sequence that is at least 95% identical to an amino acid sequence selected from SEQ ID NO: 33, 36, 39, 42, 45, 47, 50, 56, 61, 64, 67, and 70. In some embodiments, the antigen binding portion comprises: V H CDR2 has an amino acid sequence that is at least 95% identical to an amino acid sequence selected from SEQ ID NO.: 34, 37, 40, 43, 48, 51, 53, 57, 59, 62, 65, 68, and 71. In some embodiments, the antigen binding portion comprises: V H CDR3 having an amino acid sequence that is at least 95% identical to an amino acid sequence selected from SEQ ID NO.: 35, 38, 41, 44, 46, 49, 52, 54, 55, 58, 60, 63, 66, 69, 72 and 73.

[0214] In another embodiment, the light chain LCDR1 sequence of the antigen binding portion is a light chain LCDR1 sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 from Table 8. In some embodiments, the light chain LCDR2 sequence of the antigen binding portion is a light chain LCDR2 sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 from Table 8. In some embodiments, the light chain LCDR3 sequence of the antigen binding portion is the light chain LCDR3 sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 from Table 8.

[0215] In some embodiments, the antigen binding portion comprises: a variable light chain region (V L ) CDR1 having an amino acid sequence that is at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 96%, at least 98%, at least 99% or at least 100% identical to an amino acid sequence selected from SEQ ID NO.: 74, 77, 79, 82, 86, 89, 92, 95, 97, 100, 102, 105, 108, 111 and 114. In some embodiments, the antigen binding portion comprises: V L CDR2, which has an amino acid sequence that is at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 96%, at least 98%, at least 99% or at least 100% identical to an amino acid sequence selected from SEQ ID NO. 75, 80, 83, 87, 90, 93, 98, 103, 106, 109, 112 and 115. In some embodiments, the antigen binding portion comprises: V L A CDR3 having an amino acid sequence that is at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 96%, at least 98%, at least 99% or at least 100% identical to an amino acid sequence selected from SEQ ID NO.: 76, 78, 81, 84, 85, 88, 91, 94, 96, 99, 101, 104, 107, 110, 113 and 116.

[0216] In some embodiments, the antigen binding portion comprises: a variable light chain region (V L) CDR1 having an amino acid sequence that is at least 95% identical to an amino acid sequence selected from SEQ ID NO.: 74, 77, 79, 82, 86, 89, 92, 95, 97, 100, 102, 105, 108, 111, and 114. In some embodiments, the antigen binding portion comprises: V L CDR2 has an amino acid sequence that is at least 95% identical to an amino acid sequence selected from SEQ ID NO. 75, 80, 83, 87, 90, 93, 98, 103, 106, 109, 112, and 115. In some embodiments, the antigen binding portion comprises: V L A CDR3 having an amino acid sequence that is at least 95% identical to an amino acid sequence selected from SEQ ID NO.: 76, 78, 81, 84, 85, 88, 91, 94, 96, 99, 101, 104, 107, 110, 113 and 116.

[0217] In another embodiment, the antigen binding portion comprises 1, 2, 3, 4 or 5 CDR sequences, each of which independently differs from the CDR sequences of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 by 6, 5, 4, 3, 2, 1 or 0 single amino acid additions, substitutions and / or deletions, and the antigen binding protein further comprises 1, 2, 3, 4 or 5 CDR sequences, each of which independently differs from the CDR sequences by 6, 5, 4, 3, 2, 1 or 0 single amino acid additions, substitutions and / or deletions. In some embodiments, the antigen binding portion comprises 1, 2, 3, 4, or 5 CDR sequences, each of which has at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity to the CDR sequences of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11.

[0218] The nucleotide sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11, or the amino acid sequence of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 can be altered by, for example, random mutagenesis or by site-directed mutagenesis (e.g., oligonucleotide-directed site-specific mutagenesis) to create an altered polynucleotide comprising one or more specific nucleotide substitutions, deletions or insertions compared to the unmutated polynucleotide. Examples of techniques for making such changes are described in the following literature: Walder et al., 1986, Gene 42:133; Bauer et al., 1985, Gene 37:73; Craik, BioTechniques, January 1985, 12-19; ​​Smith et al., 1981, Genetic Engineering: Principles and Methods, Plenum Press; and U.S. Pat. Nos. 4,518,584 and 4,737,462. These methods and other methods can be used to make derivatives of, for example, anti-internalizing receptor antibodies that have desirable properties, for example, increased affinity, avidity or specificity for internalizing receptors, increased in vivo or in vitro activity or stability, or reduced in vivo side effects compared to underivatized antibodies.

[0219] In one embodiment, the antigen binding portion of the invention comprises an IgG1 heavy chain domain of any one of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11, or a fragment of an IgG1 heavy chain domain of any one of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11. In another embodiment, the antigen binding portion of the invention comprises a kappa light chain constant region of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11, or a fragment of the kappa light chain constant region of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11. In another embodiment, the antigen binding portion of the invention comprises an IgG1 heavy chain domain of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11, or a fragment thereof, and a kappa light chain domain of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11, or a fragment thereof.

[0220] In some embodiments, at least one antigen binding portion of the invention includes those comprising, for example, variable domain combinations 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11, which have a desired isotype (e.g., IgA, IgG1, IgG2, IgG3, IgG4, IgM, IgE, and IgD) and Fab or F(ab')2 fragments thereof. In addition, if IgG4 is desired, it may also be desirable to introduce a point mutation (CPSCP->CPPCP) in the hinge region (as described in Bloom et al., 1997, Protein Science 6:407, which is incorporated herein by reference) to mitigate the tendency to form disulfide bonds within the H chain, which may result in heterogeneity in IgG4 antibodies.

[0221] In one embodiment, the antigen binding portion of the antigen binding protein has 1x10 -4 s -1 or lower K off In another embodiment, K off 5x10 -5 s -1 In another embodiment, K off The antigen binding portion is substantially identical to an antibody having a combination of a light chain variable domain sequence and a heavy chain variable domain sequence selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 of Tables 5 to 6. In another embodiment, the antigen binding portion has a Kp1 sequence that is substantially identical to that of the antibody. off Binds to M6PR, the antibody comprising one or more CDRs from an antibody having a combination of a light chain variable domain sequence and a heavy chain variable domain sequence selected from the group consisting of 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 of Tables 7 to 8. In another embodiment, the antigen binding portion has a KD substantially the same as that of the antibody. off In another embodiment, the antigen binding portion binds to M6PR and the antibody comprises one of the amino acid sequences exemplified above. off Binding to M6PR, the antibody comprises one or more CDRs from an antibody comprising one of the amino acid sequences exemplified above.

[0222] The antigen binding proteins of the present disclosure comprising the first antigen binding moiety may have a -7 M, less than or equal to 1x 10 -7 M, less than or equal to 0.5x 10 -7 M, less than or equal to 1x 10 -8 M, less than or equal to 1x 10 -9 M, less than or equal to 1x 10 -10 M, less than or equal to 1x 10 -11 M or less than or equal to 1x 10 -12 In some embodiments, the first antigen binding moiety has a K in the range of 1 nM to 1 pM. D Binds to internalized receptors.

[0223] In some embodiments, the first antigen binding moiety has a K of less than 500 nM. D In some embodiments, the first antigen binding moiety binds to the internalized receptor with a K of less than 200 nM. D In some embodiments, the first antigen binding moiety binds to the internalized receptor with a K of less than 25 nM. D In some embodiments, the first antigen binding moiety binds to the internalized receptor with a K of less than 5 nM. D In some embodiments, the first antigen binding moiety binds to the internalized receptor with a K of less than 2 nM. D In some embodiments, the first antigen binding moiety is present at 1x10 -7 M to 5x10 -12 K in the M range D In some embodiments, the first antigen binding moiety is present at 2x10 -7 M to 9.5x10 -8 K in the M range D Binds to internalized receptors.

[0224] The affinity of an antibody or antigen binding protein for its binding partner can be determined by one of ordinary skill in the art using conventional techniques, such as those described by Scatchard et al. (Ann. NY Acad. Sci. 51: 660-672 (1949)), or by surface plasmon resonance (SPR; BIAcore, Biosensor, Piscataway, NJ). For surface plasmon resonance, the target molecule is immobilized on a solid phase and exposed to a ligand in a mobile phase running along a flow cell. If the ligand binds to the immobilized target, the local refractive index changes, resulting in a change in the SPR angle, which can be monitored in real time by detecting changes in the intensity of the reflected light. The rate of change of the SPR signal can be analyzed to derive the apparent rate constants for the association and dissociation phases of the binding reaction. The ratio of these values ​​gives the apparent equilibrium constant (affinity) (see, e.g., Wolff et al., Cancer Res. 53: 2560-65 (1993)).

[0225] The antibody or antigen binding protein according to the present disclosure can belong to any immunoglobulin class, such as IgG, IgE, IgM, IgD or IgA. It can be obtained or derived from animals (e.g., poultry (e.g., chicken) and mammals), including but not limited to mice, rats, hamsters, rabbits or other rodents, cattle, horses, sheep, goats, camels, humans or other primates.

[0226] 6.6. Monoclonal Antibodies

[0227] In another aspect, the invention provides monoclonal antibodies that bind to an internalizing receptor (eg, M6PR).The monoclonal antibodies of the invention can be generated using a variety of known techniques. In general, monoclonal antibodies that bind to a specific antigen can be obtained by methods known to those skilled in the art (see, e.g., Kohler et al., Nature 256:495, 1975; Coligan et al. (eds.), Current Protocols in Immunology, 1:2.5.12.6.7 (John Wiley & Sons 1991); U.S. Pat. Nos. RE 32,011, 4,902,614, 4,543,439, and 4,411,993; Monoclonal Antibodies, Hybridomas: A New Dimension in Biological Analyses, Plenum Press, Kennett, McKearn, and Bechtol (eds.) (1980); and Antibodies: A Laboratory Manual, Harlow and Lane (eds.), Cold Spring Harbor Laboratory Press (1988); Picksley et al., "Production of monoclonal antibodies against proteins expressed in E. coli", in DNA Cloning 2: Expression Systems, 2nd ed., Glover et al. (eds.), p. 93 (Oxford University Press 1995). Antibody fragments may be derived therefrom using any suitable standard technique, such as proteolytic digestion, or optionally by proteolytic digestion (e.g., using papain or pepsin) followed by mild reduction of disulfide bonds and alkylation. Alternatively, such fragments may also be generated by recombinant genetic engineering techniques as described herein.

[0228] Monoclonal antibodies can be obtained by injecting an animal, such as a rat, hamster, rabbit or preferably a mouse, with an immunogen comprising human M6PR or a fragment thereof, according to methods known in the art and described herein, including, for example, transgenic or knockout animals known in the art. The presence of specific antibody production can be monitored after the initial injection and / or after the booster injection by obtaining a serum sample and detecting the presence of antibodies that bind to human M6PR or peptide using any of several immunoassay methods known in the art and described herein. Lymphoid cells, most commonly cells from the spleen or lymph nodes, are removed from the animal producing the desired antibody to obtain B-lymphocytes. The B lymphocytes are then fused with a drug-sensitized myeloma cell fusion partner, preferably a partner that is homologous to the immunized animal and optionally has other desired characteristics (e.g., cannot express endogenous Ig gene products, e.g., P3X63-Ag 8.653 (ATCC No.CRL 1580); NSO, SP20), to produce hybridomas, which are immortal eukaryotic cell lines.

[0229] Lymphoid (e.g., spleen) cells and myeloma cells can be combined with a membrane fusion-promoting agent (such as polyethylene glycol or a nonionic detergent) for a few minutes and then plated at low density on a selective medium that supports the growth of hybridoma cells but does not support unfused myeloma cells. The preferred selective medium is HAT (hypoxanthine, aminopterin, thymidine). After a sufficient time (usually about one to two weeks), cell colonies are observed. Single colonies are isolated, and any of a variety of immunoassays known in the art and described herein can be used to test the binding activity of antibodies produced by the cells to human M6PR. Clone hybridomas (e.g., by limiting dilution cloning or by soft agar plaque separation), and select and culture positive clones that produce antibodies specific to M6PR. Monoclonal antibodies from hybridoma cultures can be isolated from the supernatant of hybridoma cultures.

[0230] An alternative method for producing murine monoclonal antibodies is to inject hybridoma cells into the peritoneal cavity of syngeneic mice (e.g., mice that have been treated (e.g., treated with pristane antigens) to promote the formation of ascites containing the monoclonal antibodies). Monoclonal antibodies can be isolated and purified by a variety of well-established techniques. Such separation techniques include affinity chromatography using protein-A agarose, size exclusion chromatography, and ion exchange chromatography (see, e.g., Coligan, pp. 2.7.1 to 2.7.12 and pp. 2.9.1 to 2.9.3; Baines et al., "Purification of Immunoglobulin G (IgG)", in Methods in Molecular Biology, Vol. 10, pp. 79 to 104 (The Humana Press, Inc. 1992)). Monoclonal antibodies can be purified by affinity chromatography using appropriate ligands selected based on the specific properties of the antibody (e.g., heavy or light chain isotype, binding specificity, etc.). Examples of suitable ligands immobilized on a solid support include protein A, protein G, constant region (light or heavy chain) antibodies, anti-idiotypic antibodies and M6PR binding proteins, or fragments or variants thereof.

[0231] The antibodies of the present disclosure may also be humanized or fully human monoclonal antibodies.

[0232] 6.7. pH-dependent antigen binding

[0233] In one aspect, the invention provides an antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to an internalization domain of a cation-independent mannose 6-phosphate receptor (CI-M6PR) that facilitates intracellular transport of the ABP, wherein the binding of the ABP to the CI-M6PR is pH-dependent. In such cases, the first antigen binding portion preferentially dissociates from the antigen in intracellular compartments having a lower pH.

[0234] In some embodiments, the first antigen binding moiety binds to CI-M6PR with a higher affinity at pH 7.4 than at pH 6.0. In some embodiments, the first antigen binding moiety binds to CI-M6PR with a higher affinity at pH 7.4 than at pH 5.5. In some embodiments, the first antigen binding moiety binds to CI-M6PR with a higher affinity at pH 7.4 than at pH 5.0.

[0235] In some embodiments, the antigen binding portion is released from CI-M6PR at a pH of 7.4 or less. In some embodiments, the antigen binding portion is released from CI-M6PR at a pH of 6.0 or less. In some embodiments, the antigen binding portion is released from CI-M6PR at a pH of 5.5 or less. In some embodiments, the antigen binding portion is released from CI-M6PR at a pH of 5.0 or less.

[0236] In some embodiments, the K of the first antigen binding moiety at pH 6.0 / pH 7.4 is D Ratio or K off The ratio is 1, 2, 5, 10, 20, 25, 50, 75 or 100 or more.

[0237] In some embodiments, the first antigen binding moiety has a K at pH 5.5 / pH 7.4. D Ratio or K off The ratio is 1, 2, 5, 10, 20, 25, 50, 75 or 100 or more.

[0238] In some embodiments, the K of the first antigen binding moiety at pH 5.0 / pH 7.4 is D Ratio or K off The ratio is 1, 2, 5, 10, 20, 25, 50, 75 or 100 or more.

[0239] 6.8. Internalization Domain

[0240] In some embodiments, the antigen binding domain specifically binds to the internalization domain of the target molecule. The internalization domain of the target molecule promotes, participates in, or otherwise contributes to the intracellular transport of ABP. In some embodiments, the internalization domain of the target molecule promotes, participates in, or otherwise contributes to the transport of ABP to the endolysosomal compartment. In some embodiments, when the target molecule is present on the surface of the cell, after promoting, participating in, or otherwise contributing to the internalization of ABP, the internalization domain of the target molecule promotes, participates in, or otherwise contributes to the recycling of the target molecule to the surface of the cell.

[0241] In some embodiments, when the antigen binding domain specifically binds to the internalization domain of the target molecule, the binding activates the target molecule, whereby the activated target molecule promotes, participates in, or otherwise contributes to the intracellular transport of the ABP.

[0242] 6.9. Target binding part

[0243] In one aspect, the present invention provides an antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to the internalization domain of a cation-independent mannose 6-phosphate receptor (CI-M6PR) that facilitates intracellular transport of the ABP, and a target binding portion that specifically binds to a target molecule.

[0244] In some embodiments, the ABP is an aptamer that specifically binds to a target molecule (such as a target protein). In some embodiments, the ABP is a peptide or protein (e.g., a peptide binding motif, a protein domain, an engineered polypeptide, or a glycoprotein) that specifically binds to a target molecule (such as a target protein). In some embodiments, the ABP is an antibody or antibody fragment that specifically binds to a target molecule (such as a target protein). In some embodiments, Y is a polynucleotide or oligonucleotide that specifically binds to a target molecule (such as a target protein or a target nucleic acid).

[0245] In some embodiments, peptides or proteins that specifically bind to a target molecule include, but are not limited to, peptide binding motifs, protein domains, engineered polypeptides, or glycoproteins.

[0246] In some embodiments, the target binding moiety is an aptamer that specifically binds to a target molecule. An aptamer is a nucleic acid molecule that has a specific binding affinity to a molecule through interactions other than classical Watson-Crick base pairing. An aptamer can specifically bind to a selected target and regulate the activity of the target (e.g., binding to an aptamer can block the ability of a target molecule to exert its effect). A typical aptamer is 10 to 15 kDa (30 to 45 nucleotides) in size, binds to its target with sub-nanomolar affinity, and identifies closely related targets (e.g., an aptamer will not typically bind to other proteins from the same gene family). An aptamer binds to a target molecule via interactions that drive affinity and specificity (e.g., hydrogen binding, electrostatic complementarity, hydrophobic contact, steric repulsion).

[0247] In some embodiments, the target binding moiety is a ligand that specifically binds to a target molecule. In some embodiments, a ligand is a molecule that can regulate signal transduction. In some embodiments, a ligand is a cytokine (e.g., a proinflammatory cytokine or an anti-inflammatory cytokine). Non-limiting examples of cytokines include IL-1 family cytokines (non-limiting examples include: IL-1A, IL-1B, IL-18, and IL-33), TNF family cytokines (non-limiting examples include: TNFα, TNFβ, and 4-1BBL), interferon family cytokines (non-limiting examples include: IFNα (IFNA), IFNγ (IFNG), and IFNβ (IFNB)), IL-6 family cytokines (non-limiting examples include: IL-6, IL-11, and LIF), IL-10 family cytokines (non-limiting examples include: IL-10, IL-19, IL-20, IL-22), and TGFβ family cytokines (e.g., TGFβ1, TGFβ2, and TGFβ3).

