Anti-MuSK antibodies for use in the treatment of neuromuscular disorders

Abstract

The present invention relates to an anti-MuSK antibody or antigen-binding fragment thereof for use in treating a neuromuscular disorder in a human subject. In certain embodiments, the antibody or antigen-binding fragment thereof is combined with an anticholinergic compound.

Description

[Technical field]

[0001] (Related Applications) This application claims priority to U.S. Provisional Patent Application No. 63 / 364,685, filed May 13, 2022, and EP Application No. 22154118.8, filed January 28, 2022, the entire disclosures of which are incorporated herein by reference.

[0002] (Reference to sequence listing) This application contains a Sequence Listing that has been submitted electronically in ST.26 format and is incorporated herein by reference in its entirety (the ST.26 copy created on January 27, 2023 is named "196198_SL.XML" and is 364,704 bytes in size).

[0003] FIELD OF THEINVENTION The present invention relates to an anti-MuSK antibody or antigen-binding fragment thereof for use in treating a neuromuscular disorder, such as ALS (amyotrophic lateral sclerosis), in a human subject. In one embodiment, the antibody or antigen-binding fragment thereof is combined with an anticholinergic compound. Summary of the Invention

[0004] (background) ALS is an adult-onset, non-cell-autonomous neuromuscular / neurodegenerative disorder that causes progressive loss of upper and lower motor neurons (MNs), resulting in gradual paralysis and death within 2–5 years.Denervation of the neuromuscular junction (NMJ) is a hallmark of ALS [1], is present in several disease models of ALS [2–6], and even precedes the death of MNs [1,3].

[0005] The currently approved ALS treatment (riluzole) only works for 20% of ALS patients by extending their lifespan by about 3 months. The effect of riluzole on muscle function is very limited. Furthermore, ALS is considered a genetically heterogeneous disease that appears to represent several subgroups with different underlying pathology. Currently, there is no available treatment, and it is highly likely that a tailored treatment cannot help all ALS patients due to the different underlying disease mechanisms.

[0006] Therefore, there remains a need for new treatments for ALS and other diseases that mimic ALS ("ALS-like diseases"). [Brief description of the drawings]

[0007] Description of the drawings [Figure 1] Combo treatment improves motor function in SOD1G37R mice. ARGX-119 treatment was started at P400 (pre-disease or asymptomatic) and darifenacin treatment was started at ~P425 (disease onset) and continued until sacrifice (~P520). A) Diagram showing the rotarod apparatus used to assess motor function, coordination, and balance after acceleration on a rotating wheel. B) Fall latency (seconds) on the rotarod for mice treated with ARGX-119 antibody (large grey circle), ARGX-119 + darifenacin combo treatment (black triangle), darifenacin treated mice (small grey circle) vs. double placebo treated mice (grey square) from ~400 to 520 days of age. C) Grip dynamometer used to monitor overall strength of the mice's forelimbs and hindlimbs. D) Evolution of grip strength measurements during the treatment period from ~P400 to P520 for these groups. E) Evolution of body weight measurements during the treatment period from -P400 to P520 in these groups. *p<0.05, ***p<0.001, ****p<0.0001. One-way ANOVA and multiple t-test. [Diagram 2]Treatment improves the contractile properties of the EDL muscle. A) Photograph of the setup of the muscle force transducer and nerve and muscle stimulation electrodes used to induce muscle contractions. Examples of raw data show muscle contractions induced by nerve or muscle stimulation used to calculate the contractile potential ratio. B-C) ​​Peak extension forces of the EDL muscle generated by nerve stimulation (B) or muscle stimulation (C) at different frequencies (5 Hz to 300 Hz) in mice treated with ARGX-119 antibody (grey circles) and ARGX-119 + darifenacin combo treatment (black triangles) versus mice treated with dual placebo (grey squares). D) Histogram showing the mean ± SEM of the contractile potential ratio expressed as a percentage of the EDL muscle, representing the ratio of peak forces generated by nerve stimulation (stimulation frequencies of 5 Hz to 100 Hz) compared to muscle stimulation. E) Histogram showing the mean ± SEM of EDL muscle weights in mice treated with ARGX-119 antibody, combo, and placebo. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. One-way repeated measures ANOVA and multiple t-test. [Diagram 3] Combo treatment improves the contractile properties of the soleus muscle. A-B) Peak stretch force of the soleus muscle (SOL) generated by nerve stimulation (A) or muscle stimulation (B) at different frequencies (5 Hz to 300 Hz). (B) Mice treated with ARGX-119 antibody (grey circles) and ARGX-119 + darifenacin combo treatment (black triangles) versus double placebo-treated mice (grey squares). C) Histograms showing the mean ± SEM of the contractile capacity ratio expressed as a percentage of the SOL muscle, representing the ratio of peak force generated by nerve stimulation (stimulation frequencies from 5 Hz to 100 Hz) compared to muscle stimulation. D) Histograms showing the mean ± SEM of EDL muscle weight for ARGX-119 antibody-treated, combo-treated, and placebo-treated mice. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. One-way repeated measures ANOVA and multiple t-test. [Figure 4]Combo treatment maintains muscle fatigue properties. A) Diagram showing an EDL fatigue protocol consisting of 18 trains of 10 stimuli induced at 120 Hz for 300 ms (1 train per second). Nerve stimulation alone is used 9 out of 10 times, and muscle stimulation is superimposed on the nerve stimulation every 10th stimulus. The fatigue protocol is followed by a 30 min recovery period. B-C) ​​Peak contractile forces during the fatigue protocol and recovery period expressed as a percentage of the initial baseline force generated before the fatigue protocol for nerve stimulation (B) and nerve + muscle (C) of the EDL muscle. Note the higher resistance to fatigue in placebo-treated animals (grey squares) compared to ARGX-119 + darifenacin dual-treated animals (black triangles) and ARGX-119 antibody-treated animals (grey circles). This indicates a marked change in normal fast-twitch properties that are normally highly fatigable. D-E) Peak contractile force during the fatigue protocol and recovery period expressed as a percentage of the initial baseline force generated before the fatigue protocol for nerve stimulated (B) and nerve+muscle (C) SOL muscles. *p<0.05, **p<0.01, ***p<0.001, ***p<0.0001. One-way repeated measures ANOVA and multiple t-test. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0008] Detailed Description of the Invention (General definition) The following terms or definitions are provided solely to aid in the understanding of the present invention. Unless otherwise specifically defined herein, all terms used herein have the same meaning as they would have to a person skilled in the art of the present invention. Those skilled in the art are directed to refer to, in particular, Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Press, Plainsview, New York (1989); and Ausubel et al., Current Protocols in Molecular Biology (Supplement 47), John Wiley & Sons, New York (1999), for definitions and terms in the art. The definitions provided herein should not be construed as having a scope that is not understood by those skilled in the art.

[0009] Unless otherwise indicated, all methods, steps, techniques and operations not specifically described in detail can and have been carried out in a manner known per se, as would be apparent to one skilled in the art, for example, reference is again made in this case to the standard handbooks, the general background mentioned above and the further references cited therein.

[0010] As used herein, the singular forms "a," "an," and "the" include both singular and plural referents unless the context clearly dictates otherwise.

[0011] The terms "comprising," "comprises," and "comprised of," as used herein, are synonymous with "including," "includes," or "containing," and are inclusive or open-ended and do not exclude additional members, compounds, products, elements, or method steps not listed. The phrase "consisting essentially of," when used in the context of a product or composition (a "product consisting essentially of" or a "composition consisting essentially of") means that additional molecules may be present, but such molecules do not change / alter the properties / activity / functionality of the product or composition. For example, a composition may consist essentially of an antibody or an antibody fragment if the composition itself exhibits similar properties / activity / functionality as one of the antibodies or one of the antibody fragments.

[0012] The recitation of numerical ranges by endpoints includes not only the recited endpoints but also all numbers and fractions subsumed within each range.

[0013] The term "about" as used herein when referring to a measurable value, e.g., a parameter, amount, time duration, etc., is meant to encompass variations of the specified value and of no more than ±10%, preferably no more than ±5%, more preferably no more than ±1%, and even more preferably no more than ±0.1% from the specified value, to the extent that such variations are appropriate for practice in the disclosed invention. It should be understood that the value to which the "about" modifier refers is itself specifically and preferably disclosed.

[0014] The terms "disorder" and "disease" are used interchangeably herein.

[0015] As used herein, amino acid residues are designated by their full name or according to the standard three letter or one letter amino acid code.

[0016] As used herein, the terms "polypeptide" or "protein" are used interchangeably and refer to polymeric forms of amino acids of any length, which can include coded and non-coded amino acids, chemically or biochemically modified or derivatized amino acids, and polypeptides with modified peptide backbones. A "peptide" is also a polymer of amino acids, usually up to 50 amino acids in length. A polypeptide or peptide is represented by an amino acid sequence.

[0017] As used herein, the terms "nucleic acid molecule", "polynucleotide", "polynucleic acid", and "nucleic acid" are used interchangeably and refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof. Nucleic acid molecules are represented by nucleic acid sequences, which are characterized primarily by their base sequence. Polynucleotides can have any three-dimensional structure and can perform any function, known or unknown. Non-limiting examples of polynucleotides include genes, gene fragments, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, control regions, isolated RNA of any sequence, nucleic acid probes, and primers. Nucleic acid molecules can be linear or circular.

[0018] As used herein, the term "homology" refers to the identity or similarity of at least the secondary structure between two polymers, particularly between two polypeptides or polynucleotides, from the same or different taxa, where the similarity is due to a common ancestor. Thus, the term "homolog" refers to such related polymers having similarities in the secondary structure and optionally the tertiary structure. To compare two or more nucleotide sequences, the "sequence identity (percentage)" between a first nucleotide sequence and a second nucleotide sequence can be calculated using methods known to those skilled in the art, for example, by dividing the number of nucleotides in the first nucleotide sequence that are identical to the nucleotides at the corresponding positions in the second nucleotide sequence by the total number of nucleotides in the first nucleotide sequence and multiplying by 100%, or by using known computer algorithms for sequence alignment, such as NCBI Blast. When determining the degree of sequence similarity between two amino acid sequences, those skilled in the art can take into account so-called "conservative" amino acid substitutions, which can be generally described as amino acid substitutions in which an amino acid residue is replaced with another amino acid residue of similar chemical structure and has little or essentially no effect on the function, activity, or other biological properties of a polypeptide. Possible conservative amino acid substitutions have already been exemplified herein. Amino acid sequences and nucleic acid sequences are said to be "exactly the same" if they have 100% sequence identity over their entire length.

[0019] Throughout this application, whenever a particular amino acid sequence SEQ ID NO, for example SEQ ID NO: Y, is mentioned, it can be replaced with: a polypeptide comprising an amino acid sequence having at least 80% sequence identity or similarity to amino acid sequence SEQ ID NO: Y. Throughout this application, the phrase "a sequence is at least X% identical to another sequence" can be replaced with "a sequence has at least X% sequence identity to another sequence."

[0020] Each amino acid sequence described herein by its percentage identity (at least 80%) with each given amino acid sequence has, in further preferred embodiments, at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identity with each given amino acid sequence. In preferred embodiments, sequence identity is determined by comparing the full length of the sequences identified herein. Each amino acid sequence described herein by its percentage of similarity (at least 80%) with each given amino acid sequence has, in a further preferred embodiment, at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more similarity with each given amino acid sequence. In a preferred embodiment, sequence similarity is determined by comparing the full length of the sequences identified herein. Unless otherwise indicated herein, identity or similarity with a given SEQ ID NO refers to identity or similarity based on the full length of the sequence (i.e., over its entire length or as a whole).

[0021] "Sequence identity" is defined herein as the relationship between two or more amino acid (polypeptide or protein) sequences or two or more nucleic acid (polynucleotide) sequences, as determined by comparing the sequences. The identity between two amino acid sequences is preferably defined by assessing their identity over the entire SEQ ID NO: identified herein, or over a portion thereof. The portion may mean at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the length of the SEQ ID NO:

[0022] In the art, "identity" also means the degree of sequence relatedness between amino acid sequences, sometimes as determined by the match between strings of such sequences. "Similarity" between two amino acid sequences is determined by comparing the amino acid sequence and its conserved amino acid substitutes of one polypeptide to the sequence of a second polypeptide. "Identity" and "similarity" include, but are not limited to, those used in Computational Molecular Biology, Lesk, AM, ed., Oxford University Press, New York, 1988; Biocomputing: Informatics and Genome Projects, Smith, DW, ed., Academic Press, New York, 1993; Computer Analysis of Sequence Data, Part I, Griffin, AM and Griffin, HG, eds., Humana Press, New Jersey, 1994; Sequence Analysis in Molecular Biology, von Heine, G., Academic Press, 1987; and Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York, 1991; and Carillo, H., and Lipman, This can be readily calculated by known methods, including those described in D., SIAM J. Applied Math., 48:1073 (1988).