[0248] In some embodiments, the target binding moiety is a second antigen binding moiety that specifically binds to a target molecule different from the first antigen binding moiety.

[0249] In some embodiments, the target binding moiety is a second antigen binding moiety that specifically binds to the same target molecule as the first antigen binding moiety. In such cases, the second antigen binding domain can be any of the second antigen binding domains described herein. In some embodiments, the target binding moiety is a second antigen binding moiety that specifically binds to the same target molecule as the first antigen binding moiety but to a peptide different from the first antigen binding moiety.

[0250] In some embodiments, the target binding moiety is a small molecule.

[0251] In some embodiments, the target binding moiety is optionally conjugated to the first antigen binding moiety via a linker, such as any of the linkers described herein. In some embodiments, the target binding moiety is conjugated to the first antigen binding moiety using a linker, such as any of the linkers described herein. In some embodiments, the linker is selected based on the type of target binding moiety.

[0252] In some embodiments, the target binding portion is conjugated to the first antigen binding portion using a linker selected as described in the following documents: U.S. Patent Nos. 7,837,980B2; 8,906,376B2; 10,201,615B2; 10,933,112B2; U.S. Patent Publication Nos. 2019 / 0290775A1; 2022 / 0111066A1; and PCT Publication No. WO 2021 / 259506A1.

[0253] In some embodiments in which the target binding moiety is a small molecule, the small molecule can be coupled to the antigen binding protein at an activatable site. Suitable activatable sites include conjugation points such as thiol groups, amino groups (e.g., the epsilon amino group of a lysine residue or at the N-terminus of a protein), vicinal hydroxyl groups (1,2-diols) (e.g., oxidized carbohydrates), and carboxyl groups (e.g., the C-terminus of a protein, aspartic acid and glutamic acid residues, and carbohydrates such as sialic acid residues).

[0254] In some embodiments where the target binding moiety is a small molecule, the small molecule can be directly coupled to the conjugation point on the antigen binding protein. For example, the drug can be attached by alkylation of the ε-amino group of antibody lysine, reductive amination of oxidized carbohydrates or reaction with hydrazides, transesterification between hydroxyl and carboxyl groups, amidation at amino or carboxyl groups, and conjugation with thiols (e.g., interchain thiols) or by alkylation of lysine with 2-iminothiolane for example. Suitable methods for conjugating drugs to conjugation points are disclosed in, for example, the following documents: Current Protocols in Protein Science (John Wiley & Sons, Inc.), Chapter 15: Chemical Modifications of Proteins, which is incorporated herein by reference in its entirety.

[0255] In some embodiments where the target binding moiety is a small molecule, the small molecule can be directly coupled to the antigen binding protein via another molecule (such as a joint). For example, the drug can also be conjugated via a maleimide group coupled to a thiol group in, for example, an antibody hinge region. In some embodiments, an antigen binding protein (e.g., antibody) conjugate can be manufactured in the following manner: reacting a maleimide-derived drug with an antigen binding domain (e.g., antibody). In some embodiments, an antigen binding protein (e.g., antibody) conjugate can be manufactured in the following manner: reducing the antigen binding domain (e.g., antibody) to produce reduced antibodies, producing amine drugs, derivatizing amine drugs with maleimide to produce maleimide-derived drugs, and reacting maleimide-derived drugs with antigen binding domains (e.g., antibodies).

[0256] In some embodiments where the target binding moiety is a small molecule, two or more small molecules can be coupled (eg, directly or indirectly) to the antigen binding protein.

[0257] 6.9.1. Target molecules

[0258] In some embodiments where the ABP comprises a target binding moiety that specifically binds to a target molecule, the target molecule is selected from the group consisting of: a soluble extracellular target molecule, a cell surface target molecule, a toxin, a pathogen, and a therapeutic agent.

[0259] In some embodiments, the target molecule is an immunoglobulin. In some embodiments, the target molecule is immunoglobulin E (IgE).

[0260] In some embodiments, the target molecule is a receptor in the epidermal growth factor receptor (EGFR) family.

[0261] 6.10. Goods

[0262] In one aspect, the present invention provides an antigen binding protein (ABP), comprising a first antigen binding moiety that specifically binds to an internalization domain of a cation-independent mannose 6-phosphate receptor (CI-M6PR) that promotes intracellular transport of the ABP, and a cargo portion. In some embodiments, the cargo portion is fused to the antigen binding protein. In some embodiments, the cargo portion is fused to the N-terminus of the antigen binding protein. For example, when the ABP comprises a Fab fragment and a cargo portion, the cargo portion is fused to the N-terminus of the ABP. In some embodiments, the cargo portion is fused to the C-terminus of the antigen binding protein. For example, when the ABP comprises a Fab-Fc fragment and the Fc domain is attached to the C-terminus of the Fab fragment, the cargo portion is fused to the C-terminus of the Fc domain (e.g., the domains are sorted from the N-terminus to the C-terminus: Fab-Fc-cargo portion).

[0263] In another aspect, the present invention provides an antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to the internalization domain of a cation-independent mannose 6-phosphate receptor (CI-M6PR) that facilitates intracellular transport of the ABP, a cargo portion fused to the antigen binding protein, and a target binding portion that specifically binds to a target molecule (e.g., any of the target molecules described herein).

[0264] In some embodiments, the cargo moiety is selected from the group consisting of a small molecule, a drug, a dye, a fluorophore, a monosaccharide, a disaccharide, a trisaccharide, a biomolecule, a nanoparticle, and a viral composition.

[0265] In some embodiments, the cargo moiety is a biomolecule selected from the group consisting of a polypeptide, a polynucleotide, an aptamer, a polysaccharide, a glycoprotein, a lipid, an enzyme, an antibody, and an antibody fragment.

[0266] In some embodiments, the cargo moiety is optionally conjugated to the first antigen binding moiety via a linker, such as any of the linkers described herein. In some embodiments, the cargo moiety is conjugated to the first antigen binding moiety using a linker, such as any of the linkers described herein. In some embodiments, the linker is selected based on the type of target binding moiety.

[0267] 6.11. Connectors

[0268] The terms "joint", "linking moiety" and "linking group" are used interchangeably and refer to a linking moiety that covalently connects two or more moieties or compounds (such as ligands and other moieties of interest). In some cases, the joint is divalent and connects two moieties. In some cases, the joint is a trivalent or higher multivalent branched linking group. In some cases, the joint connecting two or more moieties has a length of 500 atoms or less (such as 400 atoms or less, 300 atoms or less, 200 atoms or less, 100 atoms or less, 80 atoms or less, 60 atoms or less, 50 atoms or less, 40 atoms or less, 30 atoms or less, or even 20 atoms or less) linear or branched main chain, for example, as measured between two or more moieties. The linking moiety can be a covalent bond, which connects two groups or a linear or branched chain of length between 1 and 500 atoms (e.g., a length of about 1, 2, 3, 4, 5, 6, 8, 10, 12, 14, 16, 18, 20, 30, 40, 50, 100, 150, 200, 300, 400 or 500 carbon atoms), wherein the joint can be linear, branched, cyclic or single atom. In some cases, one, two, three, four, five or more, ten or more or even more carbon atoms of the joint main chain can be optionally replaced by heteroatoms (e.g., sulfur, nitrogen or oxygen heteroatoms). In some cases, when the joint includes a PEG group, every three atoms of this section of the joint main chain are replaced by oxygen. The key between the main chain atoms can be saturated or unsaturated, and there is usually no more than one, two or three unsaturated bonds in the joint main chain. The joint can include one or more substituent groups, such as alkyl, aryl or alkenyl groups. The linker may include, but is not limited to, one or more of the following: oligo(ethylene glycol), ether, thioether, disulfide, amide, carbonate, carbamate, tertiary amine, alkyl (may be straight or branched), such as methyl, ethyl, n-propyl, 1-methylethyl (isopropyl), n-butyl, n-pentyl, 1,1-dimethylethyl (tert-butyl), etc. The linker backbone may include a cyclic group, such as an aryl group, a heterocyclic ring, a cycloalkyl group, or a heterocyclic group, wherein 2 or more atoms (e.g., 2, 3, or 4 atoms) of the cyclic group are included in the backbone.

[0269] In some embodiments, a "linker" or connecting moiety is derived from a molecule having two reactive ends, one reactive end for conjugation with a moiety of interest, such as a biomolecule (e.g., an antibody), and the other reactive end for conjugation with a moiety that binds to a cell surface receptor M6PR. When the moiety of interest is a polypeptide, the polypeptide conjugation reactive end of the linker is in some cases a site capable of conjugation to the polypeptide through a cysteine ​​thiol or lysine amine group on the polypeptide, and thus can be a thiol-reactive group (such as maleimide or dibromomaleimide or as defined herein) or an amine-reactive group (such as an active ester (e.g., perfluorophenyl ester or tetrafluorophenyl ester) or as defined herein).

[0270] In certain embodiments of the formula described herein, the joint includes one or more straight or branched carbon moieties and / or polyether (e.g., ethylene glycol) moieties (e.g., -CH2CH2O- repeating units) and combinations thereof. In certain embodiments, these joints optionally have amide bonds, urea or thiourea bonds, carbamate bonds, ester bonds, amino bonds, ether bonds, thioether bonds, sulfhydryl bonds or other heterofunctional bonds. In certain embodiments, the joint comprises one or more of a carbon atom, a nitrogen atom, a sulfur atom, an oxygen atom and combinations thereof. In certain embodiments, the joint comprises one or more of an ether bond, a thioether bond, an amine bond, an amide bond, a carbon-carbon bond, a carbon-nitrogen bond, a carbon-oxygen bond, a carbon-sulfur bond and combinations thereof. In certain embodiments, the joint comprises a linear structure. In certain embodiments, the joint comprises a branched structure. In certain embodiments, the joint comprises a cyclic structure.

[0271] In certain embodiments, the linker includes one or more linking moieties independently selected from the group consisting of: -C1-20-alkylene-, -NHCO-C1-6-alkylene-, -CONH-C1-6-alkylene-, -NHC1-6-alkylene-, -NHCONH-C1-6-alkylene-, -NHCSNH-C1-6-alkylene-, -C1-6-alkylene-NHCO-, -C1-6-alkylene-CONH-, -C1-6-alkylene-NH-, -C1-6-alkylene-NHCONH-, -C1-6-alkylene-NHCSNH- , O(CH2)p-, -(OCH2CH2)p-, -NHCO-, -CONH-, -NHSO2-, -SO2NH-, -CO-, -SO2-, -O-, -S-, monocyclic heteroaryl (e.g., 1,2,3-triazole), monocyclic aryl (e.g., phenyl, e.g., 1,4-linked phenyl or 1,3-linked phenyl), monocyclic heterocycle (e.g., pyrrolidine-2,5-dione, piperazine or piperidine ring as described herein), amino acid residues (naturally or non-naturally occurring amino acid residues), -NH- and -NMe-, wherein each p is independently 1 to 50.

[0272] 6.11.1. Chemoselective Linking Groups

[0273] A chemoselective linking group is a group having a reactive functional group or functional groups that can be conjugated with a compatible group of a second moiety. For example, a chemoselective linking group (or a precursor thereof) can be one of a pair of groups associated with a conjugation chemistry such as azide-alkyne click chemistry, copper-free click chemistry, Staudinger ligation, tetrazine ligation, hydrazine-iso-Pictet-Spengler (HIPS) ligation, cysteine-reactive ligation chemistry (e.g., thiol-maleimide, thiol-haloacetamide, or alkyne hydrothiolation), amine-active ester coupling, reductive amination, dialkyl squarate chemistry, etc.

[0274] Chemoselective linking groups that can be used to link the two moieties include, but are not limited to, amino groups (e.g., the N-terminal amino group of a polypeptide or a lysine side chain group), azido groups, arylazide groups, alkynyl groups (e.g., ethynyl or cyclooctyne or derivatives), active esters (e.g., N-hydroxysuccinimide (NHS) esters, sulfo-NHS esters or PFP esters or thioesters), haloacetamides (e.g., iodoacetamide or bromoacetamide), chloroacetyl groups, bromoacetyl groups, hydrazides, maleimides, vinyl sulfones, 2-sulfonylpyridines, cyano-alkynes, thiols (e.g., cysteine ​​residues), disulfides or protected thiols, isocyanates, isothiocyanates, aldehydes, ketones, alkoxyamines, hydrazides, aminooxy groups, phosphines, HIPS hydrazino-indolyl groups or aza-HIPS hydrazino-pyrrolo-pyridinyl groups, tetrazines, cyclooctenes, squarates, and the like.

[0275] The conjugates of the present disclosure can be made using a variety of linkers and / or bifunctional protein coupling agents such as BMPS, EMCS, GMBS, HBVS, LC-SMCC, MBS, MPBH, SBAP, SIA, SIAB, SMCC, SMPB, SMPH, sulfo-EMCS, sulfo-GMBS, sulfo-KMUS, sulfo-MBS, sulfo-SIAB, sulfo-SMCC, sulfo-SMPB and SVSB (succinimidyl-(4-vinyl sulfone) benzoate). The present disclosure further contemplates that the conjugates described herein can be prepared using any suitable method as disclosed in the art (see, e.g., Bioconjugate Techniques (Hermanson ed., 2nd ed. 2008)).

[0276] In some embodiments, when the two groups are contacted under suitable conditions (e.g., copper-free click chemistry conditions), the chemoselective linking group can spontaneously conjugate with compatible chemical groups. In some cases, when the two groups are contacted in the presence of a catalyst or other reagent (e.g., copper-catalyzed click chemistry conditions), the chemoselective linking group can conjugate with compatible chemical groups. In some embodiments, the chemoselective linking group is a photoactive linking group. For example, upon irradiation with ultraviolet light, the diaziridinyl group can form a reactive carbenes that can be inserted into the CH, NH, and OH bonds of the second portion.

[0277] 6.12. Sequence

[0278] Antibodies 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11 comprise a heavy chain V(J)D polynucleotide and a light chain V(J)D polynucleotide (e.g., of antibody clones 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12, and 53B11). Antibodies 3C7, 6D1, 6D12, 6F1, 6H10, 8D8, 10H5-2, 11B12, 12D5, 16H4, 17F11, 18G4, 21D5, 43C8, 45A12 and 53B11 comprise the variable light chain sequences and variable heavy chain sequences listed in Tables 5 to 6. The CDR sequences in the light and heavy chains are also provided with specific SEQ ID NOs.

[0279] Table 5: Full-length M6PR ABP variable heavy chain amino acid sequence

[0280]

[0281]

[0282] Table 6: Full-length M6PR ABP variable light chain amino acid sequence

[0283]

[0284]

[0285] Table 7: M6PR variable heavy chain ABP CDR (HCDR)

[0286]

[0287]

[0288] Table 8: M6PR variable light chain ABP CDR (LCDR)

[0289]

[0290]

[0291] Table 9: Exemplary anti-EGFR antibody sequences

[0292]

[0293]

[0294]

[0295]

[0296] Table 11: M6PR variable heavy chain ABP framework region (FR) sequences

[0297]

[0298]

[0299] Table 12: M6PR variable light chain ABP framework region (FR) sequences

[0300]

[0301]

[0302]

[0303] 6.13. Pharmaceutical compositions

[0304] Also provided are pharmaceutical compositions containing the proteins and polypeptides of the invention. Such compositions comprise a therapeutically or prophylactically effective amount of the polypeptide or protein in admixture with pharmaceutically acceptable materials and physiologically acceptable formulation materials.

[0305] Pharmaceutical compositions may contain formulation materials for modifying, maintaining or preserving, for example, the pH, osmotic pressure, viscosity, clarity, color, isotonicity, odor, sterility, stability, rate of dissolution or release, adsorption or penetration of the composition.

[0306] Suitable formulation materials include, but are not limited to, amino acids (such as glycine, glutamine, asparagine, arginine or lysine); antimicrobial agents; antioxidants (such as ascorbic acid, sodium sulfite or sodium bisulfite); buffers (such as borates, bicarbonates, Tris-HCl, citrates, phosphates, other organic acids); bulking agents (such as mannitol or glycine), chelating agents (such as ethylenediaminetetraacetic acid (EDTA)); complexing agents (such as caffeine, polyvinylpyrrolidone, β-cyclodextrin or hydroxypropyl-β-cyclodextrin); fillers; monosaccharides; disaccharides and other carbohydrates (such as glucose, mannose or dextrin); proteins (such as serum albumin, gelatin or immunoglobulins); colorants; flavorings and diluents; emulsifiers; hydrophilic polymers (such as polyvinylpyrrolidone); low molecular weight polypeptides; salt-forming Counterions (such as sodium); preservatives (such as benzalkonium chloride, benzoic acid, salicylic acid, thimerosal, phenylethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid or hydrogen peroxide); solvents (such as glycerol, propylene glycol or polyethylene glycol); sugar alcohols (such as mannitol or sorbitol); suspending agents; surfactants or wetting agents (such as Pluronics, PEG, sorbitan esters, polysorbates (such as polysorbate 20, polysorbate 80), triton, tromethamine, lecithin, cholesterol, tyloxapal); stability enhancers (sucrose or sorbitol); tonicity enhancers (such as alkali metal halides (preferably sodium chloride or potassium chloride), mannitol, sorbitol); delivery vehicles; diluents; excipients and / or pharmaceutical adjuvants. Neutral buffered saline or saline mixed with the same serum albumin is an example of a suitable diluent. Preservatives such as benzyl alcohol may also be added according to appropriate industry standards. The composition may be formulated as a lyophilisate using an appropriate excipient solution (e.g., sucrose) as a diluent. Suitable components are nontoxic to the recipient at the dose and concentration employed. Further examples of components that may be used in pharmaceutical formulations are presented in the following literature: Remington's Pharmaceutical Sciences, 16th edition (1980) and 20th edition (2000), Mack Publishing Company, Easton, PA.

[0307] Optionally, the composition additionally comprises one or more physiologically active agents, for example, anti-angiogenic substances, chemotherapeutic substances (such as capecitabine, 5-fluorouracil or doxorubicin), analgesic substances, etc., non-exclusive examples of which are provided herein. In various specific embodiments, in addition to internalized receptor binding proteins, cell surface target molecules and / or soluble target molecules, the composition further comprises one, two, three, four, five or six physiologically active agents.

[0308] In another embodiment of the present invention, the compositions disclosed herein can be formulated as neutral or salt forms. Exemplary pharmaceutically acceptable salts include acid addition salts (formed with the free amino groups of proteins), and they are formed with inorganic acids (such as, for example, hydrochloric acid or phosphoric acid) or organic acids (such as acetic acid, oxalic acid, tartaric acid, mandelic acid, etc.). The salts formed by free carboxyl groups can also be derived from inorganic bases (such as, for example, sodium hydroxide, potassium hydroxide, ammonium hydroxide, calcium hydroxide or ferric hydroxide) and organic bases (such as isopropylamine, trimethylamine, histidine, procaine, etc.). When formulated, the solution will be administered in a therapeutically effective amount in a manner compatible with the dosage formulation.

[0309] The carrier may further comprise any and all solvents, dispersion media, vehicles, coatings, diluents, antibacterial and antifungal agents, isotonic and absorption slow-release agents, buffers, carrier solutions, suspensions, colloids, etc. The use of such media and agents for pharmaceutically active substances is well known in the art. Unless any conventional media or agents are incompatible with the active ingredient, their use in therapeutic compositions is considered. Supplementary active ingredients may also be incorporated into the composition. The phrase "pharmaceutically acceptable" refers to molecular entities and compositions that do not produce allergic or similar adverse reactions when administered to humans.

[0310] The optimal pharmaceutical composition will be determined by those skilled in the art based on, for example, the intended route of administration, delivery form, and desired dosage. See, for example, Remington's Pharmaceutical Sciences, supra. Such compositions may affect the physical state, stability, in vivo release rate, and in vivo clearance rate of the polypeptide. For example, a suitable composition may be water for injection, a physiological saline solution for parenteral administration.

[0311] 6.14. Usage

[0312] In some embodiments, the antigen binding proteins of the present disclosure may be used to treat a subject suffering from a disease or disorder associated with the presence of excess cell surface target molecules and / or soluble target molecules.

[0313] In some embodiments, the antigen binding proteins of the present disclosure may be used to treat subjects suffering from allergic conditions including, but not limited to, allergic asthma, nasal polyps, and urticaria.

[0314] Therapeutic antibodies that specifically bind to internalizing receptors can be used. In addition, therapeutic antibodies that specifically bind to internalizing receptors and soluble target molecules or cell surface target molecules can be used.

[0315] In vivo and / or in vitro assays may optionally be employed to help identify optimal dosage ranges. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems.

[0316] Aspects of the present disclosure include methods of producing an isolated multispecific antigen-binding protein (ABP), comprising an antigen-binding portion that specifically binds to an internalizing receptor and promotes intracellular transport of the ABP through the internalizing receptor. In some embodiments, the antigen-binding portion specifically binds to an epitope on an antigen that promotes intracellular transport of the ABP through the internalizing receptor.

[0317] Aspects of the present disclosure provide methods of treating a disease, comprising the step of administering to a subject in need thereof an effective amount of a multispecific ABP of the present disclosure or a pharmaceutical composition of the present disclosure.

[0318] Depending on the condition to be treated, the composition can be administered alone or in combination with other treatments, simultaneously or sequentially. For example, the pharmaceutical composition can be administered in combination with one or more drugs targeting different soluble or cell surface target molecules.

[0319] 6.15. Examples

[0320] Below is the example of the specific embodiment of the present invention.These examples are provided for illustrative purposes only, and are not intended to limit the scope of the present invention in any way.Efforts have been made to ensure the accuracy of the numerals (e.g., amount, temperature, etc.) used, but of course some experimental errors and deviations should be allowed.

[0321] Unless otherwise indicated, the practice of the present invention will adopt conventional methods of protein chemistry, biochemistry, recombinant DNA technology and pharmacology within the technology of the art. Such techniques are fully explained in the literature. See, for example, TECreighton, Proteins: Structures and Molecular Properties (WH Freeman and Company, 1993); A Lehninger, Biochemistry (Worth Publishers, Inc., current edition); Sambrook et al., Molecular Cloning: A Laboratory Manual (2nd edition, 1989); Methods In Enzymology (S. Colowick and N. Kaplan ed., Academic Press, Inc.); Remington's Pharmaceutical Sciences, 18th edition (Easton, Pennsylvania: Mack Publishing Company, 1990); Carey and Sundberg Advanced Organic Chemistry 3rd edition (Plenum Press) Volumes A and B (1992). In addition, the methods for generating and selecting antibodies explained in the following references can be adopted: Adler et al., A natively paired antibody library yields drug leads with higher sensitivity and specificity than a randomly paired antibody library, MAbs (2018), and Adler et al., Rare, high-affinity mouse anti-PD-1 antibodies that function in checkpoint blockade, discovered using microfluidics and molecular genomics, MAbs (2017), which are incorporated herein by reference in their entirety.

[0322] 6.15.1. Example 1: Generation of anti-M6PR antibodies

[0323] Antibody generation of monoclonal antibodies that specifically bind to the MP6R receptor uses a hybridoma-based antibody discovery protocol that uses recombinant antigens of human M6PR and various cell lines that support mouse immunization and antibody screening. Mouse immunization is performed using proprietary hyperimmune DiversimAb / DiverGimAb mice (Abveris). As part of hybridoma cryopreservation, supernatant samples, and antibody sequencing, hybridomas are characterized by flow cytometry, ELISA, and Octet biolayer interferometry (BLI).

[0324] Materials and methods

[0325] Each of the following protocols was used for the corresponding assay described herein.

[0326] Titer test (ELISA) method: Indirect ELISA protocol (brief description)

[0327] 1. Coat high binding ELISA plates with antigen overnight at 4°C.

[0328] 2. Aspirate and block the wells with 1x PBS containing 2% BSA for 1 hour.

[0329] 3 Aspirate, add antiserum dilution in blocking solution, and incubate at room temperature for 1 hour.

[0330] 4 Wash wells 4 times in 1x PBST.

[0331] 5. Add HRP-conjugated secondary antibody in blocking solution and incubate at room temperature for 45 minutes.

[0332] 6 Wash wells 5 times in 1x PBST.

[0333] 7. Add TMB substrate followed by stop solution. Read at 450 nm.

[0334] Note: (-)ctrl NMS = normal mouse serum

[0335] Titer Testing (Flow Cytometry) Cohort 1, 2 Methods: FACS Staining Protocol (Overview of Key Steps)

[0336] 1. Seed 100,000 cells per well

[0337] 2 Resuspend in 50uL titrated serum (starting from 1:100, 3-fold dilution, a total of 11 points).

[0338] 3Incubate on ice for 1 hour.

[0339] 4. Resuspend in 50uL secondary antibody (1:500 anti-muFc-A488 and 1:500 Live / Dead NIR Viability).

[0340] 5 Incubate on ice for 30 min.

[0341] 6 Read at least 5000 events on iQue.

[0342] Primary screening (flow) method: iQue flow cytometry (key steps)

[0343] 110k cells were seeded per well.

[0344] 2Resuspend in 100uL culture medium.

[0345] 3Incubate on ice for 1 hour.

[0346] 4Resuspend in 50uL secondary antibody.

[0347] 5 Incubate on ice for 30 min.

[0348] 6Fix with 1% PFA.

[0349] 7 Read at least 5000 events on iQue.

[0350] Secondary screening (flow) method: iQue flow cytometry (key step).

[0351] 110k cells were seeded per well.

[0352] 2Resuspend in 100uL culture medium.

[0353] 3Incubate on ice for 1 hour.

[0354] 4Resuspend in 50uL secondary antibody.

[0355] 5 Incubate on ice for 30 min.

[0356] 6Fix with 1% PFA.

[0357] 7 Read at least 5000 events on iQue.

[0358] Secondary Screening (ELISA) Method: Indirect ELISA Protocol

[0359] 1Coat a high binding ELISA plate with antigen overnight at 4°C.

[0360] 2. Aspirate and block the wells with 1x PBS containing 2% BSA for 1 hour.

[0361] 3 Aspirate, add suspension 1:1 in blocking buffer, and incubate at room temperature for 1 hour.

[0362] 4 Wash wells 4 times in 1x PBST.

[0363] 5. Add HRP-conjugated secondary antibody in blocking solution and incubate at room temperature for 45 minutes.

[0364] 6 Wash wells 5 times in 1x PBST.

[0365] 7. Add TMB substrate followed by stop solution. Read at 450 nm.

[0366] (-)ctrl H10 = medium negative control.

[0367] (+)ctrl H11 = positive control of cohort 1 antiserum.

[0368] (+)ctrl H12 = positive control of cohort 2 antiserum.

[0369] Characterization of K562 cells for antibody screening

[0370] Wild-type K562 cells expressing M6PR and M6PR knockout (KO) K562 cells without the M6PR gene were used for antibody screening. Figure 4 Cell surface and intracellular staining of human M6PR in WT and M6PR-KO K562 cells are shown in FIG.

[0371] Antibodies and fluorescent stains used in the antibody screening assay included anti-M6PR (clone MEM238)-Alexafluor 647 (Abcam).

[0372] Cells were frozen, subsequently stained and fixed for cell surface staining. For intracellular staining, cells were fixed, permeabilized and stained.

[0373] Table 13: Reagents used for immunization and screening:

[0374]

[0375] Mouse immunization and titer testing

[0376] The mouse cohorts were immunized according to the following method and schedule and using the reagents described in Table 15. The mice were monitored for daily health. Two cohorts of mice (n=4 / cohort) of DiverSimab and DiverGimAb each received a priming dose of 100ug / animal recombinant human M6PR protein and were boosted with 20ug / animal recombinant human M6PR protein per week for 4 weeks. After the immunization schedule was completed, the titer of each mouse was assessed using recombinant antigens and confirmed via flow-based assays using human cell lines (K562WT, K562 M6PR-KO, Hela WT, and Hela M6PR-KO), and cross-screened for two mouse cell lines (Dipak MC38 and 4T1) ( Figure 5 ). Mice with high titers progressed to fusion.

[0377] Hybridoma fusion and antibody screening

[0378] Tissues including lymph nodes and / or spleen were harvested. High-efficiency electrofusion was used, and hybridomas were seeded and grown in culture before high-throughput primary screening (flow cytometry) of clones. Primary screening (flow cytometry) was performed using K562 cells (WT and M6PR-KO) for binding confirmation. Clones were evaluated by ELISA to assess binding of each clone to human and mouse M6PR antigens. Clones that were positive for binding to the human M6PR antigen were subjected to secondary flow-based screening using human cell lines (K562 WT, K562M6PR-KO, Hela WT, and HelaM6PR-KO) and murine cell lines (Dipak MC38 and 4T1) to confirm specific antibodies that bind to human M6PR. No mouse cross-reactive monoclonal antibodies were identified; all monoclonal antibodies specifically bound to human M6PR.

[0379] The first 192 clones were subjected to affinity measurement via Octet to measure on / off rates and K against human antigens. D All 192 clones were sequenced using the Sanger sequencing process to generate paired V H / V L Variable region sequences, antibody isotype data and germline families. Each anti-mouse biosensor was loaded with mIgG (and poly-mIgG control) from the supernatant ( Figure 6 ). The baseline of each biosensor was measured. The loaded biosensors were then allowed to bind to recombinant His-tagged human M6PR and the association rate was measured. The biosensors were moved to the buffer wells to measure the dissociation rate. The results of representative clones are shown in Figure 7 The biosensor was regenerated for measuring additional analyte interactions (mouse M6PR-His6 and off-target binding to His). Clones with affinity for human M6PR and showing no binding to the His control and / or mouse M6PR were binned to determine epitope diversity.

[0380] 6.15.2. Example 2: Evaluation of anti-CI-M6PR monoclonal antibodies produced by assaying hybridoma supernatants

[0381] Screening of CI-M6PR binding antibodies for uptake activity, such as Figure 8As shown. Since CI-M6PR binds to and internalizes IGF2, a non-binding antibody (KLH) fused to IGF2 was used as a positive control for uptake activity. K562 cells were seeded at 30,000 cells per well in 96-well tissue culture plates in 100 μl RPMI (Gibco #61870143), which contained 10% v / v FBS (VWR #89510-188), 2mM L-alanyl-L-glutamine dipeptide, 100 units / ml penicillin and 100ug / ml streptomycin (Gibco #15140148). Separately, 50 μl of CI-M6PR clone hybridoma supernatant was mixed with 50 μl of Zenon pHrodo-iFL IgG labeling reagent (Thermo Fisher #Z25609) and incubated at room temperature for 15 minutes. The hybridoma supernatant / pHrodo mixture was layered on top of K562 cells and incubated for 18 hours to allow uptake and lysosomal accumulation.Uptake was measured using a Novocyte Advanteon flow cytometer (Agilent Technologies) with a 488 nM laser and FITC detection configuration.

[0382] Table 14 below shows the binding affinity and cellular internalization of various anti-CI-M6PR monoclonal antibodies measured according to the methods described above.

[0383] Table 14: CI-M6PR monoclonal antibody characteristics

[0384]

[0385]

[0386]

[0387] 6.15.3. Example 3: Anti-MP6PR Antibodies Bind to Human M6PR

[0388] Thirty-one of the 96 anti-M6PR antibodies identified in Example 2 were isolated, cloned, and recombinantly expressed as monoclonal antibodies.

[0389] Binding affinity and kinetics

[0390] Each of the 31 antibodies was evaluated for its ability to bind CI-M6PR.Each of the anti-M6PR antibodies binding to CI-M6PR was also evaluated for binding affinity and kinetics.

[0391] Anti-M6PR antibodies bound to human CI-M6PR were measured by biolayer interferometry (BLI) using a GatorPrime instrument. Anti-M6PR antibodies were prepared in PBS buffer supplemented with 0.002% Tween-20, 0.02% BSA, and 0.005% NaN3, and immobilized on a sensor coated with anti-mouse IgG Fc antibody. The immobilized anti-M6PR antibody was then incubated with purified recombinant CI-M6PR (domains 1 to 9) proteins prepared in the same buffer at different concentrations. During the experiment, the reaction plate was maintained at 30 ° C and vibrated at 1,000 rpm. GatorOne software v2.10 was used to fit the data, where the response curves were compared in the association step. The 1:1 binding model was used to globally fit the treated association and dissociation curves to obtain the kinetic constants for each antibody.

[0392] Fig. 9A The ELISA results in G to G show that 16 of the 31 anti-M6PR antibodies bound to human M6PR.

[0393] Fig.10 The results in the literature demonstrate that the top 15 anti-M6PR antibodies exhibit a range of affinities (K D ) and dynamics (K on , K off ).

[0394] Epitope binning and domain mapping

[0395] Each of the 16 anti-M6PR antibodies identified as binding to human CI-M6PR ( Figures 9A to 9G ) were evaluated by epitope binning and domain mapping to determine the binding location of each antibody on CI-M6PR ( Fig.11 ).

[0396] To determine the epitope bins, anti-M6PR antibodies and CI-M6PR protein were immobilized to anti-mouse IgG Fc sensors according to the method described above. These probes were then incubated with a secondary anti-M6PR antibody. Antibodies that resulted in any further increase in signal as detected by the sensor were determined to be in a separate epitope bin from the antibody already immobilized on the sensor. Antibodies that resulted in a decrease in signal were determined to be in the same epitope bin as the antibody already immobilized on the sensor. See Fig. 12A (a non-limiting example of such an assay) and Fig. 12B (Example graph showing signal transitions for different epitope bins). Fig.13 As summarized in , 12 of the preferred anti-M6PR antibodies represent 8 separate epitope bins.

[0397] To perform domain mapping of anti-M6PR antibodies, a series of chimeric human / mouse CI-M6PR constructs were cloned and purified. Each of the 9 domains (D1 to D9) of human CI-M6PR was replaced individually with the equivalent domains of all 9 domains of mouse CI-M6PR. The N-terminus of each chimeric construct was labeled with a HIS tag.

[0398] Each of the nine HIS-tagged CI-M6PR domains was then recombinantly produced via HEK293 expression of the extracellular domain. Fig.14 The expression of each of the HIS-tagged D1 to D9 CI-M6PR domains is shown. The recombinantly expressed HIS-tagged D1 to D9 CI-M6PR domains were purified by harvesting the HEK293 culture medium and using a HisTrap affinity column. All domains were purified as non-aggregating monovalent entities of approximately 200 kDa. The instantaneous recovery rate was approximately 1 to 2 mg / mL.

[0399] BLI experiments were performed according to the method described above to test the binding of each anti-M6PR antibody to each chimeric CI-M6PR protein. Wild-type human CI-M6PR was used as a positive control. If the replacement of the specific domain of human CI-M6PR with the specific domain of mouse CI-M6PR resulted in loss of binding, the domain was identified as the domain of human CI-M6PR that binds to the given antibody.

[0400] like Fig.15A To B and Fig.15D Anti-M6PR antibody domains were mapped for clones 3C7, 6D1, 11B12, 17F11, 18G4, 21D5, and 53B11 as shown in Figures 1 to 3. Fig. 15C ) showed that binding to each of the chimeric CI-M6PR proteins was similar, meaning that the domain of human CI-M6PR that bound to the clone 8D8 antibody could not be determined using this assay.

[0401] Fig.16 A schematic diagram summarizing the domain mapping results of anti-M6PR antibody clones 3C7, 6D1, 11B12, 17F11, 18G4, 21D5, and 53B11 is shown in . For example, 53B11 binds to domain 1 on CI-M6PR, 11B12 binds to domain 4 on CI-M6PR, 18G4 binds to domain 5 on CI-M6PR, 3C7 and 6D1 each bind to domain 6 on CI-M6PR, 17F11 binds to domain 7 on CI-M6PR, and 21D5 binds to domain 8 on CI-M6PR.

[0402] 6.15.4. Example 4: Evaluation of pH-dependent dissociation of anti-M6PR Ab from CI-M6PR

[0403] Anti-M6PR antibodies were immobilized onto anti-mouse IgG1 Fc-coated sensors according to the method described above and incubated with human CI-M6PR protein prepared in citrate buffer (pH 7.4). For dissociation, the sensors were transferred to wells containing citrate buffer at pH 7.4, pH 6.0, or pH 5.0 (see Fig.17A ). Figures 17B to 17D Clone 17F11 ( Fig. 17B )、18G4( Fig. 17C ) and 21D5( Fig.17D ), where clones 17F11 and 21D5 showed stronger pH-dependent dissociation than clone 18G4.

[0404] To quantify pH-dependent dissociation, the calculated K values ​​for each antibody at pH 5.0 and pH 6.0 were subtracted from the calculated K values ​​for each antibody at pH 5.0 and pH 6.0. off Divided by the K at pH 7.4 off , to determine the fold increase in the off-rate for each antibody at the indicated pH. Fig.18 It was shown that the off-rates of antibody clones 17F11 and 21D5 were increased at low pH (pH 5.0).