[0023] Preferred methods for determining identity are designed to give the greatest match between the sequences tested. Methods for determining identity and similarity are codified in publicly available computer programs. Preferred computer program methods for determining identity and similarity between two sequences include, for example, the GCG program package (Devereux, J. et al., Nucleic Acids Research 12(1): 387(1984)), BestFit, FASTA, BLASTN, and BLASTP (Altschul, SF et al., J. Mol. Biol. 215:403-410(1990)), EMBOSS Needle (Madeira, F. et al., Nucleic Acids Research 47(W1): W636-W641(2019)). BLAST programs are publicly available from NCBI and other sources (BLAST Manual, Altschul, S. et al., NCBI NLM NIH Bethesda, MD 20894; Altschul, S. et al., J. Mol. Biol. 215:403-410(1990)). EMBOSS programs are publicly available from EMBL-EBI. Identity can also be determined using the well-known Smith Waterman algorithm. The EMBOSS Needle program is a preferred program used.

[0024] Preferred parameters for polypeptide sequence comparison include: Algorithm: Needleman and Wunsch, J. Mol. Biol. 48(3):443-453 (1970); Comparison matrix: BLOSUM62 from Henikoff and Henikoff, Proc. Natl. Acad. Sci. USA. 89:10915-10919 (1992); Gap open penalty: 10; and Gap extension penalty: 0.5. A program useful with these parameters is publicly available as the EMBOSS Needle program from EMBL-EBI. The aforementioned parameters are the default parameters (no penalty for end gaps) for Global Pairwise Sequence alignment of proteins.

[0025] Preferred parameters for nucleic acid comparison include the following: Algorithm: Needleman and Wunsch, J. Mol. Biol. 48:443-453 (1970); Comparison matrix: DNAfull; Gap opening penalty: 10; Gap extension penalty: 0.5. A program useful with these parameters is publicly available as the EMBOSS Needle program from EMBL-EBI. The aforementioned parameters are the default parameters (no penalty for end gaps) for Global Pairwise Sequence alignment of nucleotide sequences.

[0026] As used herein, the terms "disorder" and "disease" are used interchangeably.

[0027] Also provided herein are embodiments in which any embodiment described herein may be combined with any one or more other embodiments, provided that the combinations are not mutually exclusive.

[0028] (Anti-MuSK antibody or antigen-binding fragment thereof) All antibodies or antigen-binding fragments thereof defined herein are encompassed as such by the present invention. The antibodies or antigen-binding fragments thereof are for use in the treatment of neuromuscular disorders in human subjects.

[0029] The present invention relates to anti-MuSK antibody-based molecules, including anti-MuSK antibodies, epitope-binding domains thereof, antigen-binding fragments thereof, and antibody derivatives, for treating neuromuscular diseases or conditions. In certain embodiments, the phrase "antibody-based molecules" may be replaced by the word "antibody" or by the phrase "antibody or functional fragment thereof" or by the phrase "antibody or antigen-binding fragment".

[0030] The term "anti-MuSK antibody" may be interchanged with the term "MuSK antibody."

[0031] Any anti-MuSK antibody-based molecule, including anti-MuSK antibodies, epitope-binding domains thereof, antigen-binding fragments thereof, and antibody derivatives, that can bind to muscle-specific tyrosine protein kinase (MuSK) are encompassed by the present invention. In some embodiments, such anti-MuSK antibodies can also activate MuSK signaling and / or phosphorylation. The present invention provides the insight that such antibody-based molecules are useful for treating conditions in which a subject requires increased MuSK signaling or MuSK phosphorylation, such as neuromuscular diseases or conditions. Thus, in a first aspect, an anti-MuSK antibody or antigen-binding fragment thereof is provided for use in treating a neuromuscular disorder in a human subject.

[0032] MuSK is a receptor tyrosine kinase expressed in skeletal muscle that plays a pivotal role in the formation and maintenance of neuromuscular synapses (Burden et al., "The Role of MuSK in Synapse Formation and Neuromuscular Disease," Cold Spring Harb. Perspect. Biol. 5:a009167 (2013), incorporated herein by reference in its entirety). MuSK is a 120 kDa single-spanning membrane protein composed of an extracellular region containing three Ig-like and a Frizzled (Fz)-like domain, and an intracellular region containing a juxtamembrane region, a kinase domain, and a short cytoplasmic tail (Jennings et al., "Muscle-Specific trk-Related Receptor with a Kringle Domain Defines a Distinct Class of Receptor Tyrosine Kinases," Proc. Natl. Acad. Sci. USA 90:2895-2899 (1993) and Valenzuela et al., "Receptor Tyrosine Kinase Specific for the Skeletal Muscle Lineage: Expression in Embryonic muscle, at the Neuromuscular Junction, and After Injury," which are incorporated by reference in their entireties. and After Injury," Neuron 15: 573-584 (1995)). MuSK phosphorylation is stimulated by agrin, a signal provided by motor neurons.Once activated, MuSK (1) clusters and anchors AchRs and additional muscle proteins important for synaptic transmission, (2) enhances transcription of genes encoding synaptic proteins in muscle "synaptic nuclei," and (3) stimulates pathways that promote presynaptic differentiation and the production of retrograde signals that promote attachment of motor nerve terminals to muscle. In the absence of MuSK, neuromuscular synapses do not form (Burden et al., "The Role of MuSK in Synapse Formation and Neuromuscular Disease," Cold Spring Harb. Perspect. Biol. 5:a009167 (2013), incorporated herein by reference in its entirety). In addition to its role in synapse formation, MuSK is also required for maintaining adult synapses, as inhibition of MuSK expression in adult muscle results in severe defects in pre- and postsynaptic differentiation (Kong et al., "Inhibition of Synapse Assembly in Mammalian Muscle in vivo by RNA Interference," EMBO Rep 5:183-188 (2004) and Hesser et al., "Synapse Disassembly and Formation of New Synapses in Postnatal Muscle Upon Conditional Inactivation of MuSK," Mol. Cell. Neurosci. 31:470-480 (2006), which are incorporated by reference in their entireties).Consistent with these findings in mice, mutations that impair MuSK kinase activity or inhibit signaling steps downstream from MuSK cause myasthenia gravis (CM), a condition characterized by structurally and functionally defective synapses, resulting in muscle weakness and fatigue (Beeson et al., "Dok-7 Mutations Underlie a Neuromuscular Junction Synaptopathy," Science 313:1975-1978 (2006); Muller et al., "Phenotypical Spectrum of DOK7 Mutations in Congenital Myasthenic Syndromes," Brain 130:1497-1506 (2007); and Selcen et al., "Compensatory Subpopulations of Motor Neurons in a Mouse Model of Amyotrophic Lateral Sclerosis," which are incorporated by reference in their entireties. "Compensatory Subpopulation of Motor Neurons in a Mouse Model of Amyotrophic Lateral Sclerosis", J. Comp. Neurol. 490:209-219 (2008)).

[0033] The amino acid sequence of human MuSK has the amino acid sequence of SEQ ID NO:129 below. [ka]

[0034] In accordance with the invention, the MuSK antibody-based molecules described herein bind to an epitope within the Frizzled (Fz)-like domain of the MuSK protein. The Fz-like domain of MuSK has the amino acid sequence of SEQ ID NO: 130, shown below. [ka]

[0035] The term "epitope" as used herein refers to an antigenic determinant capable of binding to an antibody. Epitopes usually comprise surface groups of molecules such as amino acids or sugar side chains, and usually have specific three-dimensional structural characteristics and specific charge characteristics. Conformational and non-conformational epitopes are distinguished in that the binding to the former, but not the latter, is lost in the presence of denaturing solvents. Epitopes can include amino acid residues that are directly involved in binding (also called the immunodominant components of the epitope) and other amino acid residues that are not directly involved in binding, e.g., amino acid residues that are effectively blocked by a specific antigen-binding peptide (in other words, the amino acid residues are within the region occupied by the specific antigen-binding peptide). An epitope typically includes at least 3, and more usually, at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more amino acids in a unique spatial conformation.

[0036] In certain embodiments, the MuSK antibody or antigen-binding fragment for use according to the invention binds to the MuSK Frizzled (Fz)-like domain. In certain embodiments, the MuSK antibody or antigen-binding fragment immunospecifically binds to an epitope within the MuSK Fz-like domain sequence of SEQ ID NO: 130 more frequently, rapidly, with longer duration, and / or with higher affinity or avidity than another epitope. In certain embodiments, the MuSK antibody-based molecules described herein immunospecifically bind to any 2, 3, 4, 5, 6, 7, 8, 9, 10, or more amino acid residues of SEQ ID NO: 130. The terms "affinity," "specific binding," "binding," "immunospecific binding," "binding activity," or "specific binding activity" as used herein refer to the degree to which an antibody or antibody fragment as defined herein binds to an epitope within the MuSK-Fz-like domain sequence of SEQ ID NO: 130.

[0037] In certain embodiments, the MuSK antibody-based molecules disclosed herein bind to the MuSK Fz-like domain with an affinity corresponding to a KD of about 10-7 M or less. For example, the MuSK antibody-based molecules disclosed herein bind to the MuSK Fz-like domain with an affinity corresponding to a KD of about 10-8 M, about 10-9 M, about 10-10 M, about 10-11 M, about 10-12 M, or less, as determined, for example, by surface plasmon resonance (SPR) techniques on a Biacore 3000 instrument (preferably using the antibody as the ligand and MuSK as the analyte). The MuSK antibody-based molecules disclosed herein bind to the MuSK Fz-like domain with an affinity that corresponds to a KD that is at least 10-fold lower, e.g., at least 100-fold lower, e.g., at least 1,000-fold lower, e.g., at least 10,000-fold lower, e.g., at least 100,000-fold lower than its affinity for binding to a non-specific antigen (e.g., bovine serum albumin, casein, etc.). The amount by which the affinity is lowered depends on the KD of the antibody, so that if the KD of the antibody is very low (i.e., the antibody is highly specific), the amount by which the affinity for the antigen is lower than the affinity for a non-specific antigen can be at least 10,000-fold. As used herein, the term "kd" (sec-1 or 1 / s) refers to the dissociation rate constant of a particular antibody-antigen interaction. This value is also referred to as the koff value. As used herein, the term "ka" (M-1 x sec-1 or 1 / M) refers to the association rate constant of a particular antibody-antigen interaction. As used herein, the term "KD" (M) refers to the dissociation equilibrium constant of a particular antibody-antigen interaction and is obtained by dividing kd by ka. As used herein, the term "KA" (M-1 or 1 / M) refers to the association equilibrium constant of a particular antibody-antigen interaction and is obtained by dividing ka by kd.

[0038] In certain embodiments, the MuSK antibody-based molecules described herein have a pH-dependent binding affinity to MuSK that allows for antibody recycling to enhance antigen binding. For example, in certain embodiments, the association or dissociation rate constants may be different under acidic versus neutral versus basic pH conditions. In certain embodiments, the MuSK antibody-based molecules described herein have a higher dissociation rate constant under acidic pH conditions, e.g., a pH of <7.0, compared to neutral pH conditions, e.g., a pH of ~7.0-7.9. In some embodiments, the MuSK antibody-based molecules described herein have a 2- to 3-fold higher dissociation rate constant under acidic pH conditions (e.g., pH ~5.5) compared to neutral pH (pH ~7.4). In certain embodiments, the MuSK antibody-based molecules bind to the MuSK Fz-like domain with a higher affinity under neutral pH conditions than under acidic pH conditions. In other words, in certain embodiments, the MuSK antibody-based molecules bind to the MuSK Fz-like domain with a higher dissociation rate under acidic pH conditions than under neutral pH conditions. Neutral pH conditions can be defined as a pH comprised between 7.0 and 7.9. Acidic pH conditions can be defined as a pH less than 7.0. Higher can mean at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300% higher. Antibodies with this pH-dependent dissociation property dissociate from antigen after binding and activation, but before lysosomal degradation. Once dissociated, the antibody is released back into circulation via the neonatal Fc receptor to bind more antigen.

[0039] In some embodiments, MuSK signaling is activated by binding of the MuSK antibodies of the present invention to their respective epitopes in the Fz-like domain. In particular, when the MuSK antibodies of the present invention bind to their respective epitopes in the MuSK Fz-like domain, this binding induces phosphorylation and activation of MuSK. The MuSK antibodies of the present invention induce about 50% to about 100% MuSK phosphorylation compared to MuSK phosphorylation induced by agrin activation (e.g., as measured by C2C12 phosphorylation assay). In some embodiments, the MuSK antibodies of the present invention induce about 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%95% MuSK phosphorylation (compared to MuSK phosphorylation induced by agrin activation). In certain embodiments, the MuSK antibody-based molecules of the invention induce about 90% to about 100% MuSK phosphorylation (compared to MuSK phosphorylation induced by agrin activation) upon MuSK binding. MuSK phosphorylation can be assessed using techniques known to those skilled in the art, such as Western blotting or C2C12 myotube phosphorylation assays. Such antibodies that activate MuSK signaling (i.e., induce MuSK dimerization, induce MuSK tyrosine phosphorylation) are agonistic antibodies.