[0405] 6.15.5. Example 5: Cellular Uptake of Anti-M6PR Antibodies

[0406] pH rodo uptake assay of 31 anti-M6PR antibodies

[0407] Recombinantly expressed anti-M6PR antibodies were evaluated for uptake activity using the pH rodo uptake assay ( Fig.19A ). In particular, mAb and Zenon pHrodo green Fab were pre-complexed at 50nM mAb and 100nM Zenono pHrodo green iFL (2x final concentration) in 100ul RPMI+10% fetal bovine serum (FBS) and 1x pen / strep for 30 minutes at room temperature. 100μl of each mAb:Zenon pHrodo Fab was added to 100μl RPMI+10% FBS and 1x pen / strep containing 100,000 K562 cells. The plates were incubated at 37°C with 5% CO2 for 24 hours. The cells were pelleted and washed once with FACS wash buffer (PBS+1% bovine serum albumin (BSA)). The uptake of anti-M6PR antibody was determined by measuring the pHrodo green signal in the FITC channel by flow cytometry.

[0408] Because CI-M6PR binds and internalizes IGF2, a non-binding antibody (KLH) fused to IGF2 was used as a positive control for uptake activity. The identified antibodies showed uptake activity greater than the KLH-IGF2 positive control. Fig.19B As shown, antibody clones 6D12, 6H10, 6D1, 17F11 and 21D5 showed greater uptake than KLH-IGF2.

[0409] Evaluation of anti-M6PR / anti-IgE bispecific antibodies for uptake

[0410] A subset of anti-M6PR antibodies were evaluated for uptake activity using the IgE-pH rodo uptake assay. Specifically, the M6PR-Oma bispecific antibody (BsAb) ( Figure 2A ) and IgE fluorescently labeled with Alexa Fluor 488 or AlexaFluor647 (IgE-AF488 or IgE-AF647) were pre-complexed for 30 minutes at 200nM each (2x final) in RPMI+10% FBS and 1x penicillin / streptomycin. A 6-fold log3 serial dilution was performed, and 100μl of each BsAb:IgE dilution was added to 100μl RPMI+10% FBS and 1x penicillin / streptomycin containing 100,000 K562 cells. The plate was incubated at 37°C with 5% CO2 for 24 hours. The cells were pelleted and washed once with FACS wash buffer (PBS+1% BSA). Anti-M6PR antibody uptake was determined by measuring the signal from AF488 and / or AF647 by flow cytometry.

[0411] A non-binding antibody (KLH) fused to omalizumab (Oma) was used as a negative control for uptake activity. The Oma-16590 (Dar4) bispecific antibody was used as a positive control for uptake activity. Figures 20A to 20F Uptake of the anti-M6PR-Oma bispecific antibody is shown. Fig. 20A and Fig. 20B The results shown in demonstrate that 6H10-Oma, 6D1-Oma, 8D8-Oma, and 17F11-Oma transport fluorescently labeled IgE (IgE-AF488 or IgE-AF647) into cells. Figures 20C to 20FThe results shown in demonstrate that 6H10-Oma and 6D1-Oma internalize pHrodo-labeled IgE (IgE-phrodo) into acidic compartments, while other M6PR-Oma bispecific antibodies do not. In particular, bispecific antibodies including 6D1-oma, 8D8-oma, 17F11-oma, and 6H10-oma showed uptake of IgE.

[0412] Evaluation of anti-M6PR / anti-C5 bispecific antibodies for C5 internalization and uptake

[0413] To evaluate whether anti-M6PR antibodies could internalize C5 as a target, a bispecific antibody with one anti-M6PR binding arm and one anti-C5 binding arm (eculizumab "ECU") was generated and assayed for uptake activity using C5 labeled with pHrodo green.

[0414] HeLa cells were plated in 96-well plates at 15,000 cells per well and incubated overnight. Anti-M6PR / anti-C5 bispecific antibodies were added to 100 μL complete growth medium (DMEM+10% FBS) at 100 nM. In a separate 96-well plate, C5 labeled with pHrodo green was added to 100 μL complete growth medium at 500 nM, and 2-fold serial dilutions were performed. The continuously diluted C5 was then combined with anti-M6PR / anti-C5 bispecific antibodies (53B11-ECU, 43C8-ECU, 12D5-ECU, 18G4-ECU) (final volume of 200 μL), added to HeLa cells, and incubated at 37 ° C for 18 hours. The internalization of C5 to lysosomes was measured by flow cytometry. Data was collected in the APC channel, and the mean fluorescence intensity was calculated.

[0415] Fig.26A (pH rodo) and Fig.26B The results shown in (AF647) demonstrated that anti-M6PR / anti-C5 bispecific antibodies (53B11-ECU, 43C8-ECU, 12D5-ECU, 18G4-ECU) transported fluorescently labeled C5 (C5-AF488 or C5-AF647) into cells.

[0416] 6.15.6. Example 6: Evaluation of internalization and degradation of anti-M6PR antibodies

[0417] Cell surface depletion and total depletion of HiBiT-tagged proteins of interest were measured using the Nano-Glo HiBiT Extracellular Detection System (Promega, Cat. No. N2422) and Nano-Glo HiBiT Lysis Detection System (Promega, Cat. No. N3050), respectively. Fig.21 Schematic diagrams of surface HiBit assays and total HiBit assays are provided, where the span indicates the extent of depletion of the target (eg, EGFR).

[0418] HiBiT assay protocol

[0419] Briefly, HCT116 cells expressing HiBiT-tagged EGFR were plated at 5,000 cells per well in 100 μL in a tissue culture treated white 96-well plate and grown for 24 hours at 37° C. Matuzumab-anti-M6PR bispecific antibody was diluted in culture medium to prepare a 3-fold dilution series from 40 to 0.002 nM (2x final concentration) and added to the cells at 100 μL per well, in addition to culture medium alone as a control.

[0420] The EGFR ligand EGF was used as a positive control and was assayed in a 3-fold dilution series from 100 to 0.005 nM (2 times final concentration). After antibody or control treatment, cells were grown at 37°C for 48 hours, and the culture medium was replaced with 50 μL of fresh culture medium and 50 μL of extracellular or lysed HiBiT detection reagent prepared according to the manufacturer's instructions. In brief, LgBiT protein and Nano-Glo HiBiT substrate were diluted in the corresponding Nano-Glo HiBiT buffer at a ratio of 1:100 and 1:50, respectively. After three minutes of shaking and incubation at ambient temperature for 10 minutes, the HiBiT signal was detected by measuring luminescence on an Envision plate reader (Perkin Elmer). The dose response was performed in duplicate and normalized to the culture medium control alone. IC was determined by fitting a 4-parameter curve in GraphPad Prism 50 value, and the span is calculated from the difference between the first and last points on the curve.

[0421] result

[0422] Fig.22A Surface depletion (ie, span) of EGFR for 16 Matuzumab-anti-M6PR bispecific antibodies is shown. Fig. 22B Total depletion (ie, span) of EGFR for 16 Matuzumab-anti-M6PR bispecific antibodies is shown. Fig. 22C The percentage of surface EGFR activity of exemplary matuzumab-anti-M6PR bispecific antibodies (3C7-matuz, 53B11-matuz) is shown. Fig.22DThe percentage of total EGFR activity of exemplary matuzumab-anti-M6PR bispecific antibodies (3C7-matuz, 53B11-matuz) is shown. In summary, the data show that anti-M6PR antibody clones 3C7, 6D1 and 17F11 are similar to EGF (positive control) in EGFR internalization and degradation with sub-nanomolar potency.

[0423] 6.15.7. Example 7: Evaluation of Recirculation of Anti-M6PR Antibodies

[0424] Recycling of the anti-M6PR-omalizumab (M6PR-Oma) bispecific antibody after internalization was assessed using a hIgE uptake assay (see, e.g., Fig.23 ). In particular, the M6PR-Oma bispecific antibody was diluted in RPMI+10% FBS to 2 times the final concentration. 100 μl of the M6PR-Oma bispecific antibody was added to 100 μl of RPMI+10% FBS and 1× penicillin / streptomycin containing 100,000 K562 cells or 100,000 HepG2 cells. The cells were contacted with the M6PR-Oma bispecific antibody to label the cells with the bispecific antibody. After labeling, the cells were washed to remove unbound (unlabeled) M6PR-Oma bispecific antibody. The cells were then divided into Fig.23 The four conditions indicated in : 0'; I-30', R-30' and R-60'.

[0425] For the O' condition, cells were incubated with hIgE-Alexa Fluor (AF) 647 ("hIgE-AF647") without first allowing the M6PR-Oma bispecific antibody to internalize. The O' control cells were then washed and assayed for the presence of the M6PR-Oma bispecific antibody at the cell surface using flow cytometry to measure AF647.

[0426] For conditions I-30', R-30' and R-60', cells were incubated at 37°C for 30 minutes to allow internalization of the M6PR-Oma bispecific antibody. After 30 minutes of incubation, the cells were exposed to hIgE-AF647. The cells in the I-30' condition were washed to remove unlabeled hIgE-AF647, and AF647 was measured using flow cytometry to determine the presence of the M6PR-Oma bispecific antibody at the cell surface. The cells in R-30' and R-60' were maintained at 37°C for the remainder of the specified incubation period to induce recycling. The cells in R-30' and R-60' were then washed to remove unlabeled hIgE-AF647, and AF647 was measured using flow cytometry to determine the presence of the M6PR-Oma bispecific antibody at the cell surface.

[0427] Fig.24A The internalization and recycling of the M6PR-Oma bispecific antibody by HepG2 cells, as measured by mean fluorescence intensity (MFI), is shown. Fig.23 Each antibody was evaluated for recycling using each of the indicated incubation schemes exemplified in . The ability of the bispecific antibodies to internalize and recycle to the surface was evaluated by comparing the MFI under I-30', R-30', and R-60' conditions with the MFI under 0' conditions ( Fig. 24B Based on this analysis, clones 12D5, 21D5, 18G4, and 53B11 showed that the recycling of the M6PR-Oma bispecific antibodies was most similar to the 0' baseline condition, indicating that these M6PR-Oma bispecific antibodies have characteristics that enable uptake and recycling in HepG2 cells.

[0428] Fig.25A Shown is the internalization and recycling of the MPR6-Oma bispecific antibody by K562 cells, as measured by mean fluorescence intensity (MFI). Fig.23 Each antibody was evaluated for recycling using each of the indicated incubation schemes exemplified in . The ability of the bispecific antibodies to internalize and recycle to the surface was evaluated by comparing the MFI under I-30', R-30', and R-60' conditions with the MFI under 0' conditions ( Fig.25B Based on this analysis, clones 12D5, 18G4, 21D5, 43C8, and 53B11 showed recycling of the M6PR-Oma bispecific antibody most similar to the 0' baseline condition, indicating that these Matuzumab-anti-M6PR bispecific antibodies promoted uptake and recycling in K562 cells.

[0429] 6.15.8. Example 8: Assessment of HER3 internalization and degradation

[0430] Cell surface depletion and total depletion of HiBiT-tagged HER3 (human epidermal growth receptor 3) were measured using the Nano-Glo HiBiT Extracellular Detection System (Promega, Cat. No. N2422) and the Nano-Glo HiBiT Lytic Detection System (Promega, Cat. No. N3050), respectively.

[0431] HiBiT assay protocol

[0432] In brief, HCC1419 cells (trastuzumab-resistant) expressing HiBiT-tagged HER3 were plated in 96-well plates treated with tissue culture at 100 μL and grown at 37°C for 24 hours. The LUM-anti-M6PR bispecific antibody was diluted in culture medium to prepare a 3-fold dilution series from 20 to 0.001 nM (2 times the final concentration) and added to the cells at 100 μL per well, with culture medium alone used as a control. Rutuzumab (LUM), humanized anti-human epidermal growth factor receptor 3 (HER3) monoclonal antibody, and NRG1 (Neuregulin 1) antibody were used as positive controls. A non-binding antibody (KLH) fused to LUM was used as a negative control for uptake activity.

[0433] After treatment with LUM-anti-M6PR bispecific antibody or control, cells were grown at 37°C for 48 hours and the medium was replaced with 50 μL of fresh medium and 50 μL of extracellular or lysed HiBiT detection reagent prepared according to the manufacturer's instructions. Briefly, LgBiT protein and Nano-Glo HiBiT substrate were diluted in the corresponding Nano-Glo HiBiT buffer at a ratio of 1:100 and 1:50, respectively. After three minutes of shaking and incubation at ambient temperature for 10 minutes, HiBiT signals were detected by measuring luminescence on an Envision plate reader (Perkin Elmer). Dose responses were performed in duplicate and normalized to medium-only controls. IC was determined by fitting a 4-parameter curve in GraphPad Prism 50 value, and the span is calculated from the difference between the first and last points on the curve.

[0434] LUM-anti-M6PR mediated HER3 internalization or degradation was assessed at 48 hours. Cell viability after LUM-anti-M6PR treatment was assessed at 72 hours. The results are shown in Figures 27A to 27B middle.

[0435] result

[0436] Fig.27AShown are surface depletion (ie, span) and total depletion (ie, span) of HER3 for a subset of LUM-anti-M6PR bispecific antibodies. Fig.27B Cell viability after treatment of a subset of LUM-anti-M6PR bispecific antibodies is shown. Fig.27A The left panel shows the percentage of surface HER3 activity of exemplary LUM-anti-M6PR bispecific antibodies (6D1, 3C7, 18G4). Fig.27A The right panel shows the percentage of total HER3 activity of exemplary LUM-anti-M6PR bispecific antibodies (6D1, 3C7, 18G4). Fig.27B Cell viability after treatment with exemplary LUM-anti-M6PR bispecific antibodies (6D1, 3C7, 18G4) is shown. Taken together, the data show that scfv LUM-anti-M6PR bispecific antibodies increase HER3 internalization compared to the parental antibody (LUM). In addition, the data demonstrate that scfv LUM-anti-M6PR bispecific antibodies effectively attenuate cell viability of trastuzumab-resistant cells.

[0437] 7. Incorporation by Reference

[0438] All references cited herein are incorporated by reference to the same extent as if each individual publication, database entry (e.g., Genbank sequence or GeneID entry), patent application, or patent was specifically and individually indicated as incorporated by reference in its entirety for all purposes. Applicants intend that this incorporation by reference statement be associated with each individual publication, database entry (e.g., Genbank sequence or GeneID entry), patent application, or patent, each of which is specifically identified in accordance with 37 CFR §1.57(b)(1), even if such reference is not directly adjacent to a specific incorporation by reference statement. The inclusion of a specific incorporation by reference statement (if any) within the specification does not in any way diminish this general incorporation by reference statement. Citation of a reference herein does not constitute an admission that the reference is relevant prior art, nor does it constitute any admission of the contents or date of these publications or documents.

[0439] 8. Terms

[0440] Clause 1. An antigen binding protein (ABP) comprising a first antigen binding moiety that specifically binds to an internalization domain of a cation-independent mannose 6-phosphate receptor (CI-M6PR) that facilitates intracellular transport of the ABP.

[0441] Item 2. The ABP according to Item 1, further comprising a target binding portion that specifically binds to a target molecule.

[0442] Clause 3. The ABP according to Clause 2, wherein the target binding moiety is a second antigen binding moiety that specifically binds to the target molecule.

[0443] Clause 4. The ABP according to Clause 2, wherein the target binding moiety is conjugated to the first antigen binding moiety, optionally via a linker.

[0444] Clause 5. The ABP according to any one of clauses 2 to 4, wherein the target molecule is a soluble extracellular target molecule or a cell surface target molecule.

[0445] Clause 6. The ABP of any preceding clause, further comprising an attached cargo portion.

[0446] Clause 7. The ABP according to Clause 6, wherein the cargo moiety is a polypeptide fused to the first antigen binding moiety or the second antigen binding moiety.

[0447] Clause 8. The ABP according to Clause 6, wherein the cargo moiety is conjugated to the first antigen binding moiety or the second antigen binding moiety, optionally via a linker.

[0448] Clause 9. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to a domain of CI-M6PR selected from the group consisting of domain 1, domain 4, domain 5, domain 6, domain 7 and domain 8.

[0449] Clause 10. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to domain 1 of CI-M6PR.

[0450] Clause 11. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to domain 4 of CI-M6PR.

[0451] Clause 12. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to domain 5 of CI-M6PR.

[0452] Clause 13. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to domain 6 of CI-M6PR.

[0453] Clause 14. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to domain 7 of CI-M6PR.

[0454] Clause 15. The ABP according to clause 1 or 2, wherein the first antigen binding portion binds to domain 8 of CI-M6PR.

[0455] Clause 16. The ABP according to any one of the preceding clauses, wherein the first antigen binding moiety binds to the CI-M6PR with high affinity.

[0456] Clause 17. The ABP according to Clause 16, wherein the first antigen binding portion has a dissociation equilibrium constant (K D ) combined with CI-M6PR.

[0457] Clause 18. The ABP according to any of the preceding clauses, wherein the first antigen binding portion has a K of between about 1 nM and about 500 nM. D Combined with CI-M6PR.

[0458] Clause 19. The ABP according to Clause 18, wherein the first antigen binding portion has a K of between about 10 nM and about 100 nM. D Combined with CI-M6PR.

[0459] Clause 20. The ABP according to Clause 18, wherein the first antigen binding portion has a K between about 100 nM and about 200 nM. D Combined with CI-M6PR.

[0460] Clause 21. The ABP according to Clause 18, wherein the first antigen binding portion has a K between about 200 nM and about 300 nM. D Combined with CI-M6PR.

[0461] Clause 22. The ABP according to Clause 18, wherein the first antigen binding portion has a K between about 300 nM and about 400 nM. D Combined with CI-M6PR.

[0462] Clause 23. The ABP according to Clause 18, wherein the first antigen binding portion has a K between about 400 nM and about 500 nM. D Combined with CI-M6PR.

[0463] Clause 24. The ABP according to any of the preceding clauses, wherein the dissociation rate (K) of the first antigen binding portion against CI-M6PR is off ) in 1x 10 -8 s -1 With 0.1s -1 between.

[0464] Clause 25. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is directed against the K of CI-M6PR. off In 1x 10 -6 s-1 With 1x 10 -2 s -1 between.

[0465] Clause 26. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is directed against the K of CI-M6PR. off About 1x 10 -5 s -1 Or slower.

[0466] Clause 27. The ABP according to any one of the preceding clauses, wherein said binding of said first antigen binding moiety to CI-M6PR is pH dependent.

[0467] Clause 28. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is released from CI-M6PR at a pH of 7.4 or lower.

[0468] Clause 29. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is released from CI-M6PR at a pH of 6.5 or lower.

[0469] Clause 30. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is released from CI-M6PR at a pH of 6.0 or lower.

[0470] Clause 31. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is released from CI-M6PR at a pH of 5.5 or lower.

[0471] Clause 32. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is released from CI-M6PR at a pH of 5.0 or less.

[0472] Clause 33. The ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a light chain CDR3 (LCDR3) having the sequence of SEQ ID NO:76 and a heavy chain CDR3 (HCDR3) having the sequence of SEQ ID NO:35.

[0473] Clause 34. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 78 and a heavy chain CDR3 having the amino acid sequence of SEQ ID NO: 38.

[0474] Clause 35. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 81 and a heavy chain CDR3 having the sequence of SEQ ID NO: 41.

[0475] Clause 36. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 84 and a heavy chain CDR3 having the sequence of SEQ ID NO: 44.

[0476] Clause 37. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 85 and a heavy chain CDR3 having the sequence of SEQ ID NO: 46.

[0477] Clause 38. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 88 and a heavy chain CDR3 having the sequence of SEQ ID NO: 49.

[0478] Clause 39. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:91 and a heavy chain CDR3 having the sequence of SEQ ID NO:52.

[0479] Clause 40. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 94 and a heavy chain CDR3 having the sequence of SEQ ID NO: 54.

[0480] Clause 41. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:96 and a heavy chain CDR3 having the sequence of SEQ ID NO:55.

[0481] Clause 42. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:99 and a heavy chain CDR3 having the sequence of SEQ ID NO:58.

[0482] Clause 43. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 101 and a heavy chain CDR3 having the sequence of SEQ ID NO: 60.

[0483] Clause 44. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 104 and a heavy chain CDR3 having the sequence of SEQ ID NO: 63.

[0484] Clause 45. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 107 and a heavy chain CDR3 having the sequence of SEQ ID NO: 66.

[0485] Clause 46. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 110 and a heavy chain CDR3 having the sequence of SEQ ID NO:69.

[0486] Clause 47. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 113 and a heavy chain CDR3 having the sequence of SEQ ID NO:72.

[0487] Clause 48. The ABP according to any one of clauses 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 116 and a heavy chain CDR3 having the sequence of SEQ ID NO:73.

[0488] Clause 49. The ABP of Clause 33, wherein the first antigen binding portion further comprises a light chain CDR1 (LCDR1) having the sequence of SEQ ID NO:74 and a heavy chain CDR1 (HCDR1) having the sequence of SEQ ID NO:33; and a light chain CDR2 (LCDR2) having the sequence of SEQ ID NO:75 and a heavy chain CDR2 (HCDR2) having the sequence of SEQ ID NO:34.

[0489] Clause 50. The ABP of Clause 34, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 77 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 36; and a LCDR2 having the sequence of SEQ ID NO: 75 and a HCDR2 having the sequence of SEQ ID NO: 37.

[0490] Clause 51. The ABP of Clause 35, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 39; and a LCDR2 having the sequence of SEQ ID NO: 80 and a HCDR2 having the sequence of SEQ ID NO: 40.

[0491] Clause 52. The ABP of Clause 36, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 82 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 42; and a LCDR2 having the sequence of SEQ ID NO: 83 and a HCDR2 having the sequence of SEQ ID NO: 43.

[0492] Clause 53. The ABP of Clause 37, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 45; and a LCDR2 having the sequence of SEQ ID NO: 80 and a HCDR2 having the sequence of SEQ ID NO: 34.

[0493] Clause 54. The ABP of Clause 38, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:86 and a heavy chain HCDR1 having the sequence of SEQ ID NO:47; and a LCDR2 having the sequence of SEQ ID NO:87 and a HCDR2 having the sequence of SEQ ID NO:48.

[0494] Clause 55. The ABP of Clause 39, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:89 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:90 and a HCDR2 having the sequence of SEQ ID NO:53.

[0495] Clause 56. The ABP of Clause 40, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:92 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:93 and a HCDR2 having the sequence of SEQ ID NO:53.

[0496] Clause 57. The ABP of Clause 41, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:95 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:93 and a HCDR2 having the sequence of SEQ ID NO:51.

[0497] Clause 58. The ABP of Clause 42, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:97 and a heavy chain HCDR1 having the sequence of SEQ ID NO:56; and a LCDR2 having the sequence of SEQ ID NO:98 and a HCDR2 having the sequence of SEQ ID NO:57.

[0498] Clause 59. The ABP of Clause 43, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 100 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 50; and a LCDR2 having the sequence of SEQ ID NO: 93 and a HCDR2 having the sequence of SEQ ID NO: 59.

[0499] Clause 60. The ABP of Clause 44, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 102 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 61; and a LCDR2 having the sequence of SEQ ID NO: 103 and a HCDR2 having the sequence of SEQ ID NO: 62.

[0500] Clause 61. The ABP of Clause 45, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 105 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 64; and a LCDR2 having the sequence of SEQ ID NO: 106 and a HCDR2 having the sequence of SEQ ID NO: 65.

[0501] Clause 62. The ABP of Clause 46, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 67; and a LCDR2 having the sequence of SEQ ID NO: 109 and a HCDR2 having the sequence of SEQ ID NO: 68.

[0502] Clause 63. The ABP of Clause 47, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 70; and a LCDR2 having the sequence of SEQ ID NO: 112 and a HCDR2 having the sequence of SEQ ID NO: 71.

[0503] Clause 64. The ABP of Clause 48, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 70; and a LCDR2 having the sequence of SEQ ID NO: 115 and a HCDR2 having the sequence of SEQ ID NO: 71.

[0504] Clause 65. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:205, LFR2 is at least 95% identical to the sequence of SEQ ID NO:206, LFR3 is at least 95% identical to the sequence of SEQ ID NO:207; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:208.

[0505] Clause 66. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:209, LFR2 is at least 95% identical to the sequence of SEQ ID NO:210, LFR3 is at least 95% identical to the sequence of SEQ ID NO:211; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:212.

[0506] Clause 67. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:213, LFR2 is at least 95% identical to the sequence of SEQ ID NO:214, LFR3 is at least 95% identical to the sequence of SEQ ID NO:215; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:216.

[0507] Clause 68. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:217, LFR2 is at least 95% identical to the sequence of SEQ ID NO:218, LFR3 is at least 95% identical to the sequence of SEQ ID NO:219; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:220.

[0508] Clause 69. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:221, LFR2 is at least 95% identical to the sequence of SEQ ID NO:222, LFR3 is at least 95% identical to the sequence of SEQ ID NO:223; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:224.

[0509] Clause 70. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:225, LFR2 is at least 95% identical to the sequence of SEQ ID NO:226, LFR3 is at least 95% identical to the sequence of SEQ ID NO:227; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:228.

[0510] Clause 71. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:229, LFR2 is at least 95% identical to the sequence of SEQ ID NO:230, LFR3 is at least 95% identical to the sequence of SEQ ID NO:231; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:232.

[0511] Clause 72. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:233, LFR2 is at least 95% identical to the sequence of SEQ ID NO:234, LFR3 is at least 95% identical to the sequence of SEQ ID NO:235; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:236.

[0512] Clause 73. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:237, LFR2 is at least 95% identical to the sequence of SEQ ID NO:238, LFR3 is at least 95% identical to the sequence of SEQ ID NO:239; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:240.

[0513] Clause 74. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:241, LFR2 is at least 95% identical to the sequence of SEQ ID NO:242, LFR3 is at least 95% identical to the sequence of SEQ ID NO:243; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:244.

[0514] Clause 75. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:245, LFR2 is at least 95% identical to the sequence of SEQ ID NO:246, LFR3 is at least 95% identical to the sequence of SEQ ID NO:247; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:248.

[0515] Clause 76. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:249, LFR2 is at least 95% identical to the sequence of SEQ ID NO:250, LFR3 is at least 95% identical to the sequence of SEQ ID NO:251; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:252.

[0516] Clause 77. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:253, LFR2 is at least 95% identical to the sequence of SEQ ID NO:254, LFR3 is at least 95% identical to the sequence of SEQ ID NO:255; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:256.

[0517] Clause 78. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:257, LFR2 is at least 95% identical to the sequence of SEQ ID NO:258, LFR3 is at least 95% identical to the sequence of SEQ ID NO:259; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:260.

[0518] Clause 79. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:261, LFR2 is at least 95% identical to the sequence of SEQ ID NO:262, LFR3 is at least 95% identical to the sequence of SEQ ID NO:263; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:264.

[0519] Clause 80. An ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:265, LFR2 is at least 95% identical to the sequence of SEQ ID NO:266, LFR3 is at least 95% identical to the sequence of SEQ ID NO:267; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:268.

[0520] Clause 81. The ABP of clause 65, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 141, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 142, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 143; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 144.

[0521] Clause 82. The ABP of clause 66, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 145, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 146, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 147; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 148; or

[0522] Clause 83. An ABP according to clause 67, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 149, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 150, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 151; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 152.

[0523] Clause 84. The ABP of clause 68, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 153, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 154, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 155; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 156.

[0524] Clause 85. The ABP of clause 69, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 157, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 158, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 159; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 160.

[0525] Clause 86. The ABP of clause 70, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 161, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 162, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 163; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 164.

[0526] Clause 87. The ABP of clause 71, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 165, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 166, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 167; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 168.

[0527] Clause 88. The ABP of clause 72, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 169, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 170, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 171; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 172.

[0528] Clause 89. The ABP of clause 73, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 173, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 174, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 175; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 176.

[0529] Clause 90. The ABP of clause 74, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 177, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 178, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 179; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 180.

[0530] Clause 91. The ABP of clause 75, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 181, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 182, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 183; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 184.

[0531] Clause 92. The ABP of clause 76, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 185, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 186, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 187; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 188.

[0532] Clause 93. The ABP of clause 77, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 189, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 190, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 191; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 192; or

[0533] Clause 94. The ABP of clause 78, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 193, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 194, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 195; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 196; or

[0534] Clause 95. The ABP of clause 79, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 197, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 198, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 199; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 200; or

[0535] Clause 96. The ABP of clause 80, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:201, HFR2 is at least 95% identical to the sequence of SEQ ID NO:202, HFR3 is at least 95% identical to the sequence of SEQ ID NO:203; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:204.

[0536] Clause 97. The ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable light chain (VL) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

[0537] Clause 98. The ABP according to any of the preceding clauses, wherein the first antigen binding portion comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

[0538] Clause 99. The ABP according to any of the preceding clauses, wherein the first antigen binding portion is an antibody fragment (eg, a single chain variable fragment (scFv) or an antigen binding fragment).

[0539] Clause 100. The ABP according to any of the preceding clauses, wherein the ABP is transported to the lysosome.

[0540] Clause 101. The ABP according to any of the preceding clauses, wherein the ABP is transported back to the cell surface after internalization.

[0541] Clause 102. The ABP of any one of clauses 1 to 100, wherein the ABP is degraded in the cell.

[0542] Clause 103. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein

[0543] The LCDR1 has a sequence of SEQ ID NO: 82,

[0544] The LCDR2 has a sequence of SEQ ID NO: 83,

[0545] The LCDR3 has a sequence of SEQ ID NO: 84,

[0546] The HCDR1 has the sequence of SEQ ID NO:42,

[0547] The HCDR2 has the sequence of SEQ ID NO: 43, and

[0548] The HCDR3 has the sequence of SEQ ID NO:44.

[0549] Clause 104. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein

[0550] The LCDR1 has a sequence of SEQ ID NO: 92,

[0551] The LCDR2 has a sequence of SEQ ID NO: 93,

[0552] The LCDR3 has a sequence of SEQ ID NO: 94,

[0553] The HCDR1 has the sequence of SEQ ID NO: 50,

[0554] The HCDR2 has the sequence of SEQ ID NO: 53, and

[0555] The HCDR3 has the sequence of SEQ ID NO:54.

[0556] Clause 105. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein

[0557] The LCDR1 has a sequence of SEQ ID NO: 97,

[0558] The LCDR2 has a sequence of SEQ ID NO: 98,

[0559] The LCDR3 has a sequence of SEQ ID NO: 99,

[0560] The HCDR1 has the sequence of SEQ ID NO:56,

[0561] The HCDR2 has the sequence of SEQ ID NO: 57, and

[0562] The HCDR3 has the sequence of SEQ ID NO:58.

[0563] Clause 106. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein

[0564] The LCDR1 has a sequence of SEQ ID NO: 100,

[0565] The LCDR2 has a sequence of SEQ ID NO: 93,

[0566] The LCDR3 has a sequence of SEQ ID NO: 101,

[0567] The HCDR1 has the sequence of SEQ ID NO: 50,

[0568] The HCDR2 has the sequence of SEQ ID NO: 59, and

[0569] The HCDR3 has the sequence of SEQ ID NO:60.

[0570] Clause 107. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein

[0571] The LCDR1 has a sequence of SEQ ID NO: 108,

[0572] The LCDR2 has a sequence of SEQ ID NO: 109,

[0573] The LCDR3 has a sequence of SEQ ID NO: 110,

[0574] The HCDR1 has the sequence of SEQ ID NO: 67,

[0575] The HCDR2 has the sequence of SEQ ID NO: 68, and

[0576] The HCDR3 has the sequence of SEQ ID NO:69.

[0577] Clause 108. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein

[0578] The LCDR1 has a sequence of SEQ ID NO: 111,

[0579] The LCDR2 has a sequence of SEQ ID NO: 112,

[0580] The LCDR3 has a sequence of SEQ ID NO: 113,

[0581] The HCDR1 has the sequence of SEQ ID NO: 70,

[0582] The HCDR2 has the sequence of SEQ ID NO: 71, and

[0583] The HCDR3 has the sequence of SEQ ID NO:72.

[0584] Clause 109. A bifunctional molecule comprising:

[0585] a first moiety that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), wherein the first moiety is an antibody or an antibody fragment; and

[0586] A second portion that specifically binds to a cell surface target molecule or an extracellular target molecule, wherein the second portion is selected from an antibody, an antigen binding fragment, a ligand, and a small molecule.

[0587] Clause 110. The bifunctional molecule of Clause 109, wherein the bifunctional molecule is a polypeptide.

[0588] Clause 111. The bifunctional molecule according to Clause 109, wherein the first portion and the second portion are covalently attached via a linker.

[0589] Clause 112. The bifunctional molecule according to clause 111, wherein the second moiety is a small molecule.

[0590] Clause 113. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 (LCDR3) having the sequence of SEQ ID NO:76 and a heavy chain CDR3 (HCDR3) having the sequence of SEQ ID NO:35.

[0591] Clause 114. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:78 and a heavy chain CDR3 having the amino acid sequence of SEQ ID NO:38.

[0592] Clause 115. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:81 and a heavy chain CDR3 having the sequence of SEQ ID NO:41.

[0593] Clause 116. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:84 and a heavy chain CDR3 having the sequence of SEQ ID NO:44.

[0594] Clause 117. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:85 and a heavy chain CDR3 having the sequence of SEQ ID NO:46.

[0595] Clause 118. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:88 and a heavy chain CDR3 having the sequence of SEQ ID NO:49.

[0596] Clause 119. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:91 and a heavy chain CDR3 having the sequence of SEQ ID NO:52.

[0597] Clause 120. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:94 and a heavy chain CDR3 having the sequence of SEQ ID NO:54.

[0598] Clause 121. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:96 and a heavy chain CDR3 having the sequence of SEQ ID NO:55.

[0599] Clause 122. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:99 and a heavy chain CDR3 having the sequence of SEQ ID NO:58.

[0600] Clause 123. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 101 and a heavy chain CDR3 having the sequence of SEQ ID NO: 60.

[0601] Clause 124. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 104 and a heavy chain CDR3 having the sequence of SEQ ID NO: 63.

[0602] Clause 125. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 107 and a heavy chain CDR3 having the sequence of SEQ ID NO: 66.

[0603] Clause 126. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 110 and a heavy chain CDR3 having the sequence of SEQ ID NO:69.

[0604] Clause 127. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 113 and a heavy chain CDR3 having the sequence of SEQ ID NO:72.

[0605] Clause 128. The bifunctional molecule according to any one of clauses 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 116 and a heavy chain CDR3 having the sequence of SEQ ID NO:73.

[0606] Clause 129. The bifunctional molecule of Clause 113, wherein the first portion further comprises a light chain CDR1 (LCDR1) having the sequence of SEQ ID NO: 74 and a heavy chain CDR1 (HCDR1) having the sequence of SEQ ID NO: 33; and a light chain CDR2 (LCDR2) having the sequence of SEQ ID NO: 75 and a heavy chain CDR2 (HCDR2) having the sequence of SEQ ID NO: 34.

[0607] Clause 130. The bifunctional molecule of clause 114, wherein the first portion further comprises a LCDR1 having the sequence of SEQ ID NO: 77 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 36; and a LCDR2 having the sequence of SEQ ID NO: 75 and a HCDR2 having the sequence of SEQ ID NO: 37.

[0608] Clause 131. The bifunctional molecule of clause 115, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 39; and a LCDR2 having a sequence of SEQ ID NO: 80 and a HCDR2 having a sequence of SEQ ID NO: 40.

[0609] Clause 132. The bifunctional molecule of clause 116, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 82 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 42; and a LCDR2 having a sequence of SEQ ID NO: 83 and a HCDR2 having a sequence of SEQ ID NO: 43.

[0610] Clause 133. The bifunctional molecule of clause 117, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 45; and a LCDR2 having a sequence of SEQ ID NO: 80 and a HCDR2 having a sequence of SEQ ID NO: 34.

[0611] Clause 134. The bifunctional molecule of Clause 118, wherein the first portion further comprises a LCDR1 having the sequence of SEQ ID NO:86 and a heavy chain HCDR1 having the sequence of SEQ ID NO:47; and a LCDR2 having the sequence of SEQ ID NO:87 and a HCDR2 having the sequence of SEQ ID NO:48.

[0612] Clause 135. The bifunctional molecule of Clause 119, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:89 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:90 and a HCDR2 having a sequence of SEQ ID NO:53.

[0613] Clause 136. The bifunctional molecule of clause 120, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:92 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:53.

[0614] Clause 137. The bifunctional molecule of clause 121, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:95 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:51.