[0040] In some embodiments, the MuSK antibodies of the invention, i.e., MuSK antibodies that bind to the Fz domain of MuSK, do not interfere with (i.e., do not block, interfere, inhibit, or reduce) natural ligand binding and stimulation of MuSK. In some embodiments, the MuSK antibodies costimulate MuSK activation with its natural ligand, i.e., agrin, resulting in an additive effect of activation, e.g., MuSK phosphorylation. Thus, in some embodiments, the MuSK antibodies of the invention enhance natural MuSK activation, i.e., phosphorylation, induced by natural ligand binding. Such MuSK antibodies are agonistic antibodies. In some embodiments, the antibodies of the invention, in combination with the natural ligand, activate MuSK (i.e., MuSK phosphorylation) to >100% of endogenous activation levels, e.g., at least 110%, 130%, 150%, 200% of endogenous activation levels.

[0041] Thus, in certain embodiments, the activity of the MuSK antibody-based molecules of the invention includes: (i) binding to an epitope of human muscle-specific tyrosine protein kinase (MuSK), which epitope is present in the MuSK Frizzled (Fz)-like domain sequence of SEQ ID NO: 130 (wherein the antibody-based molecule, upon binding to the epitope, induces MuSK phosphorylation), and / or (ii) binding to the MuSK Fz-like domain does not block, interfere with, or inhibit native or endogenous MuSK ligand-induced phosphorylation, but can enhance native or endogenous MuSK ligand-induced phosphorylation, and (iii) binding to the MuSK Fz-like domain occurs with greater affinity at neutral pH conditions than at acidic pH conditions.

[0042] All of these features are further defined herein.

[0043] Antibody-based molecules include, but are not limited to, antibodies, complete antibodies, epitope-binding fragments of whole antibodies, antigen-binding fragments of whole antibodies, and antibody derivatives. An epitope-binding fragment of an antibody can be obtained by actual fragmentation of a parent antibody (e.g., Fab or (Fab)2 fragments). Alternatively, an epitope-binding fragment is an amino acid sequence that comprises a portion of the amino acid sequence of such a parent antibody. As used herein, a molecule is said to be a "derivative" of an antibody (or a relevant portion thereof) if it is obtained by actual chemical modification of the parent antibody or a portion thereof, or if it comprises an amino acid sequence that is substantially similar to the amino acid sequence of such a parent antibody or a relevant portion thereof (e.g., differs from such parent antibody or such relevant portion thereof by less than 30%, less than 20%, less than 10%, less than 5%, or less than 10 amino acid residues, or less than 10, 9, 8, 7, 6, 5, 4, 3, or 2 amino acid residues, from such parent antibody or such relevant portion thereof).

[0044] In certain embodiments, the antibody-based molecules of the invention are intact immunoglobulins or molecules with epitope-binding domains encoding such fragments in recombinant cells (see, e.g., Evans et al., "Rapid Expression Of An Anti-Human C5 Chimeric Fab Utilizing A Vector That Replicates In COS And 293 Cells," J. Immunol. Meth. 184:123-38 (1995), incorporated herein by reference in its entirety). For example, a chimeric gene encoding a portion of the F(ab')2 fragment can include DNA sequences encoding the CH1 domain and hinge region of the heavy chain followed by a translation stop codon to obtain such a truncated antibody fragment molecule. Suitable fragments capable of binding the desired epitope can be readily screened for utility in the same manner as intact antibodies.

[0045] Antibody derivatives include molecules that contain at least one epitope-binding domain of an antibody and are usually formed using recombinant techniques. One exemplary antibody derivative includes single-chain Fv (scFv). scFv is formed from the two domains of the Fv fragment, VL and VH, which may be encoded by separate genes. Such gene sequences or their encoding cDNAs are linked by a flexible linker (usually about 10, 12, 15 or more amino acid residues) that allows them to be engineered into a single protein chain in which VL and VH associate to form monovalent epitope-binding molecules using recombinant techniques (see, for example, Bird et al., "Single-Chain Antigen-Binding Proteins," Science 242:423-426 (1988); and Huston et al., "Protein Engineering Of Antibody Binding Sites: Recovery Of Specific Activity In An Anti-Digoxin Single-Chain Fv Analogue Produced In Escherichia coli," Proc. Natl. Acad. Sci. (USA), incorporated herein by reference in their entireties. 85:5879-5883 (1988)). Alternatively, bispecific antibodies with binding specificities for two different epitopes can be formed by utilizing a flexible linker that is not too short (e.g., not less than about 9 residues) such that the VL and VH of different single polypeptide chains can associate together.

[0046] In another embodiment, the antibody derivative is a divalent or bivalent single chain variable fragment engineered by linking two scFvs in tandem (i.e., tandem scFvs) or by dimerization to form a diabody (Holliger et al., "'Diabodies': Small Bivalent And Bispecific Antibody Fragments," Proc. Natl. Acad. Sci. (USA) 90(14), 6444-8 (1993), incorporated herein by reference in its entirety). In yet another embodiment, the antibody is a triabody, i.e., a trivalent single chain variable fragment engineered by linking three scFvs in tandem or by trimerization to form a triabody. In another embodiment, the antibody is a tetrabody of four single chain variable fragments. In another embodiment, the antibody is a "linear antibody," which is an antibody that comprises a pair of tandem Fd segments (VH-CH1-VH-CH1) that form a pair of antigen-binding regions (see Zapata et al., Protein Eng. 8(10):1057-1062 (1995), which is incorporated herein by reference in its entirety). In another embodiment, the antibody derivative is a minibody consisting of a single chain Fv region attached to a CH3 region (i.e., scFv-CH3).

[0047] These and other useful antibody fragments and derivatives in the context of the present invention are discussed further herein. The term antibody-based molecule should also be understood to include, unless otherwise specified, antibody-like polypeptides, such as chimeric antibodies and humanized antibodies, as well as antigen-binding fragments and antibody fragments (epitope-binding fragments, antigen-binding fragments, or functional fragments) that retain the ability to specifically bind to an antigen, provided by any known technique, such as enzymatic cleavage, peptide synthesis, and recombinant techniques.

[0048] The antibodies produced herein may be of any isotype. As used herein, "isotype" refers to the immunoglobulin class (e.g., IgG1, IgG2, IgG3, IgG4, IgD, IgA, IgE, or IgM) encoded by heavy chain constant region genes. The choice of isotype is usually guided by the desired effector function, such as antibody-dependent cellular cytotoxicity (ADCC) induction. Exemplary isotypes are IgG1, IgG2, IgG3, and IgG4. Particularly useful isotypes of the MuSK antibodies disclosed herein include IgG1 and IgG2.

[0049] Either the human light chain constant region, kappa or lambda, can be used. If desired, the class of the MuSK antibody of the present invention can be switched by known methods. For example, an antibody of the present invention that is originally an IgM can be class switched to an IgG antibody of the present invention. Furthermore, class switching techniques can be used to convert one IgG subclass to another, for example, from IgG1 to IgG2. Thus, the effector function of the antibody of the present invention can be altered by isotype switching, for example, to an IgG1, IgG2, IgG3, IgG4, IgD, IgA, IgE, or IgM antibody, for various therapeutic applications.

[0050] In certain embodiments, one, two or more amino acid substitutions are introduced into the IgG constant region Fc region to alter the effector functions of the antibody-based molecule. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 238, 239, 243, 265, 267, 268, 292, 297, 300, 318, 320, 322, 327, 328, 329, 330, 331, 332, and 396, numbered according to the EU numbering system (https: / / www.imgt.org / IMGTScientificChart / Numbering / Hu_IGHGnber.html#notes and Edelman, GM et al., Proc. Natl. Acad. USA, 63, 78-85(1969). PMID: 5257969), can be replaced with a different amino acid residue such that the antibody-based molecule has an altered affinity for an effector ligand but retains antigen binding ability. In some embodiments, amino acid 234 or 235 is replaced. In another embodiment, amino acids 234 and 235 are replaced. In this context, preferred amino acid sequences of human IgG constant Fc regions include SEQ ID NO: 266 or 267. In this context, for example, amino acids 234 and 235 numbered according to the EU numbering system correspond to amino acids 7 and 8 of SEQ ID NO: 266 and 267 (i.e., human IgG constant Fc regions of the antibody-based molecules disclosed herein), or amino acids 234 and 235 numbered according to the EU numbering system correspond to amino acids 238 and 239 of SEQ ID NO: 268 and 270 (i.e., human full-length heavy chains of the antibody-based molecules disclosed herein). Since the variable regions are of different lengths, the positions are usually different, which introduces a "delta" between the numberings. In the cases shown above, delta is 4.Thus, when identifying the corresponding positions in SEQ ID NO: 266 or 267 or 268 or 270, the same is true for the other amino acid positions identified above numbered according to the EU numbering system (i.e., 236, 237, 238, 239, 243, 265, 267, 268, 292, 297, 300, 318, 320, 322, 327, 328, 329, 330, 331, 332, and 396). In the present application as filed, amino acid positions may also be referenced using the EU numbering system or using their actual position in a given Fc region (e.g., SEQ ID NO: 266 or 267) or full length heavy chain (e.g., SEQ ID NO: 268 or 270).

[0051] Thus, in certain embodiments, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17 amino acid substitutions are introduced into SEQ ID NO: 266 or 267. In certain embodiments, 1, 2, 3, 4 amino acid substitutions are introduced into SEQ ID NO: 266 or 267. In certain embodiments, 1 or 2 amino acid substitutions are introduced into SEQ ID NO: 266 or 267. Thus, in some embodiments, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 or 17 amino acid substitutions are introduced into SEQ ID NO: 266 or 267, and the substitutions are introduced into the amino acid positions selected from amino acid residues 234, 235, 236, 237, 239, 243, 267, 292, 297, 300, 318, 320, 322, 328, 330, 332 and 396 numbered according to the EU numbering system of said sequence. In some embodiments, 1 or 2 amino acid substitutions are introduced into SEQ ID NO: 266 or 267. In some embodiments, amino acid 234 or 235 numbered according to the EU numbering system of SEQ ID NO: 266 or 267 is replaced. In another embodiment, amino acid 234 and 235 numbered according to the EU numbering system of SEQ ID NO: 266 or 267 are replaced.

[0052] The effector ligand for which affinity is modified can be, for example, an Fc receptor or the C1 component of complement. This approach is described in further detail in U.S. Patent Nos. 5,624,821 and 5,648,260, each of which is fully incorporated herein by reference. In certain embodiments, one or more amino acid substitutions can be introduced into the Fc region of the antibody-based molecules described herein to remove potential glycosylation sites on the Fc region that may reduce Fc receptor binding (see, for example, Shields RL et al. (2001) J Biol Chem 276: 6591-604, which is incorporated herein by reference in its entirety). In certain embodiments, binding to an effector ligand is reduced by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or is no longer detectable compared to binding to the same ligand by an antibody that does not have any amino acid substitutions in its human IgG constant Fc region.

[0053] In a first embodiment, the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions have been introduced into the constant region of an antibody-based molecule described herein: G236R substitution; G236K substitution; G237A substitution; P238A substitution; F243L substitution; D265A substitution; S267E substitution; H268A substitution; R292P substitution; Y300L substitution; K322A substitution; K322Q substitution; A327Q substitution; L328F substitution; L328R substitution; P329A substitution; P329G substitution; A330L substitution; A330S substitution; P331S substitution; I332E substitution; or P396L substitution.