[0615] Clause 138. The bifunctional molecule of clause 122, wherein the first portion further comprises a LCDR1 having the sequence of SEQ ID NO:97 and a heavy chain HCDR1 having the sequence of SEQ ID NO:56; and a LCDR2 having the sequence of SEQ ID NO:98 and a HCDR2 having the sequence of SEQ ID NO:57.

[0616] Clause 139. The bifunctional molecule of clause 123, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 100 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 50; and a LCDR2 having a sequence of SEQ ID NO: 93 and a HCDR2 having a sequence of SEQ ID NO: 59.

[0617] Clause 140. The bifunctional molecule of clause 124, wherein the first portion further comprises a LCDR1 having the sequence of SEQ ID NO: 102 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 61; and a LCDR2 having the sequence of SEQ ID NO: 103 and a HCDR2 having the sequence of SEQ ID NO: 62.

[0618] Clause 141. The bifunctional molecule of clause 125, wherein the first portion further comprises a LCDR1 having the sequence of SEQ ID NO: 105 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 64; and a LCDR2 having the sequence of SEQ ID NO: 106 and a HCDR2 having the sequence of SEQ ID NO: 65.

[0619] Clause 142. The bifunctional molecule of clause 126, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 67; and a LCDR2 having a sequence of SEQ ID NO: 109 and a HCDR2 having a sequence of SEQ ID NO: 68.

[0620] Clause 143. The bifunctional molecule of clause 127, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 112 and a HCDR2 having a sequence of SEQ ID NO: 71.

[0621] Clause 144. The bifunctional molecule of clause 128, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 115 and a HCDR2 having a sequence of SEQ ID NO: 71.

[0622] Clause 145. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding moiety comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 205, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 206, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 207; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 208; or

[0623] Clause 146. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 209, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 210, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 211; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 212.

[0624] Clause 147. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 213, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 214, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 215; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 216.

[0625] Clause 148. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 217, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 218, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 219; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 220.

[0626] Clause 149. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 221, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 222, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 223; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 224.

[0627] Clause 150. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 225, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 226, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 227; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 228.

[0628] Clause 151. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 229, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 230, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 231; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 232.

[0629] Clause 152. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 233, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 234, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 235; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 236.

[0630] Clause 153. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 237, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 238, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 239; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 240.

[0631] Clause 154. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 241, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 242, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 243; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 244.

[0632] Clause 155. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 245, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 246, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 247; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 248.

[0633] Clause 156. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 249, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 250, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 251; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 252.

[0634] Clause 157. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 253, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 254, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 255; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 256.

[0635] Clause 158. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 257, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 258, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 259; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 260.

[0636] Clause 159. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 261, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 262, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 263; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 264.

[0637] Clause 160. A bifunctional molecule according to any one of clauses 109 to 144, wherein the first binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 265, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 266, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 267; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 268.

[0638] Item 161. A bifunctional molecule according to item 145, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 141, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 142, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 143; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 144.

[0639] Clause 162. A bifunctional molecule according to clause 146, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 145, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 146, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 147; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 148.

[0640] Clause 163. A bifunctional molecule according to clause 147, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 149, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 150, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 151; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 152.

[0641] Clause 164. A bifunctional molecule according to clause 148, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 153, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 154, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 155; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 156.

[0642] Clause 165. A bifunctional molecule according to clause 149, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 157, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 158, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 159; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 160.

[0643] Clause 166. A bifunctional molecule according to clause 150, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 161, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 162, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 163; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 164.

[0644] Item 167. A bifunctional molecule according to item 151, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:165, HFR2 is at least 95% identical to the sequence of SEQ ID NO:166, HFR3 is at least 95% identical to the sequence of SEQ ID NO:167; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:168.

[0645] Item 168. A bifunctional molecule according to item 152, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 169, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 170, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 171; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 172.

[0646] Clause 169. A bifunctional molecule according to clause 153, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 173, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 174, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 175; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 176.

[0647] Clause 170. A bifunctional molecule according to clause 154, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 177, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 178, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 179; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 180.

[0648] Clause 171. A bifunctional molecule according to clause 155, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 181, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 182, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 183; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 184.

[0649] Clause 172. A bifunctional molecule according to clause 156, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 185, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 186, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 187; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 188.

[0650] Clause 173. A bifunctional molecule according to clause 157, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 189, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 190, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 191; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 192.

[0651] Clause 174. A bifunctional molecule according to clause 158, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 193, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 194, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 195; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 196.

[0652] Clause 175. A bifunctional molecule according to clause 159, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 197, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 198, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 199; and HFR4 is at least 95% identical to the sequence of SEQ ID NO: 200.

[0653] Clause 176. A bifunctional molecule according to clause 160, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:201, HFR2 is at least 95% identical to the sequence of SEQ ID NO:202, HFR3 is at least 95% identical to the sequence of SEQ ID NO:203; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:204.

[0654] Clause 177. The bifunctional molecule according to any one of clauses 109 to 176, wherein the first portion comprises a variable light chain (VL) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

[0655] Clause 178. The bifunctional molecule according to any one of clauses 109 to 177, wherein the first portion comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

[0656] Clause 179. A method for degrading a soluble target molecule or a cell surface target molecule, the method comprising:

[0657] (a) contacting the target molecule with a multispecific antigen binding protein (ABP), wherein the ABP comprises a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR) on the surface of a cell; and

[0658] (b) transporting the ABP, the target molecule and the CI-M6PR to a lysosome in a cell, wherein the target molecule is degraded in the lysosome.

[0659] Clause 180. The method of Clause 179, wherein the ABP further comprises a cargo moiety attached.

[0660] Clause 181. The method according to clause 180, wherein the cargo moiety is a polypeptide fused to the ABP.

[0661] Clause 182. The method according to clause 181, wherein the cargo moiety is conjugated to the ABP, optionally via a linker.

[0662] Clause 183. A method according to any one of clauses 179 to 182, wherein the first antigen binding moiety binds to a domain of CI-M6PR selected from the group consisting of domain 1, domain 4, domain 5, domain 6, domain 7 and domain 8.

[0663] Clause 184. A method according to any one of clauses 179 to 182, wherein the first antigen binding moiety has a dissociation equilibrium constant (K D ) combined with CI-M6PR.

[0664] Clause 185. A method according to any one of clauses 179 to 182, wherein the first antigen binding moiety has a dissociation equilibrium constant (K) between about 1 nM and about 500 nM. D ) combined with CI-M6PR.

[0665] Clause 186. A method according to any one of clauses 179 to 185, wherein the dissociation rate (k off ) in 1x 10 -8 s-1 With 0.1s -1 between.

[0666] Clause 187. The method according to any one of clauses 179 to 186, wherein the binding of the first antigen binding moiety to CI-M6PR is pH dependent.

[0667] Clause 188. The method according to any one of clauses 179 to 187, wherein the first antigen binding moiety is released from CI-M6PR at a pH of 7.4 or lower.

[0668] Clause 189. The method according to any one of clauses 179 to 188, wherein the first antigen binding moiety is released from CI-M6PR at a pH of 6.0 or lower.

[0669] Clause 190. The method according to any one of clauses 179 to 189, wherein the first antigen binding moiety is released from CI-M6PR at a pH of 5.5 or lower.

[0670] Clause 191. The method according to any one of clauses 179 to 190, wherein the first antigen binding moiety is released from CI-M6PR at a pH of 5.0 or lower.

[0671] Clause 192. A method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 (LCDR3) having the sequence of SEQ ID NO:76 and a heavy chain CDR3 (HCDR3) having the sequence of SEQ ID NO:35.

[0672] Clause 193. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:78 and a heavy chain CDR3 having the amino acid sequence of SEQ ID NO:38.

[0673] Clause 194. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:81 and a heavy chain CDR3 having the sequence of SEQ ID NO:41.

[0674] Clause 195. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:84 and a heavy chain CDR3 having the sequence of SEQ ID NO:44.

[0675] Clause 196. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:85 and a heavy chain CDR3 having the sequence of SEQ ID NO:46.

[0676] Clause 197. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:88 and a heavy chain CDR3 having the sequence of SEQ ID NO:49.

[0677] Clause 198. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:91 and a heavy chain CDR3 having the sequence of SEQ ID NO:52.

[0678] Clause 199. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:94 and a heavy chain CDR3 having the sequence of SEQ ID NO:54.

[0679] Clause 200. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:96 and a heavy chain CDR3 having the sequence of SEQ ID NO:55.

[0680] Clause 201. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:99 and a heavy chain CDR3 having the sequence of SEQ ID NO:58.

[0681] Clause 202. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 101 and a heavy chain CDR3 having the sequence of SEQ ID NO:60.

[0682] Clause 203. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 104 and a heavy chain CDR3 having the sequence of SEQ ID NO:63.

[0683] Clause 204. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 107 and a heavy chain CDR3 having the sequence of SEQ ID NO: 66.

[0684] Clause 205. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 110 and a heavy chain CDR3 having the sequence of SEQ ID NO:69.

[0685] Clause 206. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 113 and a heavy chain CDR3 having the sequence of SEQ ID NO:72.

[0686] Clause 207. The method according to any one of clauses 179 to 191, wherein the first antigen binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 116 and a heavy chain CDR3 having the sequence of SEQ ID NO:73.

[0687] Clause 208. A method according to clause 192, wherein the first antigen binding portion further comprises a light chain CDR1 (LCDR1) having the sequence of SEQ ID NO:74 and a heavy chain CDR1 (HCDR1) having the sequence of SEQ ID NO:33; and a light chain CDR2 (LCDR2) having the sequence of SEQ ID NO:75 and a heavy chain CDR2 (HCDR2) having the sequence of SEQ ID NO:34.

[0688] Clause 209. A method according to clause 193, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:77 and a heavy chain HCDR1 having a sequence of SEQ ID NO:36; and a LCDR2 having a sequence of SEQ ID NO:75 and a HCDR2 having a sequence of SEQ ID NO:37.

[0689] Clause 210. A method according to clause 194, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:79 and a heavy chain HCDR1 having a sequence of SEQ ID NO:39; and a LCDR2 having a sequence of SEQ ID NO:80 and a HCDR2 having a sequence of SEQ ID NO:40.

[0690] Clause 211. A method according to clause 195, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:82 and a heavy chain HCDR1 having a sequence of SEQ ID NO:42; and a LCDR2 having a sequence of SEQ ID NO:83 and a HCDR2 having a sequence of SEQ ID NO:43.

[0691] Clause 212. A method according to clause 196, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:79 and a heavy chain HCDR1 having a sequence of SEQ ID NO:45; and a LCDR2 having a sequence of SEQ ID NO:80 and a HCDR2 having a sequence of SEQ ID NO:34.

[0692] Clause 213. A method according to clause 197, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:86 and a heavy chain HCDR1 having a sequence of SEQ ID NO:47; and a LCDR2 having a sequence of SEQ ID NO:87 and a HCDR2 having a sequence of SEQ ID NO:48.

[0693] Clause 214. A method according to clause 198, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:89 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:90 and a HCDR2 having a sequence of SEQ ID NO:53.

[0694] Clause 215. A method according to clause 199, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:92 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:53.

[0695] Clause 216. A method according to clause 200, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:95 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:51.

[0696] Clause 217. A method according to clause 201, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:97 and a heavy chain HCDR1 having a sequence of SEQ ID NO:56; and a LCDR2 having a sequence of SEQ ID NO:98 and a HCDR2 having a sequence of SEQ ID NO:57.

[0697] Clause 218. A method according to clause 202, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 100 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 50; and a LCDR2 having a sequence of SEQ ID NO: 93 and a HCDR2 having a sequence of SEQ ID NO: 59.

[0698] Clause 219. A method according to clause 203, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 102 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 61; and a LCDR2 having a sequence of SEQ ID NO: 103 and a HCDR2 having a sequence of SEQ ID NO: 62.

[0699] Clause 220. A method according to clause 204, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 105 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 64; and a LCDR2 having a sequence of SEQ ID NO: 106 and a HCDR2 having a sequence of SEQ ID NO: 65.

[0700] Clause 221. A method according to clause 205, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 67; and a LCDR2 having a sequence of SEQ ID NO: 109 and a HCDR2 having a sequence of SEQ ID NO: 68.

[0701] Clause 222. A method according to clause 206, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 112 and a HCDR2 having a sequence of SEQ ID NO: 71.

[0702] Clause 223. A method according to clause 207, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 115 and a HCDR2 having a sequence of SEQ ID NO: 71.

[0703] Clause 224. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:205, LFR2 is at least 95% identical to the sequence of SEQ ID NO:206, LFR3 is at least 95% identical to the sequence of SEQ ID NO:207; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:208.

[0704] Clause 225. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 209, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 210, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 211; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 212.

[0705] Clause 226. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:213, LFR2 is at least 95% identical to the sequence of SEQ ID NO:214, LFR3 is at least 95% identical to the sequence of SEQ ID NO:215; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:216.

[0706] Clause 227. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 217, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 218, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 219; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 220.

[0707] Clause 228. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 221, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 222, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 223; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 224.

[0708] Clause 229. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 225, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 226, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 227; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 228.

[0709] Clause 230. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 229, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 230, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 231; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 232.

[0710] Clause 231. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 233, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 234, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 235; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 236.

[0711] Clause 232. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 237, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 238, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 239; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 240.

[0712] Clause 233. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 241, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 242, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 243; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 244.

[0713] Clause 234. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 245, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 246, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 247; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 248.

[0714] Clause 235. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 249, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 250, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 251; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 252.

[0715] Clause 236. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 253, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 254, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 255; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 256.

[0716] Clause 237. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 257, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 258, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 259; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 260.

[0717] Clause 238. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 261, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 262, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 263; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 264.

[0718] Clause 239. A method according to any one of clauses 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 265, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 266, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 267; and LFR4 is at least 95% identical to the sequence of SEQ ID NO: 268.

[0719] Item 240. A method according to item 224, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:141, HFR2 is at least 95% identical to the sequence of SEQ ID NO:142, HFR3 is at least 95% identical to the sequence of SEQ ID NO:143; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:144.

[0720] Item 241. A method according to item 225, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:145, HFR2 is at least 95% identical to the sequence of SEQ ID NO:146, HFR3 is at least 95% identical to the sequence of SEQ ID NO:147; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:148.

[0721] Item 242. A method according to item 226, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:149, HFR2 is at least 95% identical to the sequence of SEQ ID NO:150, HFR3 is at least 95% identical to the sequence of SEQ ID NO:151; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:152.

[0722] Item 243. A method according to item 227, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:153, HFR2 is at least 95% identical to the sequence of SEQ ID NO:154, HFR3 is at least 95% identical to the sequence of SEQ ID NO:155; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:156.

[0723] Clause 244. A method according to clause 228, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:157, HFR2 is at least 95% identical to the sequence of SEQ ID NO:158, HFR3 is at least 95% identical to the sequence of SEQ ID NO:159; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:160.

[0724] Item 245. A method according to item 229, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:161, HFR2 is at least 95% identical to the sequence of SEQ ID NO:162, HFR3 is at least 95% identical to the sequence of SEQ ID NO:163; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:164.

[0725] Item 246. A method according to item 230, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:165, HFR2 is at least 95% identical to the sequence of SEQ ID NO:166, HFR3 is at least 95% identical to the sequence of SEQ ID NO:167; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:168.

[0726] Clause 247. A method according to clause 231, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:169, HFR2 is at least 95% identical to the sequence of SEQ ID NO:170, HFR3 is at least 95% identical to the sequence of SEQ ID NO:171; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:172.

[0727] Item 248. A method according to item 232, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:173, HFR2 is at least 95% identical to the sequence of SEQ ID NO:174, HFR3 is at least 95% identical to the sequence of SEQ ID NO:175; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:176.

[0728] Item 249. A method according to item 233, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:177, HFR2 is at least 95% identical to the sequence of SEQ ID NO:178, HFR3 is at least 95% identical to the sequence of SEQ ID NO:179; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:180.

[0729] Item 250. A method according to item 234, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:181, HFR2 is at least 95% identical to the sequence of SEQ ID NO:182, HFR3 is at least 95% identical to the sequence of SEQ ID NO:183; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:184.

[0730] Item 251. A method according to item 235, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:185, HFR2 is at least 95% identical to the sequence of SEQ ID NO:186, HFR3 is at least 95% identical to the sequence of SEQ ID NO:187; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:188.

[0731] Item 252. A method according to item 236, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:189, HFR2 is at least 95% identical to the sequence of SEQ ID NO:190, HFR3 is at least 95% identical to the sequence of SEQ ID NO:191; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:192.

[0732] Item 253. A method according to item 237, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:193, HFR2 is at least 95% identical to the sequence of SEQ ID NO:194, HFR3 is at least 95% identical to the sequence of SEQ ID NO:195; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:196.

[0733] Item 254. A method according to item 238, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:197, HFR2 is at least 95% identical to the sequence of SEQ ID NO:198, HFR3 is at least 95% identical to the sequence of SEQ ID NO:199; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:200.

[0734] Item 255. A method according to item 239, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:201, HFR2 is at least 95% identical to the sequence of SEQ ID NO:202, HFR3 is at least 95% identical to the sequence of SEQ ID NO:203; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:204.

[0735] Clause 256. The method according to any one of clauses 179 to 255, wherein the first antigen binding moiety comprises a variable light chain (VL) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

[0736] Clause 257. The method according to any one of clauses 179 to 256, wherein the first antigen binding moiety comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

[0737] Clause 258. The method according to any one of clauses 179 to 257, wherein the first antigen binding moiety is an antibody fragment.

[0738] Clause 259. A method according to any one of clauses 179 to 257, further comprising

[0739] (c) transporting the ABP to the cell surface.

[0740] Clause 260. A method of internalizing a target molecule, the method comprising:

[0741] (a) contacting the target molecule with an antigen binding protein (ABP) according to any one of clauses 1 to 108 or a bifunctional molecule according to any one of clauses 109 to 178; and

[0742] (b) The ABP is internalized into cells.

[0743] 9. Equivalents

[0744] While the invention has been particularly shown and described with reference to a preferred embodiment and various alternative embodiments, it will be understood by those skilled in the relevant art that various changes in form and details may be made therein without departing from the spirit and scope of the invention.

Claims

1. An antigen binding protein (ABP) comprising a first antigen binding moiety that specifically binds to an internalization domain of a cation-independent mannose 6-phosphate receptor (CI-M6PR) that facilitates intracellular transport of the ABP.

2. The ABP of claim 1, further comprising a target binding portion that specifically binds to a target molecule.

3. The ABP of claim 2, wherein the target binding moiety is a second antigen binding moiety that specifically binds to the target molecule.

4. The ABP of claim 2, wherein the target binding moiety is conjugated to the first antigen binding moiety, optionally via a linker.

5. The ABP according to any one of claims 2 to 4, wherein the target molecule is a soluble extracellular target molecule or a cell surface target molecule.

6. The ABP of any preceding claim, further comprising an attached cargo moiety.

7. The ABP of claim 6, wherein the cargo moiety is a polypeptide fused to the first antigen binding moiety or the second antigen binding moiety.

8. The ABP of claim 6, wherein the cargo moiety is conjugated to the first antigen binding moiety or the second antigen binding moiety, optionally via a linker.

9. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to a domain of CI-M6PR selected from the group consisting of domain 1, domain 4, domain 5, domain 6, domain 7, and domain 8.

10. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to domain 1 of CI-M6PR.

11. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to domain 4 of CI-M6PR.

12. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to domain 5 of CI-M6PR.

13. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to domain 6 of CI-M6PR.

14. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to domain 7 of CI-M6PR.

15. The ABP of claim 1 or 2, wherein the first antigen binding portion binds to domain 8 of CI-M6PR.

16. The ABP of any of the preceding claims, wherein the first antigen binding portion binds the CI-M6PR with high affinity.

17. The ABP of claim 16, wherein the first antigen binding moiety has a dissociation equilibrium constant (K) of about 10 nM or less. D ) combined with CI-M6PR.

18. The ABP of any of the preceding claims, wherein the first antigen binding portion has a K of between about 1 nM and about 500 nM. D Combined with CI-M6PR.

19. The ABP of claim 18, wherein the first antigen binding portion has a K between about 10 nM and about 100 nM. D Combined with CI-M6PR.

20. The ABP of claim 18, wherein the first antigen binding moiety has a K between about 100 nM and about 200 nM. D Combined with CI-M6PR.

21. The ABP of claim 18, wherein the first antigen binding portion has a K between about 200 nM and about 300 nM. D Combined with CI-M6PR.

22. The ABP of claim 18, wherein the first antigen binding portion has a K between about 300 nM and about 400 nM. D Combined with CI-M6PR.

23. The ABP of claim 18, wherein the first antigen binding portion has a K between about 400 nM and about 500 nM. D Combined with CI-M6PR.

24. The ABP according to any one of the preceding claims, wherein the dissociation rate (K) of the first antigen binding portion to CI-M6PR is off ) in 1x 10 -8 s -1 With 0.1s -1 between.

25. The ABP according to any of the preceding claims, wherein the first antigen binding portion is directed against the K of CI-M6PR. off In 1x 10 -6 s -1 With 1x 10 -2 s -1 between.

26. The ABP according to any of the preceding claims, wherein the first antigen binding moiety is directed against the K of CI-M6PR. off About 1x 10 -5 s -1 Or slower.

27. The ABP of any of the preceding claims, wherein the binding of the first antigen binding moiety to CI-M6PR is pH dependent.

28. The ABP of any of the preceding claims, wherein the first antigen binding portion is released from CI-M6PR at a pH of 7.4 or less.

29. The ABP of any of the preceding claims, wherein the first antigen binding portion is released from CI-M6PR at a pH of 6.5 or less.

30. The ABP of any of the preceding claims, wherein the first antigen binding portion is released from CI-M6PR at a pH of 6.0 or less.

31. The ABP of any of the preceding claims, wherein the first antigen binding portion is released from CI-M6PR at a pH of 5.5 or less.

32. The ABP of any of the preceding claims, wherein the first antigen binding portion is released from CI-M6PR at a pH of 5.0 or less.

33. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a light chain CDR3 (LCDR3) having the sequence of SEQ ID NO: 76 and a heavy chain CDR3 (HCDR3) having the sequence of SEQ ID NO:

35.

34. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 78 and a heavy chain CDR3 having the amino acid sequence of SEQ ID NO:

38.

35. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 81 and a heavy chain CDR3 having the sequence of SEQ ID NO:

41.

36. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 84 and a heavy chain CDR3 having the sequence of SEQ ID NO:

44.

37. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 85 and a heavy chain CDR3 having the sequence of SEQ ID NO:

46.

38. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 88 and a heavy chain CDR3 having the sequence of SEQ ID NO:

49.

39. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 91 and a heavy chain CDR3 having the sequence of SEQ ID NO:

52.

40. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 94 and a heavy chain CDR3 having the sequence of SEQ ID NO:

54.

41. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 96 and a heavy chain CDR3 having the sequence of SEQ ID NO:

55.

42. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 99 and a heavy chain CDR3 having the sequence of SEQ ID NO:

58.

43. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 101 and a heavy chain CDR3 having the sequence of SEQ ID NO:

60.

44. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 104 and a heavy chain CDR3 having the sequence of SEQ ID NO:

63.

45. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 107 and a heavy chain CDR3 having the sequence of SEQ ID NO:

66.

46. ​​The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 110 and a heavy chain CDR3 having the sequence of SEQ ID NO:

69.

47. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 113 and a heavy chain CDR3 having the sequence of SEQ ID NO:

72.

48. The ABP of any one of claims 1 to 32, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 116 and a heavy chain CDR3 having the sequence of SEQ ID NO:

73.

49. The ABP of claim 33, wherein the first antigen binding portion further comprises a light chain CDR1 (LCDR1) having the sequence of SEQ ID NO: 74 and a heavy chain CDR1 (HCDR1) having the sequence of SEQ ID NO: 33; and a light chain CDR2 (LCDR2) having the sequence of SEQ ID NO: 75 and a heavy chain CDR2 (HCDR2) having the sequence of SEQ ID NO:

34.

50. The ABP of claim 34, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 77 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 36; and a LCDR2 having the sequence of SEQ ID NO: 75 and a HCDR2 having the sequence of SEQ ID NO:

37.

51. The ABP of claim 35, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 39; and a LCDR2 having the sequence of SEQ ID NO: 80 and a HCDR2 having the sequence of SEQ ID NO:

40.

52. The ABP of claim 36, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 82 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 42; and a LCDR2 having the sequence of SEQ ID NO: 83 and a HCDR2 having the sequence of SEQ ID NO:

43.

53. The ABP of claim 37, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 45; and a LCDR2 having the sequence of SEQ ID NO: 80 and a HCDR2 having the sequence of SEQ ID NO:

34.

54. The ABP of claim 38, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 86 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 47; and a LCDR2 having the sequence of SEQ ID NO: 87 and a HCDR2 having the sequence of SEQ ID NO:

48.

55. The ABP of claim 39, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:89 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:90 and a HCDR2 having the sequence of SEQ ID NO:

53.

56. The ABP of claim 40, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:92 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:93 and a HCDR2 having the sequence of SEQ ID NO:

53.

57. The ABP of claim 41, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:95 and a heavy chain HCDR1 having the sequence of SEQ ID NO:50; and a LCDR2 having the sequence of SEQ ID NO:93 and a HCDR2 having the sequence of SEQ ID NO:

51.

58. The ABP of claim 42, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO:97 and a heavy chain HCDR1 having the sequence of SEQ ID NO:56; and a LCDR2 having the sequence of SEQ ID NO:98 and a HCDR2 having the sequence of SEQ ID NO:

57.

59. The ABP of claim 43, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 100 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 50; and a LCDR2 having the sequence of SEQ ID NO: 93 and a HCDR2 having the sequence of SEQ ID NO:

59.

60. The ABP of claim 44, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 102 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 61; and a LCDR2 having the sequence of SEQ ID NO: 103 and a HCDR2 having the sequence of SEQ ID NO:

62.

61. The ABP of claim 45, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 105 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 64; and a LCDR2 having the sequence of SEQ ID NO: 106 and a HCDR2 having the sequence of SEQ ID NO:

65.

62. The ABP of claim 46, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 67; and a LCDR2 having the sequence of SEQ ID NO: 109 and a HCDR2 having the sequence of SEQ ID NO:

68.

63. The ABP of claim 47, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 70; and a LCDR2 having the sequence of SEQ ID NO: 112 and a HCDR2 having the sequence of SEQ ID NO:

71.

64. The ABP of claim 48, wherein the first antigen binding portion further comprises a LCDR1 having the sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having the sequence of SEQ ID NO: 70; and a LCDR2 having the sequence of SEQ ID NO: 115 and a HCDR2 having the sequence of SEQ ID NO:

71.

65. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:205, LFR2 is at least 95% identical to the sequence of SEQ ID NO:206, LFR3 is at least 95% identical to the sequence of SEQ ID NO:207; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

208.

66. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:209, LFR2 is at least 95% identical to the sequence of SEQ ID NO:210, LFR3 is at least 95% identical to the sequence of SEQ ID NO:211; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

212.

67. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:213, LFR2 is at least 95% identical to the sequence of SEQ ID NO:214, LFR3 is at least 95% identical to the sequence of SEQ ID NO:215; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

216.

68. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:217, LFR2 is at least 95% identical to the sequence of SEQ ID NO:218, LFR3 is at least 95% identical to the sequence of SEQ ID NO:219; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

220.

69. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:221, LFR2 is at least 95% identical to the sequence of SEQ ID NO:222, LFR3 is at least 95% identical to the sequence of SEQ ID NO:223; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

224.

70. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:225, LFR2 is at least 95% identical to the sequence of SEQ ID NO:226, LFR3 is at least 95% identical to the sequence of SEQ ID NO:227; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

228.

71. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:229, LFR2 is at least 95% identical to the sequence of SEQ ID NO:230, LFR3 is at least 95% identical to the sequence of SEQ ID NO:231; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

232.

72. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 233, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 234, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 235; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

236.

73. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:237, LFR2 is at least 95% identical to the sequence of SEQ ID NO:238, LFR3 is at least 95% identical to the sequence of SEQ ID NO:239; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

240.

74. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:241, LFR2 is at least 95% identical to the sequence of SEQ ID NO:242, LFR3 is at least 95% identical to the sequence of SEQ ID NO:243; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

244.

75. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:245, LFR2 is at least 95% identical to the sequence of SEQ ID NO:246, LFR3 is at least 95% identical to the sequence of SEQ ID NO:247; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

248.

76. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:249, LFR2 is at least 95% identical to the sequence of SEQ ID NO:250, LFR3 is at least 95% identical to the sequence of SEQ ID NO:251; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

252.

77. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:253, LFR2 is at least 95% identical to the sequence of SEQ ID NO:254, LFR3 is at least 95% identical to the sequence of SEQ ID NO:255; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

256.

78. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 257, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 258, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 259; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

260.

79. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:261, LFR2 is at least 95% identical to the sequence of SEQ ID NO:262, LFR3 is at least 95% identical to the sequence of SEQ ID NO:263; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

264.

80. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:265, LFR2 is at least 95% identical to the sequence of SEQ ID NO:266, LFR3 is at least 95% identical to the sequence of SEQ ID NO:267; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

268.

81. The ABP of claim 65, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 141, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 142, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 143; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

144.

82. The ABP of claim 66, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 145, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 146, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 147; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

148.

83. The ABP of claim 67, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 149, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 150, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 151; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

152.

84. The ABP of claim 68, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 153, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 154, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 155; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

156.

85. The ABP of claim 69, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 157, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 158, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 159; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

160.

86. The ABP of claim 70, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 161, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 162, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 163; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

164.

87. The ABP of claim 71, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 165, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 166, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 167; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

168.

88. The ABP of claim 72, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 169, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 170, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 171; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

172.

89. The ABP of claim 73, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 173, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 174, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 175; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

176.

90. The ABP of claim 74, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 177, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 178, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 179; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

180.

91. The ABP of claim 75, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 181, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 182, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 183; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

184.

92. The ABP of claim 76, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 185, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 186, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 187; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

188.

93. The ABP of claim 77, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 189, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 190, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 191; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

192.

94. The ABP of claim 78, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 193, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 194, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 195; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

196.

95. The ABP of claim 79, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 197, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 198, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 199; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

200.

96. The ABP of claim 80, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:201, HFR2 is at least 95% identical to the sequence of SEQ ID NO:202, HFR3 is at least 95% identical to the sequence of SEQ ID NO:203; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

204.

97. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable light chain (VL) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

98. The ABP of any of the preceding claims, wherein the first antigen binding portion comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

99. The ABP of any of the preceding claims, wherein the first antigen binding moiety is an antibody fragment (eg, a single chain variable fragment (scFv) or an antigen binding fragment).

100. The ABP of any of the preceding claims, wherein the ABP is transported to the lysosome.

101. The ABP of any of the preceding claims, wherein the ABP is transported back to the cell surface following internalization.

102. The ABP of any one of claims 1 to 100, wherein the ABP is degraded in the cell.

103. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein The LCDR1 has a sequence of SEQ ID NO: 82, The LCDR2 has a sequence of SEQ ID NO: 83, The LCDR3 has a sequence of SEQ ID NO: 84, The HCDR1 has the sequence of SEQ ID NO:42, The HCDR2 has the sequence of SEQ ID NO: 43, and The HCDR3 has the sequence of SEQ ID NO:

44.

104. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein The LCDR1 has a sequence of SEQ ID NO: 92, The LCDR2 has a sequence of SEQ ID NO: 93, The LCDR3 has a sequence of SEQ ID NO: 94, The HCDR1 has the sequence of SEQ ID NO: 50, The HCDR2 has the sequence of SEQ ID NO: 53, and The HCDR3 has the sequence of SEQ ID NO:

54.

105. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein The LCDR1 has a sequence of SEQ ID NO: 97, The LCDR2 has a sequence of SEQ ID NO: 98, The LCDR3 has a sequence of SEQ ID NO: 99, The HCDR1 has the sequence of SEQ ID NO: 56, The HCDR2 has the sequence of SEQ ID NO: 57, and The HCDR3 has the sequence of SEQ ID NO:

58.

106. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein The LCDR1 has a sequence of SEQ ID NO: 100, The LCDR2 has a sequence of SEQ ID NO: 93, The LCDR3 has a sequence of SEQ ID NO: 101, The HCDR1 has the sequence of SEQ ID NO: 50, The HCDR2 has the sequence of SEQ ID NO: 59, and The HCDR3 has the sequence of SEQ ID NO:

60.

107. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein The LCDR1 has a sequence of SEQ ID NO: 108, The LCDR2 has a sequence of SEQ ID NO: 109, The LCDR3 has a sequence of SEQ ID NO: 110, The HCDR1 has the sequence of SEQ ID NO: 67, The HCDR2 has the sequence of SEQ ID NO: 68, and The HCDR3 has the sequence of SEQ ID NO:

69.

108. An antigen binding protein (ABP) comprising a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), the first antigen binding portion comprising a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), a light chain CDR3 (LCDR3), a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2) and a heavy chain CDR3 (HCDR3), wherein The LCDR1 has a sequence of SEQ ID NO: 111, The LCDR2 has a sequence of SEQ ID NO: 112, The LCDR3 has a sequence of SEQ ID NO: 113, The HCDR1 has the sequence of SEQ ID NO: 70, The HCDR2 has the sequence of SEQ ID NO: 71, and The HCDR3 has the sequence of SEQ ID NO:

72.

109. A bifunctional molecule comprising: A first portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR), wherein The first moiety is an antibody or antibody fragment; and A second portion that specifically binds to a cell surface target molecule or an extracellular target molecule, wherein the second portion is selected from an antibody, an antigen binding fragment, a ligand, and a small molecule.

110. The bifunctional molecule of claim 109, wherein the bifunctional molecule is a polypeptide.

111. The bifunctional molecule of claim 109, wherein the first portion and the second portion are covalently attached via a linker.

112. The bifunctional molecule of claim 111, wherein the second moiety is a small molecule.

113. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 (LCDR3) having the sequence of SEQ ID NO: 76 and a heavy chain CDR3 (HCDR3) having the sequence of SEQ ID NO:

35.

114. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 78 and a heavy chain CDR3 having the amino acid sequence of SEQ ID NO:

38.

115. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 81 and a heavy chain CDR3 having the sequence of SEQ ID NO:

41.

116. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 84 and a heavy chain CDR3 having the sequence of SEQ ID NO:

44.

117. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 85 and a heavy chain CDR3 having the sequence of SEQ ID NO:

46.

118. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 88 and a heavy chain CDR3 having the sequence of SEQ ID NO:

49.

119. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:91 and a heavy chain CDR3 having the sequence of SEQ ID NO:

52.

120. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 94 and a heavy chain CDR3 having the sequence of SEQ ID NO:

54.

121. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO:96 and a heavy chain CDR3 having the sequence of SEQ ID NO:

55.

122. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 99 and a heavy chain CDR3 having the sequence of SEQ ID NO:

58.

123. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 101 and a heavy chain CDR3 having the sequence of SEQ ID NO:

60.

124. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 104 and a heavy chain CDR3 having the sequence of SEQ ID NO:

63.

125. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 107 and a heavy chain CDR3 having the sequence of SEQ ID NO:

66.

126. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 110 and a heavy chain CDR3 having the sequence of SEQ ID NO:

69.

127. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 113 and a heavy chain CDR3 having the sequence of SEQ ID NO:

72.

128. The bifunctional molecule of any one of claims 109 to 112, wherein the first binding moiety comprises a light chain CDR3 having the sequence of SEQ ID NO: 116 and a heavy chain CDR3 having the sequence of SEQ ID NO:

73.

129. The bifunctional molecule of claim 113, wherein the first portion further comprises a light chain CDR1 (LCDR1) having the sequence of SEQ ID NO: 74 and a heavy chain CDR1 (HCDR1) having the sequence of SEQ ID NO: 33; and a light chain CDR2 (LCDR2) having the sequence of SEQ ID NO: 75 and a heavy chain CDR2 (HCDR2) having the sequence of SEQ ID NO:

34.

130. The bifunctional molecule of claim 114, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:77 and a heavy chain HCDR1 having a sequence of SEQ ID NO:36; and a LCDR2 having a sequence of SEQ ID NO:75 and a HCDR2 having a sequence of SEQ ID NO:

37.

131. The bifunctional molecule of claim 115, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 39; and a LCDR2 having a sequence of SEQ ID NO: 80 and a HCDR2 having a sequence of SEQ ID NO:

40.

132. The bifunctional molecule of claim 116, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 82 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 42; and a LCDR2 having a sequence of SEQ ID NO: 83 and a HCDR2 having a sequence of SEQ ID NO:

43.

133. The bifunctional molecule of claim 117, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 79 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 45; and a LCDR2 having a sequence of SEQ ID NO: 80 and a HCDR2 having a sequence of SEQ ID NO:

34.