[0054] In a second embodiment, the following mutations (all numbered according to the EU numbering system): L234A and / or L235A substitutions; L234A and L235A substitutions; L234A, L235A, and P329G substitutions; L234A, L235A, and G236K substitutions; L234A, L235A, and G236E substitutions; L234A, L235A, and G236R substitutions; L234A and G236R substitutions; L234A, L235S, and G236R substitutions; L234A, L235T, and G236R substitutions; L234D, L235H, and G236R substitutions; L234D, L235K, and G236R substitutions; L234D and G236R substitutions; L234D, L235Q, and G236R substitutions; L234D, L235S, and G236R substitutions; L234E, L235D, and G236R substitutions; L234E, L235H, and G236R substitutions; L234E, L235I, and G236R substitutions; L234G, L235H, and G236R substitutions; L234G, L235Q, and G236R substitutions; L234G, L235S, and G236R substitutions; L234H, L235I, and G236R substitutions; L234H, L235S, and G236R substitutions; L234K, L235Q, and G236R substitutions; L234K, L235R, and G236R substitutions; L234K, L235S, and G236R substitutions; L234K, L235T, and G236R substitutions; L234K, L235V, and G236R substitutions; L234Q, L235A, and G236R substitutions; L234Q, L235D, and G236R substitutions; L234Q, L235H, and G236R substitutions; L234Q and G236R substitutions; L234Q, L235Q, and G236R substitutions; L234Q, L235R, and G236R substitutions; L234Q, L235S, and G236R substitutions; L234Q, L235T, and G236R substitutions; L234Q, L235V, and G236R substitutions; L234R, L235D, and G236R substitutions; L234R, L235E, and G236R substitutions; L234R, L235H, and G236R substitutions; L234R, L235I, and G236R substitutions; L234R, L235K, and G236R substitutions; L234R and G236R substitutions; L234R, L235Q, and G236R substitutions;L234R, L235R, and G236R substitutions; L234R, L235T, and G236R substitutions; L234S, L235E, and G236R substitutions; L234S, L235G, and G236R substitutions; L234S, L235H, and G236R substitutions; L234S, L235I, and G236R substitutions; L234S and G236R substitutions; L234S, L235R, and G236R substitutions; L234S, L235T, and G236R substitutions; L234S, L235V, and G236R substitutions; L234T, L235A, and G236R substitutions; L234T, L235D, and G236R, L234T, L235H, and G236R substitutions; L234T, L235I, and G236R substitutions; L234T, L235K, and G236R substitutions; L234T, L235Q, and G236R substitutions; L234T, L235R, and G236R substitutions; L234T, L235S, and G236R substitutions; L234T, L235T, and G236R substitutions; L234T, L235V, and G236R substitutions; G236R and an L328R substitution; L234A, L235A, G237A, P238S, H268A, A330S, and P331S substitutions; One or more of the following substitutions have been introduced into the constant region of the antibody-based molecules described herein: E233P, L234V, L235A, G326 deletion, A327G, A330S, and P331S substitutions; L235A and G236R substitutions; L235S and G236R substitutions;

[0055] In a third embodiment, the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions have been introduced into the Fc region (SEQ ID NO: 266 or SEQ ID NO: 267) of an antibody-based molecule described herein: G236R substitution; G236K substitution; G237A substitution; P238A substitution; F243L substitution; D265A substitution; S267E substitution; H268A substitution; R292P substitution; Y300L substitution; K322A substitution; K322Q substitution; A327Q substitution; L328F substitution; L328R substitution; P329A substitution; P329G substitution; A330L substitution; A330S substitution; P331S substitution; I332E substitution; or P396L substitution.

[0056] In a fourth embodiment, the following mutations (all numbered according to the EU numbering system): L234A and / or L235A substitutions; L234A and L235A substitutions; L234A, L235A, and P329G substitutions; L234A, L235A, and G236K substitutions; L234A, L235A, and G236E substitutions; L234A, L235A, and G236R substitutions; L234A and G236R substitutions; L234A, L235S, and G236R substitutions; L234A, L235T, and G236R substitutions; L234D, L235H, and G236R substitutions; L234D, L235K, and G236R substitutions; L234D and G236R substitutions; L234D, L235Q, and G236R substitutions; L234D, L235S, and G236R substitutions; L234E, L235D, and G236R substitutions; L234E, L235H, and G236R substitutions; L234E, L235I, and G236R substitutions; L234G, L235H, and G236R substitutions; L234G, L235Q, and G236R substitutions; L234G, L235S, and G236R substitutions; L234H, L235I, and G236R substitutions; L234H, L235S, and G236R substitutions; L234K, L235Q, and G236R substitutions; L234K, L235R, and G236R substitutions; L234K, L235S, and G236R substitutions; L234K, L235T, and G236R substitutions; L234K, L235V, and G236R substitutions; L234Q, L235A, and G236R substitutions; L234Q, L235D, and G236R substitutions; L234Q, L235H, and G236R substitutions; L234Q and G236R substitutions; L234Q, L235Q, and G236R substitutions; L234Q, L235R, and G236R substitutions; L234Q, L235S, and G236R substitutions; L234Q, L235T, and G236R substitutions; L234Q, L235V, and G236R substitutions; L234R, L235D, and G236R substitutions; L234R, L235E, and G236R substitutions; L234R, L235H, and G236R substitutions; L234R, L235I, and G236R substitutions; L234R, L235K, and G236R substitutions; L234R and G236R substitutions; L234R, L235Q, and G236R substitutions;L234R, L235R, and G236R substitutions; L234R, L235T, and G236R substitutions; L234S, L235E, and G236R substitutions; L234S, L235G, and G236R substitutions; L234S, L235H, and G236R substitutions; L234S, L235I, and G236R substitutions; L234S and G236R substitutions; L234S, L235R, and G236R substitutions; L234S, L235T, and G236R substitutions; L234S, L235V, and G236R substitutions; L234T, L235A, and G236R substitutions; L234T, L235D, and G236R, L234T, L235H, and G236R substitutions; L234T, L235I, and G236R substitutions; L234T, L235K, and G236R substitutions; L234T, L235Q, and G236R substitutions; L234T, L235R, and G236R substitutions; L234T, L235S, and G236R substitutions; L234T, L235T, and G236R substitutions; L234T, L235V, and G236R substitutions; G236R and L328R substitutions; L234A, L235A, G237A, P238S, H268A, A330S, and P331S substitutions; One or more of the following substitutions have been introduced into the Fc region of the antibody-based molecules described herein (SEQ ID NO: 266 or SEQ ID NO: 267): E233P, L234V, L235A, G326 deletion, A327G, A330S, and P331S substitutions; L235A and G236R substitutions; L235S and G236R substitutions.

[0057] In certain embodiments, one or more of the following mutations (all numbered according to the EU numbering system): L234A and / or L235A substitutions are introduced into the Fc region of an antibody-based molecule described herein (SEQ ID NO:266 or SEQ ID NO:267). In certain embodiments, the following mutations: L234A and L235A numbered according to the EU numbering system are introduced into the Fc region of an antibody-based molecule described herein (SEQ ID NO:266 or SEQ ID NO:267). This embodiment results in an antibody-based molecule having a heavy chain represented by SEQ ID NO:268 or 270.

[0058] Such antibodies with altered, reduced or even abolished effector functions are attractive in the context of the present invention.

[0059] In one embodiment, - an agonist MuSK antibody, and / or -has reduced or absent effector function, Anti-MuSK antibodies or antigen-binding fragments thereof are provided.

[0060] The antibody or antigen-binding fragment thereof is preferably for use in treating a neuromuscular disorder in a human subject.

[0061] In one embodiment, - binds to the MuSK Frizzled (Fz)-like domain sequence of SEQ ID NO: 129, - an agonist MuSK antibody, and -has reduced or absent effector function, Anti-MuSK antibodies or antigen-binding fragments thereof are provided.

[0062] The antibody or antigen-binding fragment thereof is preferably for use in treating a neuromuscular disorder in a human subject.

[0063] Reduced or eliminated effector functions can be obtained by introducing mutations into the human IgG constant Fc region as previously described herein. Preferably, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17 amino acid substitutions are introduced into the Fc region. Preferably, at least 1, 2, 3, 4 amino acid substitutions are introduced into the Fc region.

[0064] The Fc region can comprise SEQ ID NO: 266 or 267, wherein the substitutions are introduced at amino acid positions selected from amino acid residues 234, 235, 236, 237, 238, 239, 243, 265, 267, 268, 292, 297, 300, 318, 320, 322, 327, 328, 329, 330, 331, 332, and 396, numbered according to the EU numbering system of said sequences.

[0065] In certain embodiments, the Fc region may comprise SEQ ID NO: 266 or 267, and the substitution is introduced at an amino acid position selected from amino acid residue 234 or 235 of said sequence, numbered according to the EU numbering system.

[0066] In certain embodiments, the Fc region may comprise SEQ ID NO: 266 or 267, and the substitution is introduced at an amino acid position selected from amino acid residues 234 and 235, numbered according to the EU numbering system of said sequence.

[0067] In certain embodiments, the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions are introduced into the human IgG constant Fc region (SEQ ID NO: 266 or SEQ ID NO: 267) of the antibody-based molecules described herein: G236R substitution; G236K substitution; G237A substitution; P238A substitution; F243L substitution; D265A substitution; S267E substitution; H268A substitution; R292P substitution; Y300L substitution; K322A substitution; K322Q substitution; A327Q substitution; L328F substitution; L328R substitution; P329A substitution; P329G substitution; A330L substitution; A330S substitution; P331S substitution; I332E substitution; or P396L substitution.

[0068] In one embodiment, each of the combinations of mutations previously described in the fourth embodiment of the present application in the human IgG constant Fc region of the antibody-based molecules described herein can be made.

[0069] In a preferred embodiment, the L234A or L235A substitution is introduced into the human IgG constant Fc region of the antibody-based molecules described herein. In a more preferred embodiment, the L234A and L235A substitutions are introduced into the human IgG constant Fc region of the antibody-based molecules described herein. This embodiment results in an antibody-based molecule having a heavy chain represented by SEQ ID NO: 268 or 270.

[0070] In an even more preferred embodiment, the anti-MuSK antibody or antigen-binding fragment thereof comprises: a) a heavy chain variable domain (VH) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234; and b) a light chain variable domain (VL) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235 Contains :.

[0071] In this context, identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%.

[0072] Preferred anti-MuSK antibodies or antigen-binding fragments thereof include a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains :.

[0073] Preferred anti-MuSK antibodies or antigen-binding fragments thereof include c) a full-length heavy chain comprising SEQ ID NO: 270, and d) a full length light chain comprising SEQ ID NO: 271 Contains :.

[0074] In certain embodiments, the antibody-based molecules of the invention are "humanized", especially when they are to be utilized for therapeutic purposes. The term "humanized" refers to chimeric molecules, usually prepared using recombinant techniques, having an antigen-binding site derived from an immunoglobulin from a non-human species and the remaining immunoglobulin structure based on the structure and / or sequence of a human immunoglobulin. The antigen-binding site can comprise either a complete non-human antibody variable domain fused to a human constant domain, or only the complementarity determining regions (CDRs) of such a variable domain grafted into appropriate human framework regions of the human variable domain. The framework residues of such humanized molecules may be wild-type (e.g., fully human) or they may be modified to contain one or more amino acid substitutions not found in the human antibody whose sequence serves as the basis for humanization. Although humanization reduces or eliminates the possibility that the constant regions of the molecule may act as immunogens in human individuals, the possibility of an immune response to the foreign variable regions remains (LoBuglio, AF et al., Mouse / Human Chimeric Monoclonal Antibody In Man: Kinetics And Immune Response, Proc. Natl. Acad. Sci. 2003, 144:1311-1323, which is incorporated herein by reference in its entirety).USA 86:4220-4224(1989)). Another approach focuses not only on providing constant regions of human origin, but also on reshaping the variable regions to be as close as possible to the human form. The variable regions of both the heavy and light chains contain three complementarity determining regions (CDRs) that vary in response to the antigen in question and determine the binding capacity. The CDRs are flanked by four framework regions (FRs) that are relatively conserved in a given species and are presumed to provide a scaffold for the CDRs. When a non-human antibody is prepared for a particular antigen, the variable region can be "reshaped" or "humanized" by grafting the CDRs from the non-human antibody into the FRs present in the human antibody to be modified. Suitable methods for humanizing the non-human antibodies described herein are known in the art, e.g., see Sato, K. et al., Cancer Res 53:851-856 (1993); Riechmann, L. et al., Reshaping Human Antibodies for Therapy, Nature 332:323-327 (1988); Verhoeyen, M. et al., Reshaping Human Antibodies: Grafting An Antilysozyme Activity, Science 239:1534-1536 (1988); Kettleborough, CA et al., Humanization Of A Mouse Monoclonal Antibody By CDR-Grafting: The Importance Of Framework Residues On Loop Conformation, all of which are incorporated by reference in their entireties. Protein Engineering 4:773-3783(1991); Maeda, H.See, for example, Gorman, SD et al., "Construction Of Reshaped Human Antibodies With HIV-Neutralizing Activity," Human Antibodies Hybridoma 2:124-134 (1991); Gorman, SD et al., "Reshaping A Therapeutic CD4 Antibody," Proc. Natl. Acad. Sci. USA 88:4181-4185 (1991); Tempest, PR et al., "Reshaping A Human Monoclonal Antibody To Inhibit Human Respiratory Syncytial Virus Infection In Vivo," Bio / Technology 9:266-271 (1991); Co, MS et al., "Humanized Antibodies For Antiviral Therapy," , "Humanization Of An Anti-p185her2 Antibody For Human Cancer Therapy," Proc. Natl. Acad. Sci. USA 88:2869-2873 (1991); Carter, P. et al., "Humanization Of An Anti-p185her2 Antibody For Human Cancer Therapy," Proc. Natl. Acad. Sci. USA 89:4285-4289 (1992); and Co, MS et al., "Chimeric And Humanized Antibodies With Specificity For The CD33 Antigen," J. Immunol.148:1149-1154 (1992). In some embodiments, the humanized MuSK antibodies of the invention retain all CDR sequences (e.g., a humanized antibody containing all six CDRs from a llama or mouse antibody). In other embodiments, the humanized MuSK antibodies of the invention have one or more CDRs (one, two, three, four, five, six) that are altered relative to the original antibody. Methods of humanizing antibodies are well known in the art and are suitable for humanizing the antibodies disclosed herein (see, e.g., U.S. Patent No. 5,225,539 to Winter; U.S. Patent Nos. 5,530,101 and 5,585,089 to Queen and Selick; U.S. Patent No. 5,859,205 to Robert et al.; U.S. Patent No. 6,407,213 to Carter; and U.S. Patent No. 6,881,557 to Foote, which are incorporated herein by reference in their entireties).

[0075] In some antibodies, only a portion of the CDRs, i.e., the subset of CDR residues necessary for binding, called "specificity determining residues" ("SDRs"), are required to retain antibody binding. CDR residues that do not contact antigen and are not in the SDRs may be selected from regions of the Kabat CDRs that are outside of the Chothia hypervariable loops, based on prior work (see Kabat et al., SEQUENCES OF PROTEINS OF IMMUNOLOGICAL INTEREST, National Institutes of Health Publication No. 91-3242 (1992); Chothia, C. et al., Canonical Structures For The Hypervariable Regions Of Immunoglobulins, J. Mol. Biol. 196:901-917 (1987)), by molecular modeling and / or empirically, or from the methods described in Gonzales, NR et al., SDR Grafting Of A Murine Antibody Using Multiple Human Germline Templates to Minimize Its Immunogenicity, which are incorporated herein by reference in their entireties. Humanized antibodies can be identified as described in "Humanized Antibody Templates To Minimize Its Immunogenicity," Mol. Immunol. 41:863-872 (2004). In such humanized antibodies, at positions where one or more donor CDR residues are absent or where the entire donor CDR is deleted, the amino acid residue occupying that position can be the amino acid residue occupying the corresponding position (according to Kabat numbering) in the acceptor antibody sequence. The number of such acceptor and donor amino acid substitutions in the CDRs to be included reflects a balance of competing considerations. Such substitutions are potentially advantageous in that they reduce the number of non-human amino acids in the humanized antibody and, as a result, reduce potential immunogenicity.However, the substitution may cause a change in affinity, and a significant decrease in affinity is preferably avoided. The substitution may cause a change in activity. Such a substitution that causes a significant decrease in activity is preferably avoided. In this context, the antibody or antibody fragment should still exhibit detectable activity of the antibody as defined herein above, or at least some activity of the antibody. The position of the substitution within the CDR and the amino acid to be substituted can also be selected empirically.

[0076] Phage display technology can alternatively be used to increase (or decrease) the CDR affinity of the antibody-based molecules of the present invention. This technology, called affinity maturation, utilizes mutagenesis or "CDR walking" and reselection with the target antigen or its antigenic fragment to identify antibodies with CDRs that bind with higher (or lower) affinity to the antigen compared to the initial or parent antibody (see, for example, Glaser et al., "Antibody Engineering By Codon-Based Mutagenesis In A Filamentous Phage Vector System," J. Immunology 149:3903-3913 (1992), which is incorporated herein by reference in its entirety). Mutating entire codons, rather than single nucleotides, results in a semi-randomized repertoire of amino acid mutations. Libraries consist of a pool of mutant clones, each of which differs from other members of such a library by a single amino acid alteration in a single CDR, and which contain mutants potentially representing every possible amino acid substitution for each CDR residue. Mutants with increased (or decreased) binding affinity for the antigen can be screened by contacting the immobilized mutants with labeled antigen.Any screening method known in the art can be used to identify mutant antibody-based binding molecules with increased or decreased affinity for the antigen (e.g., ELISA) (Wu, H. et al., Stepwise In Vitro Affinity Maturation Of Vitaxin, An Alphav Beta3-Specific Humanized mAb, Proc. Natl. Acad. Sci. USA 95:6037-6042 (1998); Yelton et al., Affinity Maturation Of The BR96 Anti-Carcinoma Antibody By Codon-Based Mutagenesis, J. Immunology 155:1994 (1995), which are incorporated by reference in their entireties). CDR walking, which randomizes the light chain, may also be used (see Schier, R. et al., "Isolation Of Picomolar Affinity Anti-c-erbB-2 Single-Chain Fv By Molecular Evolution Of The Complementarity Determining Regions In The Center Of The Antibody Binding Site," J. Mol. Biol. 263:551-567 (1996), which is incorporated herein by reference in its entirety).

[0077] Methods for affinity maturation of MuSK antibody molecules are described herein, e.g., Krause, JC et al., An Insertion Mutation That Distorts Antibody Binding Site Architecture Enhances Function of a Human Antibody, MBio. 2(1): e00345-10 (2011); Kuan, CT et al., Affinity-Matured Anti-Glycoprotein NMB Recombinant Immunotoxins Targeting Malignant Gliomas And Melanomas, Int. J. Cancer 10.1002 / ijc.25645 (2010); Hackel, BJ et al., Stability And CDR Composition Biases Enrich Binder Functionality Landscape, all of which are incorporated by reference herein in their entireties. "HIV-1 Landscapes," J. Mol. Biol. 401(1):84-96(2010); Montgomery, DL et al., "Affinity Maturation And Characterization Of A Human Monoclonal Antibody Against HIV-1 gp41," MAbs 1(5):462-474(2009); Gustchina, E.Finlay, WJ et al., "Affinity Maturation By Targeted Diversification Of The CDR-H2 Loop Of A Monoclonal Fab Derived From A Synthetic Naive Human Antibody Library And Directed Against The Internal Trimeric Coiled-Coil Of Gp41 Yields A Set Of Fabs With Improved HIV-1 Neutralization Potency And Breadth," Virology 393(1):112-119 (2009); Finlay, WJ et al., "Affinity Maturation Of A Humanized Rat Antibody For Anti-RAGE Therapy: Comprehensive Mutagenesis Demonstrates High Levels of Mutational Plasticity Both Within and Outside the Complementarity Determining Regions," Mutagenesis Reveals A High Level Of Mutational Plasticity Both Inside And Outside The Complementarity-Determining Regions," J. Mol. Biol. 388(3):541-558(2009); Bostrom, J. et al., "Improving Antibody Binding Affinity And Specificity For Therapeutic Development," Methods Mol. Biol. 525:353-376(2009); Steidl, S.et al., "In Vitro Affinity Maturation Of Human GM-CSF Antibodies By Targeted CDR-Diversification," Mol. Immunol. 46(1):135-144 (2008); and Barderas, R. et al., "Affinity Maturation Of Antibodies Assisted By In Silico Modeling," Proc. Natl. Acad. Sci. USA 105(26):9029-9034 (2008).

[0078] In the context of this application, an amino acid alteration (change or modification) can be an amino acid substitution, addition, deletion, or chemical modification.

[0079] In certain embodiments, the MuSK antibody-based molecules described herein comprise the amino acid sequences of any one, any two, any three, any four, any five, or any six of the CDRs as provided in Tables 1 and 2 herein.

[0080] In one embodiment, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: (i) a complementarity determining region 1 (CDR-H1) comprising any one of the amino acid sequences of SEQ ID NOs: 1 to 16, 135, 136, 147 to 149, or any one of the modified amino acid sequences of SEQ ID NOs: 1 to 16, 135, 136, or 147 to 149 (the modified sequence has 1, 2, 3, 4, or 5 amino acid alterations compared to SEQ ID NOs: 1 to 16, 135, 136, or 147 to 149); (ii) a complementarity determining region 2 (CDR-H2) comprising any one of SEQ ID NOs: 17 to 32, 137, 138, 150 to 155, or any one of SEQ ID NOs: 17 to 32, 137, 138, or 150 to 155, a modified amino acid sequence (the modified sequence having 1, 2, 3, 4, or 5 amino acid alterations compared to SEQ ID NOs: 17 to 32, 137, 138, or 150 to 155); and (iii) Complementarity determining region 3 (CDR-H3) comprising any one of SEQ ID NOs: 33 to 48, 139, 140, 156 to 158, and 240 to 251, or any one of SEQ ID NOs: 33 to 48, 139, 140, 156 to 158, and 240 to 251 (the modified sequence has 1, 2, 3, 4, or 5 amino acid alterations compared to SEQ ID NOs: 33 to 48, 139, 140, 156 to 158, and 240 to 251). Contains :.

[0081] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain comprising: (i) a CDR-H1 of SEQ ID NO: 1 or a CDR-H1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 1, a CDR-H2 of SEQ ID NO: 17 or a CDR-H2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 17, and a CDR-H3 of SEQ ID NO: 33 or a CDR-H3 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 33; or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 34 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 34; (iii) a CDR-H1 of SEQ ID NO: 3 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 3, a CDR-H1 of SEQ ID NO: 19, (iv) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 4 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 4, a CDR-H2 of SEQ ID NO: 20 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 20, and a CDR-H3 of SEQ ID NO: 36. (v) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 5 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 5, a CDR-H2 of SEQ ID NO: 21 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 21, and a CDR-H3 of SEQ ID NO: 37 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 37;(vi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 6 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 6, a CDR-H2 of SEQ ID NO: 22 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 22, and a CDR-H3 of SEQ ID NO: 38 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 38; (vii) a CDR-H1 of SEQ ID NO: 7 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 7. (viii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 8 or a CDR-H1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 8, a CDR-H2 of SEQ ID NO: 24 or a CDR-H2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 24, and a CDR-H3 of SEQ ID NO: 39 or a CDR-H3 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 39; (ix) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 9 or a CDR-H1 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 9, a CDR-H2 of SEQ ID NO: 25 or a CDR-H2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 25, and a CDR-H3 of SEQ ID NO: 41 or a CDR-H3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 41. (x) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 10 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 10, a CDR-H2 of SEQ ID NO: 26 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 26, and a CDR-H3 of SEQ ID NO: 42 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 42;(xi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 11 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 11, a CDR-H2 of SEQ ID NO: 27 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 27, and a CDR-H3 of SEQ ID NO: 43 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 43; (xii) a CDR-H1 of SEQ ID NO: 12 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 12. (xiii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 13 or a CDR-H1 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 13, a CDR-H2 of SEQ ID NO: 29 or a CDR-H2 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 29, and a CDR-H3 of SEQ ID NO: 44 or a CDR-H3 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 44; (xiv) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 14 or a CDR-H1 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 14, a CDR-H2 of SEQ ID NO: 30 or a CDR-H2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 30, and a CDR-H3 of SEQ ID NO: 46 or a CDR-H3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 46. (xv) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 15 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 15, a CDR-H2 of SEQ ID NO: 31 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 31, and a CDR-H3 of SEQ ID NO: 47 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 47;(xvi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 16 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 16, CDR-H2 of SEQ ID NO: 32 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 32, and CDR-H3 of SEQ ID NO: 48 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 48; (xvii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 137. and (xviii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 136 or a CDR-H1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 136, a CDR-H2 of SEQ ID NO: 138 or a CDR-H2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 138, and a CDR-H3 of SEQ ID NO: 140 or a CDR-H3 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 140. The sequences of the heavy chain CDRs are provided in Table 1.

[0082] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain (X2m1) comprising: (ii.a) CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 240 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 240; (ii.b) CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 241 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 241. (ii.c) a heavy chain variable domain comprising a CDR-H3 (X2m2); (ii.c) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 242 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 242. (ii.d) a heavy chain variable domain (X2m4) comprising a CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 243 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 243; (ii.e) a heavy chain variable domain (X2m5) comprising CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 244 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 244; (ii.f) a heavy chain variable domain (X2m6) comprising CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 245 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 245; (ii. g) a heavy chain variable domain (X2m7) comprising CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 246 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 246; (ii.h) a heavy chain variable domain (X2m8) comprising CDR-H1 of SEQ ID NO: 2 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 247 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 247.

[0083] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises: (xvii.a) a CDR-H1 of SEQ ID NO: 135 or a CDR-H1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 135; a CDR-H2 of SEQ ID NO: 137 or a CDR-H2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 137; and a CDR-H3 of SEQ ID NO: 248 or a CDR-H3 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 248. (xvii.b) a heavy chain variable domain comprising a CDR-H3 having 5 amino acid modifications (X17m1); (xvii.b) a CDR-H1 of SEQ ID NO: 135 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 135, a CDR-H2 of SEQ ID NO: 137 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 137, and a CDR-H3 of SEQ ID NO: 249 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 249. (xvii.c) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 135 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 135, a CDR-H2 of SEQ ID NO: 137 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 137, and a CDR-H3 of SEQ ID NO: 250 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 250. (xvii.d) a heavy chain variable domain (X17m6) comprising CDR-H1 of SEQ ID NO: 135 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 251 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 251.