134. The bifunctional molecule of claim 118, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 86 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 47; and a LCDR2 having a sequence of SEQ ID NO: 87 and a HCDR2 having a sequence of SEQ ID NO:

48.

135. The bifunctional molecule of claim 119, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:89 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:90 and a HCDR2 having a sequence of SEQ ID NO:

53.

136. The bifunctional molecule of claim 120, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:92 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:

53.

137. The bifunctional molecule of claim 121, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO:95 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:

51.

138. The bifunctional molecule of claim 122, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 97 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 56; and a LCDR2 having a sequence of SEQ ID NO: 98 and a HCDR2 having a sequence of SEQ ID NO:

57.

139. The bifunctional molecule of claim 123, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 100 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 50; and a LCDR2 having a sequence of SEQ ID NO: 93 and a HCDR2 having a sequence of SEQ ID NO:

59.

140. The bifunctional molecule of claim 124, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 102 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 61; and a LCDR2 having a sequence of SEQ ID NO: 103 and a HCDR2 having a sequence of SEQ ID NO:

62.

141. The bifunctional molecule of claim 125, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 105 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 64; and a LCDR2 having a sequence of SEQ ID NO: 106 and a HCDR2 having a sequence of SEQ ID NO:

65.

142. The bifunctional molecule of claim 126, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 67; and a LCDR2 having a sequence of SEQ ID NO: 109 and a HCDR2 having a sequence of SEQ ID NO:

68.

143. The bifunctional molecule of claim 127, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 112 and a HCDR2 having a sequence of SEQ ID NO:

71.

144. The bifunctional molecule of claim 128, wherein the first portion further comprises a LCDR1 having a sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 115 and a HCDR2 having a sequence of SEQ ID NO:

71.

145. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:205, LFR2 is at least 95% identical to the sequence of SEQ ID NO:206, LFR3 is at least 95% identical to the sequence of SEQ ID NO:207; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

208.

146. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 209, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 210, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 211; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

212.

147. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 213, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 214, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 215; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

216.

148. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 217, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 218, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 219; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

220.

149. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 221, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 222, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 223; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

224.

150. The bifunctional molecule of any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 225, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 226, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 227; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

228.

151. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 229, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 230, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 231; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

232.

152. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 233, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 234, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 235; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

236.

153. The bifunctional molecule of any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 237, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 238, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 239; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

240.

154. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 241, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 242, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 243; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

244.

155. The bifunctional molecule of any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 245, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 246, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 247; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

248.

156. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 249, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 250, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 251; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

252.

157. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 253, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 254, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 255; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

256.

158. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 257, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 258, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 259; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

260.

159. A bifunctional molecule according to any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 261, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 262, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 263; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

264.

160. The bifunctional molecule of any one of claims 109 to 144, wherein the first portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 265, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 266, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 267; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

268.

161. The bifunctional molecule of claim 145, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 141, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 142, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 143; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

144.

162. The bifunctional molecule of claim 146, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 145, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 146, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 147; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

148.

163. The bifunctional molecule of claim 147, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 149, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 150, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 151; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

152.

164. The bifunctional molecule of claim 148, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 153, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 154, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 155; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

156.

165. The bifunctional molecule of claim 149, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 157, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 158, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 159; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

160.

166. The bifunctional molecule of claim 150, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 161, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 162, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 163; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

164.

167. The bifunctional molecule of claim 151, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 165, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 166, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 167; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

168.

168. The bifunctional molecule of claim 152, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 169, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 170, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 171; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

172.

169. The bifunctional molecule of claim 153, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 173, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 174, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 175; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

176.

170. The bifunctional molecule of claim 154, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 177, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 178, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 179; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

180.

171. The bifunctional molecule of claim 155, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 181, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 182, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 183; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

184.

172. The bifunctional molecule of claim 156, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 185, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 186, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 187; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

188.

173. The bifunctional molecule of claim 157, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 189, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 190, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 191; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

192.

174. The bifunctional molecule of claim 158, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 193, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 194, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 195; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

196.

175. The bifunctional molecule of claim 159, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO: 197, HFR2 is at least 95% identical to the sequence of SEQ ID NO: 198, HFR3 is at least 95% identical to the sequence of SEQ ID NO: 199; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

200.

176. The bifunctional molecule of claim 160, wherein the first portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:201, HFR2 is at least 95% identical to the sequence of SEQ ID NO:202, HFR3 is at least 95% identical to the sequence of SEQ ID NO:203; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

204.

177. The bifunctional molecule of any one of claims 109 to 176, wherein the first portion comprises a variable light chain (VL) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

178. The bifunctional molecule of any one of claims 109 to 177, wherein the first portion comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

179. A method for degrading a soluble target molecule or a cell surface target molecule, the method comprising: (a) contacting the target molecule with a multispecific antigen binding protein (ABP), wherein the ABP comprises a first antigen binding portion that specifically binds to a cation-independent mannose 6-phosphate receptor (CI-M6PR) on the surface of a cell; and (b) transporting the ABP, the target molecule and the CI-M6PR to a lysosome in a cell, wherein the target molecule is degraded in the lysosome.

180. The method of claim 179, wherein the ABP further comprises a cargo moiety attached.

181. The method of claim 180, wherein the cargo moiety is a polypeptide fused to the ABP.

182. The method of claim 181, wherein the cargo moiety is conjugated to the ABP, optionally via a linker.

183. The method of any one of claims 179 to 182, wherein the first antigen binding portion binds to a domain of CI-M6PR selected from the group consisting of domain 1, domain 4, domain 5, domain 6, domain 7, and domain 8.

184. The method of any one of claims 179 to 182, wherein the first antigen binding moiety has a dissociation equilibrium constant (K) of about 10 nM or less. D ) combined with CI-M6PR.

185. The method of any one of claims 179 to 182, wherein the first antigen binding moiety has a dissociation equilibrium constant (K) between about 1 nM and about 500 nM. D ) combined with CI-M6PR.

186. The method of any one of claims 179 to 185, wherein the dissociation rate (k) of the first antigen binding moiety to CI-M6PR is off ) in 1x 10 -8 s -1 With 0.1s -1 between.

187. The method of any one of claims 179 to 186, wherein the binding of the first antigen binding moiety to CI-M6PR is pH dependent.

188. The method of any one of claims 179 to 187, wherein the first antigen binding portion is released from CI-M6PR at a pH of 7.4 or less.

189. The method of any one of claims 179 to 188, wherein the first antigen binding portion is released from CI-M6PR at a pH of 6.0 or less.

190. The method of any one of claims 179 to 189, wherein the first antigen binding portion is released from CI-M6PR at a pH of 5.5 or less.

191. The method of any one of claims 179 to 190, wherein the first antigen binding portion is released from CI-M6PR at a pH of 5.0 or less.

192. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 (LCDR3) having the sequence of SEQ ID NO:76 and a heavy chain CDR3 (HCDR3) having the sequence of SEQ ID NO:

35.

193. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:78 and a heavy chain CDR3 having the amino acid sequence of SEQ ID NO:

38.

194. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:81 and a heavy chain CDR3 having the sequence of SEQ ID NO:

41.

195. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:84 and a heavy chain CDR3 having the sequence of SEQ ID NO:

44.

196. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:85 and a heavy chain CDR3 having the sequence of SEQ ID NO:

46.

197. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:88 and a heavy chain CDR3 having the sequence of SEQ ID NO:

49.

198. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:91 and a heavy chain CDR3 having the sequence of SEQ ID NO:

52.

199. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:94 and a heavy chain CDR3 having the sequence of SEQ ID NO:

54.

200. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:96 and a heavy chain CDR3 having the sequence of SEQ ID NO:

55.

201. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO:99 and a heavy chain CDR3 having the sequence of SEQ ID NO:

58.

202. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 101 and a heavy chain CDR3 having the sequence of SEQ ID NO:

60.

203. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 104 and a heavy chain CDR3 having the sequence of SEQ ID NO:

63.

204. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 107 and a heavy chain CDR3 having the sequence of SEQ ID NO:

66.

205. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 110 and a heavy chain CDR3 having the sequence of SEQ ID NO:

69.

206. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 113 and a heavy chain CDR3 having the sequence of SEQ ID NO:

72.

207. The method of any one of claims 179 to 191, wherein the first antigen binding portion comprises a light chain CDR3 having the sequence of SEQ ID NO: 116 and a heavy chain CDR3 having the sequence of SEQ ID NO:

73.

208. The method of claim 192, wherein the first antigen binding portion further comprises a light chain CDR1 (LCDR1) having a sequence of SEQ ID NO:74 and a heavy chain CDR1 (HCDR1) having a sequence of SEQ ID NO:33; and a light chain CDR2 (LCDR2) having a sequence of SEQ ID NO:75 and a heavy chain CDR2 (HCDR2) having a sequence of SEQ ID NO:

34.

209. The method of claim 193, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:77 and a heavy chain HCDR1 having a sequence of SEQ ID NO:36; and a LCDR2 having a sequence of SEQ ID NO:75 and a HCDR2 having a sequence of SEQ ID NO:

37.

210. The method of claim 194, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:79 and a heavy chain HCDR1 having a sequence of SEQ ID NO:39; and a LCDR2 having a sequence of SEQ ID NO:80 and a HCDR2 having a sequence of SEQ ID NO:

40.

211. The method of claim 195, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:82 and a heavy chain HCDR1 having a sequence of SEQ ID NO:42; and a LCDR2 having a sequence of SEQ ID NO:83 and a HCDR2 having a sequence of SEQ ID NO:

43.

212. The method of claim 196, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:79 and a heavy chain HCDR1 having a sequence of SEQ ID NO:45; and a LCDR2 having a sequence of SEQ ID NO:80 and a HCDR2 having a sequence of SEQ ID NO:

34.

213. The method of claim 197, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:86 and a heavy chain HCDR1 having a sequence of SEQ ID NO:47; and a LCDR2 having a sequence of SEQ ID NO:87 and a HCDR2 having a sequence of SEQ ID NO:

48.

214. The method of claim 198, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:89 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:90 and a HCDR2 having a sequence of SEQ ID NO:

53.

215. The method of claim 199, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:92 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:

53.

216. The method of claim 200, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:95 and a heavy chain HCDR1 having a sequence of SEQ ID NO:50; and a LCDR2 having a sequence of SEQ ID NO:93 and a HCDR2 having a sequence of SEQ ID NO:

51.

217. The method of claim 201, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO:97 and a heavy chain HCDR1 having a sequence of SEQ ID NO:56; and a LCDR2 having a sequence of SEQ ID NO:98 and a HCDR2 having a sequence of SEQ ID NO:

57.

218. The method of claim 202, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 100 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 50; and a LCDR2 having a sequence of SEQ ID NO: 93 and a HCDR2 having a sequence of SEQ ID NO:

59.

219. The method of claim 203, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 102 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 61; and a LCDR2 having a sequence of SEQ ID NO: 103 and a HCDR2 having a sequence of SEQ ID NO:

62.

220. The method of claim 204, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 105 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 64; and a LCDR2 having a sequence of SEQ ID NO: 106 and a HCDR2 having a sequence of SEQ ID NO:

65.

221. The method of claim 205, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 108 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 67; and a LCDR2 having a sequence of SEQ ID NO: 109 and a HCDR2 having a sequence of SEQ ID NO:

68.

222. The method of claim 206, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 111 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 112 and a HCDR2 having a sequence of SEQ ID NO:

71.

223. The method of claim 207, wherein the first antigen binding portion further comprises a LCDR1 having a sequence of SEQ ID NO: 114 and a heavy chain HCDR1 having a sequence of SEQ ID NO: 70; and a LCDR2 having a sequence of SEQ ID NO: 115 and a HCDR2 having a sequence of SEQ ID NO:

71.

224. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:205, LFR2 is at least 95% identical to the sequence of SEQ ID NO:206, LFR3 is at least 95% identical to the sequence of SEQ ID NO:207; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

208.

225. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 209, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 210, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 211; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

212.

226. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:213, LFR2 is at least 95% identical to the sequence of SEQ ID NO:214, LFR3 is at least 95% identical to the sequence of SEQ ID NO:215; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

216.

227. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:217, LFR2 is at least 95% identical to the sequence of SEQ ID NO:218, LFR3 is at least 95% identical to the sequence of SEQ ID NO:219; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

220.

228. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:221, LFR2 is at least 95% identical to the sequence of SEQ ID NO:222, LFR3 is at least 95% identical to the sequence of SEQ ID NO:223; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

224.

229. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:225, LFR2 is at least 95% identical to the sequence of SEQ ID NO:226, LFR3 is at least 95% identical to the sequence of SEQ ID NO:227; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

228.

230. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:229, LFR2 is at least 95% identical to the sequence of SEQ ID NO:230, LFR3 is at least 95% identical to the sequence of SEQ ID NO:231; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

232.

231. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:233, LFR2 is at least 95% identical to the sequence of SEQ ID NO:234, LFR3 is at least 95% identical to the sequence of SEQ ID NO:235; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

236.

232. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 237, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 238, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 239; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

240.

233. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:241, LFR2 is at least 95% identical to the sequence of SEQ ID NO:242, LFR3 is at least 95% identical to the sequence of SEQ ID NO:243; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

244.

234. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 245, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 246, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 247; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

248.

235. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:249, LFR2 is at least 95% identical to the sequence of SEQ ID NO:250, LFR3 is at least 95% identical to the sequence of SEQ ID NO:251; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

252.

236. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:253, LFR2 is at least 95% identical to the sequence of SEQ ID NO:254, LFR3 is at least 95% identical to the sequence of SEQ ID NO:255; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

256.

237. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:257, LFR2 is at least 95% identical to the sequence of SEQ ID NO:258, LFR3 is at least 95% identical to the sequence of SEQ ID NO:259; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

260.

238. A method according to any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO:261, LFR2 is at least 95% identical to the sequence of SEQ ID NO:262, LFR3 is at least 95% identical to the sequence of SEQ ID NO:263; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

264.

239. The method of any one of claims 179 to 223, wherein the first antigen binding portion comprises a variable light chain (VL) comprising framework regions LFR1, LFR2, LFR3 and LFR4, wherein LFR1 is at least 95% identical to the sequence of SEQ ID NO: 265, LFR2 is at least 95% identical to the sequence of SEQ ID NO: 266, LFR3 is at least 95% identical to the sequence of SEQ ID NO: 267; and LFR4 is at least 95% identical to the sequence of SEQ ID NO:

268.

240. The method of claim 224, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3, and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:141, HFR2 is at least 95% identical to the sequence of SEQ ID NO:142, HFR3 is at least 95% identical to the sequence of SEQ ID NO:143; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

144.

241. A method according to claim 225, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:145, HFR2 is at least 95% identical to the sequence of SEQ ID NO:146, HFR3 is at least 95% identical to the sequence of SEQ ID NO:147; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

148.

242. A method according to claim 226, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:149, HFR2 is at least 95% identical to the sequence of SEQ ID NO:150, HFR3 is at least 95% identical to the sequence of SEQ ID NO:151; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

152.

243. A method according to claim 227, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:153, HFR2 is at least 95% identical to the sequence of SEQ ID NO:154, HFR3 is at least 95% identical to the sequence of SEQ ID NO:155; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

156.

244. A method according to claim 228, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:157, HFR2 is at least 95% identical to the sequence of SEQ ID NO:158, HFR3 is at least 95% identical to the sequence of SEQ ID NO:159; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

160.

245. A method according to claim 229, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:161, HFR2 is at least 95% identical to the sequence of SEQ ID NO:162, HFR3 is at least 95% identical to the sequence of SEQ ID NO:163; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

164.

246. A method according to claim 230, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:165, HFR2 is at least 95% identical to the sequence of SEQ ID NO:166, HFR3 is at least 95% identical to the sequence of SEQ ID NO:167; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

168.

247. A method according to claim 231, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:169, HFR2 is at least 95% identical to the sequence of SEQ ID NO:170, HFR3 is at least 95% identical to the sequence of SEQ ID NO:171; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

172.

248. A method according to claim 232, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:173, HFR2 is at least 95% identical to the sequence of SEQ ID NO:174, HFR3 is at least 95% identical to the sequence of SEQ ID NO:175; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

176.

249. A method according to claim 233, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:177, HFR2 is at least 95% identical to the sequence of SEQ ID NO:178, HFR3 is at least 95% identical to the sequence of SEQ ID NO:179; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

180.

250. A method according to claim 234, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:181, HFR2 is at least 95% identical to the sequence of SEQ ID NO:182, HFR3 is at least 95% identical to the sequence of SEQ ID NO:183; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

184.

251. A method according to claim 235, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:185, HFR2 is at least 95% identical to the sequence of SEQ ID NO:186, HFR3 is at least 95% identical to the sequence of SEQ ID NO:187; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

188.

252. A method according to claim 236, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:189, HFR2 is at least 95% identical to the sequence of SEQ ID NO:190, HFR3 is at least 95% identical to the sequence of SEQ ID NO:191; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

192.

253. A method according to claim 237, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:193, HFR2 is at least 95% identical to the sequence of SEQ ID NO:194, HFR3 is at least 95% identical to the sequence of SEQ ID NO:195; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

196.

254. A method according to claim 238, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:197, HFR2 is at least 95% identical to the sequence of SEQ ID NO:198, HFR3 is at least 95% identical to the sequence of SEQ ID NO:199; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

200.

255. A method according to claim 239, wherein the first antigen binding portion comprises a variable heavy chain (VH) comprising framework regions HFR1, HFR2, HFR3 and HFR4, wherein HFR1 is at least 95% identical to the sequence of SEQ ID NO:201, HFR2 is at least 95% identical to the sequence of SEQ ID NO:202, HFR3 is at least 95% identical to the sequence of SEQ ID NO:203; and HFR4 is at least 95% identical to the sequence of SEQ ID NO:

204.

256. The method of any one of claims 179 to 255, wherein the first antigen binding portion comprises a variable light chain (VL) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 17-32.

257. The method of any one of claims 179 to 256, wherein the first antigen binding portion comprises a variable heavy chain (VH) having an amino acid sequence that is at least 90% identical to an amino acid sequence selected from SEQ ID NOs: 1-16.

258. The method of any one of claims 179 to 257, wherein the first antigen binding moiety is an antibody fragment.

259. The method of any one of claims 179 to 257, further comprising (c) transporting the ABP to the cell surface.

260. A method for internalizing a target molecule, the method comprising: (a) contacting the target molecule with an antigen binding protein (ABP) according to any one of claims 1 to 108 or a bifunctional molecule according to any one of claims 109 to 178; and (b) The ABP is internalized into cells.

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