[0084] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: (xix) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 147 or a CDR-H1 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 147, a CDR-H2 of SEQ ID NO: 150 or a CDR-H2 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 150, and a CDR-H3 of SEQ ID NO: 156 or a CDR-H3 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 156; (xx) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 148 or a CDR-H1 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 148. (xxi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 149 or a CDR-H1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 149, a CDR-H2 of SEQ ID NO: 152 or a CDR-H2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 152, and a CDR-H3 of SEQ ID NO: 158 or a CDR-H3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 158.

[0085] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: (xxii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147 or a CDR-H1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 147, a CDR-H2 of SEQ ID NO: 153 or a CDR-H2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 153, and a CDR-H3 of SEQ ID NO: 156 or a CDR-H3 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g1m1 / 3B2g2m1); (xxiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147 or a CDR-H1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 147, a CDR-H2 of SEQ ID NO: 153 or a CDR-H2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 153 (xxiv) a heavy chain variable domain comprising a CDR-H2 of SEQ ID NO: 154 or a CDR-H2 having one, two, three, four or five amino acid modifications compared to SEQ ID NO: 154, and a CDR-H3 of SEQ ID NO: 156 or a CDR-H3 having one, two, three, four or five amino acid modifications compared to SEQ ID NO: 156 (3B2g1m2 / 3B2g2m2); (xxiv) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 147 or a CDR-H1 having one, two, three, four or five amino acid modifications compared to SEQ ID NO: 147, a CDR-H2 of SEQ ID NO: 155 or a CDR-H2 having one, two, three, four or five amino acid modifications compared to SEQ ID NO: 155, and a CDR-H3 of SEQ ID NO: 156 or a CDR-H3 having one, two, three, four or five amino acid modifications compared to SEQ ID NO: 156 (3B2g1m4 / 3B2g2m4). The sequences of the heavy chain CDRs are provided in Table 1.

[0086] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises: (i) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 1, CDR-H2 of SEQ ID NO: 17, and CDR-H3 of SEQ ID NO: 33; (ii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 34; (iii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 3, CDR-H2 of SEQ ID NO: 19, and CDR-H3 of SEQ ID NO: 35; (iv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 4, CDR-H2 of SEQ ID NO: 20, and CDR-H3 of SEQ ID NO: 21; and CDR-H2 of SEQ ID NO: 36; (v) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 5, a CDR-H2 of SEQ ID NO: 21, and a CDR-H3 of SEQ ID NO: 37; (vi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 6, a CDR-H2 of SEQ ID NO: 22, and a CDR-H3 of SEQ ID NO: 38; (vii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 7, a CDR-H2 of SEQ ID NO: 23, and a CDR-H3 of SEQ ID NO: 39; (viii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 8, a CDR-H2 of SEQ ID NO: 24, and a CDR-H3 of SEQ ID NO: (ix) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 9, a CDR-H2 of SEQ ID NO: 25, and a CDR-H3 of SEQ ID NO: 41; (x) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 10, a CDR-H2 of SEQ ID NO: 26, and a CDR-H3 of SEQ ID NO: 42; (xi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 11, a CDR-H2 of SEQ ID NO: 27, and a CDR-H3 of SEQ ID NO: 43; (xii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 12, a CDR-H2 of SEQ ID NO: 28, and a CDR-H3 of SEQ ID NO: 44. (xiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 13, CDR-H2 of SEQ ID NO: 29, and CDR-H3 of SEQ ID NO: 45; (xiv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 14, CDR-H2 of SEQ ID NO: 30, and CDR-H3 of SEQ ID NO: 46; (xv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 15, CDR-H2 of SEQ ID NO: 31, and CDR-H3 of SEQ ID NO: 47; (xvi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 16, CDR-H2 of SEQ ID NO: 32, and CDR-H3 of SEQ ID NO: 48;(xvii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 139; and (xviii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 136, CDR-H2 of SEQ ID NO: 138, and CDR-H3 of SEQ ID NO: 140. The sequences of the heavy chain CDR sequences are provided in Table 1 below.

[0087] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises: (ii.a) a heavy chain variable domain (X2m1) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 240; (ii.b) a heavy chain variable domain (X2m2) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 241; (ii.c) a heavy chain variable domain (X2m3) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 242; (ii.d) a heavy chain variable domain (X2m4) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 243. (ii.e) a heavy chain variable domain (X2m5) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 244; (ii.f) a heavy chain variable domain (X2m6) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 245; (ii.g) a heavy chain variable domain (X2m7) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 246; (ii.h) a heavy chain variable domain (X2m8) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18, and a CDR-H3 of SEQ ID NO: 247.

[0088] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises: (xvii.a) a heavy chain variable domain (X17m1) comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 248; (xvii.b) a heavy chain variable domain (X17m2) comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 249; (xvii.c) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 250 (X17m3); (xvii.d) a heavy chain variable domain (X17m6) comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 251.

[0089] In one embodiment, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: (xix) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156; (xx) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 148, CDR-H2 of SEQ ID NO: 151, and CDR-H3 of SEQ ID NO: 157; (xxi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158.

[0090] In certain embodiments, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: (xxii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 153, and CDR-H3 of SEQ ID NO: 156 (3B2g1m1 / 3B2g2m1); (xxiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 154, and CDR-H3 of SEQ ID NO: 156 (3B2g1m2 / 3B2g2m2); (xxiv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 155, and CDR-H3 of SEQ ID NO: 156 (3B2g1m4 / 3B2g2m4). The sequences of the heavy chain CDR sequences are provided in Table 1 below.

[0091] In one embodiment, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises a CDR-H1 of SEQ ID NO: 147, a CDR-H2 of SEQ ID NO: 153, or a CDR-H2 amino acid sequence having at least 0, 1, 2, 3, 4, or 5 modifications compared to SEQ ID NO: 153, and a CDR-H3 of SEQ ID NO: 156 (3B2g2m1). In one embodiment, the CDR-H2 amino acid sequence has at least 0, 1, 2, 3, 4, or 5 modifications compared to SEQ ID NO: 153. According to this embodiment, the CDR-H2 amino acid sequence has at least 0, 1, 2, 3, 4, or 5 modifications compared to SEQ ID NO: 153, wherein the modifications are present at residues 1, 2, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or any combination thereof.

[0092] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains :.

[0093] In certain embodiments, the CDR-H2 of the antibody comprises a proline (P) at position 3, a tryptophan (W) at position 4, and a serine (S) or asparagine (N) at position 5.

[0094] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain, wherein the heavy chain variable domain comprises: - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains :.

[0095] The sequences of the heavy chain CDR sequences are provided in Table 1 below. [Table 1] TIFF2025504935000004.tif219170TIFF2025504935000005.tif118170

[0096] In some embodiments, the MuSK antibody-based molecules disclosed herein further comprise a light chain variable domain. The light chain variable domain comprises: (i) a complementarity determining region 1 (CDR-L1) having an amino acid sequence of any one of SEQ ID NOs: 49 to 64, 141, 142, 159 to 169, or a modified amino acid sequence of any one of SEQ ID NOs: 49 to 64, 141, 142, or 159 to 169 (the modified sequence has at least 80% sequence identity to any one of SEQ ID NOs: 49 to 64, 141, 142, or 159 to 169); (ii) a complementarity determining region 2 (CDR-L2) having any one of the amino acid sequences of SEQ ID NOs: 65 to 80, 143, 144, and 170 to 179, or any one of the modified amino acid sequences of SEQ ID NOs: 65 to 80, 143, 144, and 170 to 179 (the modified sequence has at least 80% sequence identity with any one of SEQ ID NOs: 65 to 80, 143, 144, and 170 to 179); and (iii) Complementarity determining region 3 (CDR-L3) having any one of the amino acid sequences of SEQ ID NOs: 81 to 96, 145, 146, and 180 to 195, or any one of the modified amino acid sequences of SEQ ID NOs: 81 to 96, 145, 146, and 180 to 195 (the modified sequence has at least 80% sequence identity with any one of SEQ ID NOs: 81 to 96, 145, 146, and 180 to 195). Includes.

[0097] In some embodiments, the MuSK antibody-based molecules disclosed herein further comprise a light chain variable domain. The light chain variable domain comprises: (iv) a complementarity determining region 1 (CDR-L1) having an amino acid sequence of any one of SEQ ID NOs: 49 to 64, 141, 142, 159 to 169, or a modified amino acid sequence of any one of SEQ ID NOs: 49 to 64, 141, 142, or 159 to 169 (the modified sequence has 1, 2, 3, 4, or 5 amino acid modifications compared to any one of SEQ ID NOs: 49 to 64, 141, 142, or 159 to 169); (v) a complementarity determining region 2 (CDR-L2) having any one of the amino acid sequences of SEQ ID NOs: 65 to 80, 143, 144, 170 to 179, or any one of the modified amino acid sequences of SEQ ID NOs: 65 to 80, 143, 144, or 170 to 179 (the modified sequence has 1, 2, 3, 4, or 5 amino acid alterations compared to any one of SEQ ID NOs: 65 to 80, 143, 144, or 170 to 179); and (vi) Complementarity determining region 3 (CDR-L3) having any one of the amino acid sequences of SEQ ID NOs: 81 to 96, 145, 146, 180 to 195, or any one of the modified amino acid sequences of SEQ ID NOs: 81 to 96, 145, 146, or 180 to 195 (the modified sequence has 1, 2, 3, 4, or 5 amino acid modifications compared to any one of SEQ ID NOs: 81 to 96, 145, 146, or 180 to 195). Includes.

[0098] In certain embodiments, the light chain variable domain of the MuSK antibody-based molecules disclosed herein comprises a light chain variable domain comprising: (i) a CDR-L1 of SEQ ID NO: 49 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 49, a CDR-L2 of SEQ ID NO: 65 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 65, and a CDR-L3 of SEQ ID NO: 81 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 81; (ii) a CDR-L1 of SEQ ID NO: 50 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 50. -L1 or CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 50, CDR-L2 of SEQ ID NO: 66 or CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 66, and CDR-L3 of SEQ ID NO: 82 or CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 82; (iii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 51 or CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 51, CDR-L3 of SEQ ID NO: 67 (iv) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 52 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 52, a CDR-L2 of SEQ ID NO: 68 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 68, and a CDR-L3 of SEQ ID NO: 84. (v) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 53 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 53, a CDR-L2 of SEQ ID NO: 69 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 69, and a CDR-L3 of SEQ ID NO: 85 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 85;(vi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 54 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 54, a CDR-L2 of SEQ ID NO: 70 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 70, and a CDR-L3 of SEQ ID NO: 86 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 86; (vii) a CDR-L1 of SEQ ID NO: 55 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 55. (viii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO:56 or a CDR-L1 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO:56, a CDR-L2 of SEQ ID NO:72 or a CDR-L2 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO:72; (ix) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 57 or a CDR-L1 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 57, a CDR-L2 of SEQ ID NO: 73 or a CDR-L2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 73, and a CDR-L3 of SEQ ID NO: 89 or a CDR-L3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 89. (x) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 58 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 58, a CDR-L2 of SEQ ID NO: 74 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 74, and a CDR-L3 of SEQ ID NO: 90 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 90;(xi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 59 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 59, a CDR-L2 of SEQ ID NO: 75 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 75, and a CDR-L3 of SEQ ID NO: 91 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 91; (xii) a CDR-L1 of SEQ ID NO: 60 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 60. (xiii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 61 or a CDR-L1 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 61, a CDR-L2 of SEQ ID NO: 77 or a CDR-L2 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 77, and a CDR-L3 of SEQ ID NO: 92 or a CDR-L3 with 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 92; (xiv) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 62 or a CDR-L1 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 62, a CDR-L2 of SEQ ID NO: 78 or a CDR-L2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 78, and a CDR-L3 of SEQ ID NO: 94 or a CDR-L3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 94; (xv) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 63 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 63, a CDR-L2 of SEQ ID NO: 79 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 79, and a CDR-L3 of SEQ ID NO: 95 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 95;(xvi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 64 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 64, CDR-L2 of SEQ ID NO: 80 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 80, and CDR-L3 of SEQ ID NO: 96 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 96; (xvii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 143. (xviii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 142 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 142, a CDR-L2 of SEQ ID NO: 144 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 144, and a CDR-L3 of SEQ ID NO: 146 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 146. The sequences of the light chain CDRs are provided in Table 2 below.

[0099] In certain embodiments, the light chain variable domain of the MuSK antibody-based molecules disclosed herein comprises a light chain variable domain comprising: (xix) a CDR-L1 of SEQ ID NO: 159 or a CDR-L1 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 159, a CDR-L2 of SEQ ID NO: 170 or a CDR-L2 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 170, and a CDR-L3 of SEQ ID NO: 180 or a CDR-L3 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 180; (xx) a CDR-L1 of SEQ ID NO: 159 or a CDR-L1 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 159, a CDR-L2 of SEQ ID NO: 171 or a CDR-L2 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 171, and a CDR-L3 of SEQ ID NO: 181 or a CDR-L3 having one, two, three, four, or five amino acid modifications compared to SEQ ID NO: 181. (xxi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 160 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 160, a CDR-L2 of SEQ ID NO: 172 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 172, and a CDR-L3 of SEQ ID NO: 182 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications relative to SEQ ID NO: 182. (xxii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 159 or a CDR-L1 having one, two, three, four, or five amino acid modifications relative to SEQ ID NO: 159, a CDR-L2 of SEQ ID NO: 172 or a CDR-L2 having one, two, three, four, or five amino acid modifications relative to SEQ ID NO: 172, and a CDR-L3 of SEQ ID NO: 183 or a CDR-L3 having one, two, three, four, or five amino acid modifications relative to SEQ ID NO: 183;(xxiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 184 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 184; (xxiv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 159. (xxv) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 159 or a CDR-L1 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159, a CDR-L2 of SEQ ID NO: 173 or a CDR-L2 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 173, and a CDR-L3 of SEQ ID NO: 185 or a CDR-L3 with 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 185; (xxvi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 161 or a CDR-L1 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 161, a CDR-L2 of SEQ ID NO: 174 or a CDR-L2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 174, and a CDR-L3 of SEQ ID NO: 187 or a CDR-L3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 187; (xxvii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 162 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 162, a CDR-L2 of SEQ ID NO: 174 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 174, and a CDR-L3 of SEQ ID NO: 188 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 188;(xxviii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 163 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 163, CDR-L2 of SEQ ID NO: 174 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 188 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 188; (xxix) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 164 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 164. a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 165 or a CDR-L2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 165, a CDR-L2 of SEQ ID NO: 175 or a CDR-L2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 175, and a CDR-L3 of SEQ ID NO: 189 or a CDR-L3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 189; (xxxi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 166 or a CDR-L1 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 166, a CDR-L2 of SEQ ID NO: 176 or a CDR-L2 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 176, and a CDR-L3 of SEQ ID NO: 191 or a CDR-L3 having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 191; (xxxi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 167 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 167, a CDR-L2 of SEQ ID NO: 177 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 177, and a CDR-L3 of SEQ ID NO: 192 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 192;(xxxii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 168 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 168, a CDR-L2 of SEQ ID NO: 178 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 178, and a CDR-L3 of SEQ ID NO: 193 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 193; (xxxiii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 169 or a CDR-L1 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 169, a CDR-L2 of SEQ ID NO: 179 or a CDR-L2 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 179, and a CDR-L3 of SEQ ID NO: 194 or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 194;

[0100] In certain embodiments, the light chain variable domain of the MuSK antibody-based molecules disclosed herein comprises: (i) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 49, CDR-L2 of SEQ ID NO: 65, and CDR-L3 of SEQ ID NO: 81; (ii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 50, CDR-L2 of SEQ ID NO: 66, and CDR-L3 of SEQ ID NO: 82; (iii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 51, CDR-L2 of SEQ ID NO: 67, and CDR-L3 of SEQ ID NO: 83; (iv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 52, CDR-L2 of SEQ ID NO: 68, and CDR-L3 of SEQ ID NO: 83; (v) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 53, CDR-L2 of SEQ ID NO: 69, and CDR-L3 of SEQ ID NO: 85; (vi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 54, CDR-L2 of SEQ ID NO: 70, and CDR-L3 of SEQ ID NO: 86; (vii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 55, CDR-L2 of SEQ ID NO: 71, and CDR-L3 of SEQ ID NO: 87; (viii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 56, CDR-L2 of SEQ ID NO: 72, and CDR-L3 of SEQ ID NO: (ix) a light chain variable domain comprising a CDR-L1 of SEQ ID NO:57, a CDR-L2 of SEQ ID NO:73, and a CDR-L3 of SEQ ID NO:89; (x) a light chain variable domain comprising a CDR-L1 of SEQ ID NO:58, a CDR-L2 of SEQ ID NO:74, and a CDR-L3 of SEQ ID NO:90; (xi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO:59, a CDR-L2 of SEQ ID NO:75, and a CDR-L3 of SEQ ID NO:91; (xii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO:60, a CDR-L2 of SEQ ID NO:76, and a CDR-L3 of SEQ ID NO:92. (xiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 61, CDR-L2 of SEQ ID NO: 77, and CDR-L3 of SEQ ID NO: 93; (xiv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 62, CDR-L2 of SEQ ID NO: 78, and CDR-L3 of SEQ ID NO: 94; (xv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 63, CDR-L2 of SEQ ID NO: 79, and CDR-L3 of SEQ ID NO: 95; (xvi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 64, CDR-L2 of SEQ ID NO: 80, and CDR-L3 of SEQ ID NO: 96;(xvii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145; (xviii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 142, CDR-L2 of SEQ ID NO: 144, and CDR-L3 of SEQ ID NO: 146. The sequences of the light chain CDRs are provided in Table 2 below.

[0101] In certain embodiments, the light chain variable domain of the MuSK antibody-based molecules disclosed herein comprises: (xix) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 170, and CDR-L3 of SEQ ID NO: 180; (xx) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 181; (xxi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 160, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 182; (xii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183; (xxiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 184; (xxiv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 173, and CDR-L3 of SEQ ID NO: 185; (xxv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 173, and CDR-L3 of SEQ ID NO: 186. (xxvi) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 161, a CDR-L2 of SEQ ID NO: 174, and a CDR-L3 of SEQ ID NO: 187; (xxvii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 162, a CDR-L2 of SEQ ID NO: 174, and a CDR-L3 of SEQ ID NO: 188; (xxviii) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 163, a CDR-L2 of SEQ ID NO: 174, and a CDR-L3 of SEQ ID NO: 188; (xxix) a light chain variable domain comprising a CDR-L1 of SEQ ID NO: 164, , CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 189; (xxx) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 165, CDR-L2 of SEQ ID NO: 175, and CDR-L3 of SEQ ID NO: 190; (xxxi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 166, CDR-L2 of SEQ ID NO: 176, and CDR-L3 of SEQ ID NO: 191; (xxxi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 167, CDR-L2 of SEQ ID NO: 177, and CDR-L3 of SEQ ID NO: 192;(xxxii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 168, CDR-L2 of SEQ ID NO: 178, and CDR-L3 of SEQ ID NO: 193; (xxxiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 169, CDR-L2 of SEQ ID NO: 179, and CDR-L3 of SEQ ID NO: 194;

[0102] In one embodiment, the light chain variable domain of a MuSK antibody-based molecule disclosed herein comprises a CDR-L1 of SEQ ID NO: 159, a CDR-L2 of SEQ ID NO: 172, and a CDR-L3 of SEQ ID NO: 195, or a CDR-L3 having 1, 2, 3, 4, or 5 amino acid modifications compared to the amino acid sequence of SEQ ID NO: 195, wherein the modifications are present at residues 1, 2, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or any combination thereof.

[0103] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a light chain variable domain, wherein the light chain variable domain comprises: - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159 - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0104] In certain embodiments, the CDR-L1, CDR-L2, CDR-L3 amino acid sequences have at least 0, 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 159, 172, or 195 (respectively).

[0105] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a light chain variable domain, wherein the light chain variable domain comprises: - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0106] The sequences of the light chain CDRs are provided in Table 2 below. [Table 2] TIFF2025504935000007.tif160170

[0107] Suitable amino acid modifications to the heavy and / or light chain CDR sequences of the MuSK antibody-based molecules disclosed herein include, for example, conservative substitutions or functionally equivalent amino acid residue substitutions that result in variant CDR sequences with similar or enhanced binding characteristics to those of the CDR sequences disclosed herein as described above. Encompassed by the present invention are CDRs of Tables 1 and 2 that contain one, two, three, four, five or more amino acid alterations (depending on the length of the CDR) that maintain or enhance MuSK binding of the antibody. Suitable amino acid modifications to the heavy chain CDR sequences of Tables 1 and / or the light chain CDR sequences of Tables 1 and 2 include, for example, conservative substitutions or functionally equivalent amino acid residue substitutions that result in variant CDR sequences with similar or enhanced binding characteristics to those of the CDR sequences of Tables 1 and 2. Conservative substitutions are those that take place within a family of amino acids whose side chains are related. Genetically encoded amino acids can be classified into four families: (1) acidic (aspartate, glutamate); (2) basic (lysine, arginine, histidine); (3) nonpolar (alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan); and (4) uncharged polar (glycine, asparagine, glutamine, cysteine, serine, threonine, tyrosine). Phenylalanine, tryptophan, and tyrosine are sometimes classified jointly as aromatic amino acids. Alternatively, the amino acid repertoire can be grouped into: (1) acidic (aspartate, glutamate); (2) basic (lysine, arginine, histidine); (3) aliphatic (glycine, alanine, valine, leucine, isoleucine, serine, threonine), with serine and threonine optionally grouped separately as aliphatic hydroxyls; (4) aromatic (phenylalanine, tyrosine, tryptophan); (5) amide (asparagine, glutamine); and (6) sulfur-containing (cysteine ​​and methionine) (Stryer (Ed.), Biochemistry, 2nd Edition, WH Freeman and Co., 1981, incorporated herein by reference in its entirety). Non-conservative substitutions can also be made to the heavy chain CDR sequences in Table 1 and the light chain CDR sequences in Table 2.Non-conservative substitutions involve replacing one or more amino acid residues in a CDR with one or more amino acid residues from a different class of amino acids to improve or enhance the binding properties of the CDR. The amino acid sequences of the heavy chain CDRs of Table 1 and / or the light chain CDRs of Table 2 may further comprise one or more internal neutral amino acid insertions or deletions that maintain or enhance MuSK binding.

[0108] In one embodiment, the MuSK antibody-based molecule comprises: (i) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 1, CDR-H2 of SEQ ID NO: 17, and CDR-H3 of SEQ ID NO: 33, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 49, CDR-L2 of SEQ ID NO: 65, and CDR-L3 of SEQ ID NO: 81; (ii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 34, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 50, CDR-L2 of SEQ ID NO: 66, and CDR-L3 of SEQ ID NO: 82; (iii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 3, CDR-H2 of SEQ ID NO: 19, and CDR-H3 of SEQ ID NO: 35, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 51, CDR-L2 of SEQ ID NO: 67, and CDR-L3 of SEQ ID NO: 83; (iv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 4, CDR-H2 of SEQ ID NO: 20, and CDR-H3 of SEQ ID NO: 36, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 52, CDR-L2 of SEQ ID NO: 68, and CDR-L3 of SEQ ID NO: 84; (v) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:5, CDR-H2 of SEQ ID NO:21, and CDR-H3 of SEQ ID NO:37, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:53, CDR-L2 of SEQ ID NO:69, and CDR-L3 of SEQ ID NO:85; (vi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 6, CDR-H2 of SEQ ID NO: 22, and CDR-H3 of SEQ ID NO: 38, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 54, CDR-L2 of SEQ ID NO: 70, and CDR-L3 of SEQ ID NO: 86; (vii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 7, CDR-H2 of SEQ ID NO: 23, and CDR-H3 of SEQ ID NO: 39, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 55, CDR-L2 of SEQ ID NO: 71, and CDR-L3 of SEQ ID NO: 87; (viii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 8, CDR-H2 of SEQ ID NO: 24, and CDR-H3 of SEQ ID NO: 40, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 56, CDR-L2 of SEQ ID NO: 72, and CDR-L3 of SEQ ID NO: 88; (ix) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 9, CDR-H2 of SEQ ID NO: 25, and CDR-H3 of SEQ ID NO: 41, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 57, CDR-L2 of SEQ ID NO: 73, and CDR-L3 of SEQ ID NO: 89; (x) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 10, CDR-H2 of SEQ ID NO: 26, and CDR-H3 of SEQ ID NO: 42, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 58, CDR-L2 of SEQ ID NO: 74, and CDR-L3 of SEQ ID NO: 90; (xi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 11, CDR-H2 of SEQ ID NO: 27, and CDR-H3 of SEQ ID NO: 43, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 59, CDR-L2 of SEQ ID NO: 75, and CDR-L3 of SEQ ID NO: 91; (xii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 12, CDR-H2 of SEQ ID NO: 28, and CDR-H3 of SEQ ID NO: 44, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 60, CDR-L2 of SEQ ID NO: 76, and CDR-L3 of SEQ ID NO: 92; (xiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 13, CDR-H2 of SEQ ID NO: 29, and CDR-H3 of SEQ ID NO: 45, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 61, CDR-L2 of SEQ ID NO: 77, and CDR-L3 of SEQ ID NO: 93; (xiv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 14, CDR-H2 of SEQ ID NO: 30, and CDR-H3 of SEQ ID NO: 46, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 62, CDR-L2 of SEQ ID NO: 78, and CDR-L3 of SEQ ID NO: 94; (xv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 15, CDR-H2 of SEQ ID NO: 31, and CDR-H3 of SEQ ID NO: 47, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 63, CDR-L2 of SEQ ID NO: 79, and CDR-L3 of SEQ ID NO: 95; (xvi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 16, CDR-H2 of SEQ ID NO: 32, and CDR-H3 of SEQ ID NO: 48, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 64, CDR-L2 of SEQ ID NO: 80, and CDR-L3 of SEQ ID NO: 96; (xvii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 139, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145; and (xviii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 136, CDR-H2 of SEQ ID NO: 138, and CDR-H3 of SEQ ID NO: 140, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 142, CDR-L2 of SEQ ID NO: 144, and CDR-L3 of SEQ ID NO: 146. Contains :.

[0109] In one embodiment, the MuSK antibody-based molecule comprises: (ii.a) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:2, CDR-H2 of SEQ ID NO:18, and CDR-H3 of SEQ ID NO:240, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:50, CDR-L2 of SEQ ID NO:66, and CDR-L3 of SEQ ID NO:82 (X2m1); (ii.b) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 241, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 50, CDR-L2 of SEQ ID NO: 66, and CDR-L3 of SEQ ID NO: 82 (X2m2); (ii.c) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:2, CDR-H2 of SEQ ID NO:18, and CDR-H3 of SEQ ID NO:242, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:50, CDR-L2 of SEQ ID NO:66, and CDR-L3 of SEQ ID NO:82 (X2m3); (ii.d) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:2, CDR-H2 of SEQ ID NO:18, and CDR-H3 of SEQ ID NO:243, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:50, CDR-L2 of SEQ ID NO:66, and CDR-L3 of SEQ ID NO:82 (X2m4); (ii.e) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:2, CDR-H2 of SEQ ID NO:18, and CDR-H3 of SEQ ID NO:244, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:50, CDR-L2 of SEQ ID NO:66, and CDR-L3 of SEQ ID NO:82 (X2m5); (ii.f) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:2, CDR-H2 of SEQ ID NO:18, and CDR-H3 of SEQ ID NO:245, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:50, CDR-L2 of SEQ ID NO:66, and CDR-L3 of SEQ ID NO:82 (X2m6); (ii.g) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO:2, CDR-H2 of SEQ ID NO:18, and CDR-H3 of SEQ ID NO:246, and a light chain variable domain comprising CDR-L1 of SEQ ID NO:50, CDR-L2 of SEQ ID NO:66, and CDR-L3 of SEQ ID NO:82 (X2m7); (ii.f) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 247, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 50, CDR-L2 of SEQ ID NO: 66, and CDR-L3 of SEQ ID NO: 82 (X2m8). Contains :.

[0110] In one embodiment, the MuSK antibody-based molecule comprises: (xvii.a) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 248, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145 (X17m1); (xvii.b) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 249, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145 (X17m2); (xvii.c) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 250, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145 (X17m3); (xvii.d) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 251, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145 (X17m6). Contains :.

[0111] In one embodiment, the MuSK antibody-based molecule comprises: (i) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 170, and CDR-L3 of SEQ ID NO: 180 (14D10); (ii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 181 (7G4); (iii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 160, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 182 (3C4); (iv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183 (3B2); (v) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 184 (3G3); (vi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 173, and CDR-L3 of SEQ ID NO: 185 (31G2); (vii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 173, and CDR-L3 of SEQ ID NO: 186 (31B7); (viii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 148, CDR-H2 of SEQ ID NO: 151, and CDR-H3 of SEQ ID NO: 157, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 161, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 187 (17H10); (ix) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 148, CDR-H2 of SEQ ID NO: 151, and CDR-H3 of SEQ ID NO: 157, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 162, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 188 (23B6); (x) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 148, CDR-H2 of SEQ ID NO: 151, and CDR-H3 of SEQ ID NO: 157, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 163, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 188 (30E1); (xi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 148, CDR-H2 of SEQ ID NO: 151, and CDR-H3 of SEQ ID NO: 157, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 164, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 189 (30A11); (xii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 165, CDR-L2 of SEQ ID NO: 175, and CDR-L3 of SEQ ID NO: 190 (16F11); (xiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 166, CDR-L2 of SEQ ID NO: 176, and CDR-L3 of SEQ ID NO: 191 (4C11); (xiv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 167, CDR-L2 of SEQ ID NO: 177, and CDR-L3 of SEQ ID NO: 192 (7A12); (xv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 168, CDR-L2 of SEQ ID NO: 178, and CDR-L3 of SEQ ID NO: 193 (7G12); (xvi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 169, CDR-L2 of SEQ ID NO: 179, and CDR-L3 of SEQ ID NO: 194 (7B8); (xvii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 153, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183 (3B2g1m1); (xviii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 154, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183 (3B2g1m2); (xvix) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 155, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183 (3B2g1m4); (xx) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 153, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 195 (3B2g2m1); (xxi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 154, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 195 (3B2g2m2); and (xxii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 155, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 195 (3B2g2m4) Contains :.

[0112] In a preferred embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises: - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0113] In certain embodiments, the CDR-H2 of the antibody comprises a proline (P) at position 3, a tryptophan (W) at position 4, and a serine (S) or asparagine (N) at position 5.

[0114] In a more preferred embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0115] The MuSK antibody-based molecules described herein can comprise a variable light (VL) chain, a variable heavy (VH) chain, or a combination of a VL chain and a VH chain. In some embodiments, the VH chain of the MuSK antibody-based molecule has a sequence similar to any one of the VH amino acid sequences provided in Table 3 below, or any one of the VH amino acid sequences listed in Table 3 by at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 78%, at least 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 97%, at least 98%, at least 99%, at least 100%, at least 101%, at least 102%, at least 103%, at least 104%, at least 105%, at least 106%, at least 107%, at least 108%, at least 109%, at least 110%, at least 111%, at least 112%, at least 113%, at least 114%, at least 115%, at least 116%, at least 117%, at least 118%, at least 119%, at least 120%, at least 121%, at least 122%, at least 123%, at least 124%, at least 125%, at least 12 In some embodiments, the VL chain of the MuSK antibody-based molecule comprises an amino acid sequence that is at least 76%, at least 77%, at least 78%, at least 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical or similar to any one of the VL amino acid sequences provided in Table 3 below, or any one of the VL amino acid sequences listed in Table 3.In some embodiments, the identity or similarity is at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%. [Table 3] TIFF2025504935000009.tif216170TIFF2025504935000010.tif215170TIFF2025504935000011.tif216170TIFF2025504935 000012.tif216170TIFF2025504935000013.tif216170TIFF2025504935000014.tif216170TIFF2025504935000015.tif60170

[0116] In certain embodiments, the MuSK antibody-based molecules disclosed herein are (ii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 97 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 98; (ii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to any one of SEQ ID NOs: 99 and 252 to 259, and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 100; (iii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 101 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 102; (iv) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 103 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 104; (v) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 105 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 106; (vi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 107 or (vii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 109 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 110; (viii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 111 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 112; (ix) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 113 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 114; (x) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 115 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 116; (xi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 117 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 118;(xii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 119 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 120; (xiii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 121 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 122; (xiv) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 123 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 124; (xv) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 125 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 126. (xvi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 127 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 128; (xvii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to any one of SEQ ID NOs: 131 and 260 to 263 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 132; and (xviii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 133 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 134; Contains :.

[0117] In some embodiments, the MuSK antibody-based molecules disclosed herein comprise: (i) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 196 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 197; (ii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 198 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 199; (iii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 200 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 201; (iv) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 202 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 203; (v) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 204 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 205; (vi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 206. (vii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 208 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 209; (viii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 210 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 211; (vix) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 212 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 213; (x) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 214 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 215; (xi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 216 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 217;(xii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 218 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 219; (xiii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 220 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 221; (xiv) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 222 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 223; (xv) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 224 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 225; (xvi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 226 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 227; (xvii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 228 and and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 229; (xviii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 230 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 231; (xix) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 232 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 233; (xx) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 234 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 235; (xxi) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 236 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 237; (xxii) a heavy chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 238 and a light chain variable domain comprising an amino acid sequence at least 80% identical to SEQ ID NO: 239.

[0118] In a preferred embodiment, the MuSK antibody-based molecules (or anti-MuSK antibodies or antigen-binding fragments thereof) disclosed herein are A heavy chain variable domain (VH) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234 and a light chain variable domain (VL) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235. Includes.

[0119] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0120] In a preferred embodiment, the MuSK antibody-based molecules disclosed herein comprise: It comprises a heavy chain variable domain comprising the amino acid sequence of SEQ ID NO:234 ​​and a light chain variable domain comprising the amino acid sequence of SEQ ID NO:235.

[0121] In a preferred embodiment, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0122] In a more preferred embodiment, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235; wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0123] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises at least 80% sequence identity to SEQ ID NO: 266 or 267, wherein the heavy chain variable domain comprises an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 235; wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0124] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0125] In one embodiment, the CDR-H2 of the antibody comprises a proline (P) at position 3, a tryptophan (W) at position 4, and a serine (S) or an asparagine (N) at position 5.

[0126] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235; wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0127] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0128] In one embodiment, the CDR-H2 of the antibody comprises a proline (P) at position 3, a tryptophan (W) at position 4, and a serine (S) or an asparagine (N) at position 5.

[0129] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, wherein the heavy chain variable domain comprises SEQ ID NO: 234, and the light chain variable domain comprises SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0130] In one embodiment, the CDR-H2 of the antibody comprises a proline (P) at position 3, a tryptophan (W) at position 4, and a serine (S) or an asparagine (N) at position 5.

[0131] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, wherein the heavy chain variable domain comprises SEQ ID NO: 234, and the light chain variable domain comprises SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - the CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0132] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain; wherein the wild type human IgG constant Fc region comprises at least 80% sequence identity to SEQ ID NO: 266 or 267; wherein a) the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; one or more of a G236R substitution; a G236K substitution; a G237A substitution; a P238A substitution; a F243L substitution; a D265A substitution; a S267E substitution; a H268A substitution; a R292P substitution; a Y300L substitution; a K322A substitution; a K322Q substitution; a A327Q substitution; a L328F substitution; a L328R substitution; a P329A substitution; a P329G substitution; a A330L substitution; a A330S substitution; a P331S substitution; a I332E substitution; a P396L substitution; or each of the combinations of mutations previously described in the fourth embodiment of the present application are introduced into the full length heavy chain, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A, and wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0133] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0134] In one embodiment, the antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain; wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, wherein an L234A and / or L235A substitution, numbered according to the EU numbering system, is introduced into the Fc region; and wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235; wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0135] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0136] In certain embodiments, an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, wherein L234A and L235A substitutions, numbered according to the EU numbering system, are introduced into the Fc region; and wherein the heavy chain variable domain comprises SEQ ID NO: 234 and the light chain variable domain comprises SEQ ID NO: 235; and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0137] In one embodiment, the anti-MuSK antibody, or antigen-binding fragment thereof, a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 268, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 269. Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions are introduced into the full length heavy chain: G236R; G236K; G237A; P238A; F243L; D265A; S267E; H268A; R292P; Y300L; K322A; K322Q; A327Q; L328F; L328R; P329A; P329G; A330L; A330S; P331S; I332E; P396L or each of the combinations of mutations previously described in the fourth embodiment of the present application, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A.

[0138] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0139] In one embodiment, the anti-MuSK antibody, or antigen-binding fragment thereof, a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0140] In one embodiment, the anti-MuSK antibody, or antigen-binding fragment thereof, a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 270, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 271 Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions are introduced into the full length heavy chain: G236R; G236K; G237A; P238A; F243L; D265A; S267E; H268A; R292P; Y300L; K322A; K322Q; A327Q; L328F; L328R; P329A; P329G; A330L; A330S; P331S; I332E; P396L or each of the combinations of mutations previously described in the fourth embodiment of the present application, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A.

[0141] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0142] In one embodiment, the anti-MuSK antibody, or antigen-binding fragment thereof, a) a full length heavy chain comprising SEQ ID NO: 270, and b) a full-length light chain comprising SEQ ID NO: 271 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0143] (Polynucleotide) Another aspect of the invention is directed to an isolated polynucleotide encoding a MuSK antibody-based molecule described herein. In one embodiment, a polynucleotide encoding a MuSK antibody of the invention comprises a nucleotide sequence encoding any one, any two, any three, any four, any five, or any six of the above CDRs, including the heavy chain CDRs of SEQ ID NOs: 1-48, 135-140, 147-158, 240-251 and the light chain CDRs of SEQ ID NOs: 49-96, 141-146, and 159-195.

[0144] Thus, the invention provides a polynucleotide for use in treating a neuromuscular disease in a human subject, the polynucleotide comprising a nucleotide sequence encoding an anti-MuSK antibody or antigen-binding fragment or its VH, VL, or CDR domains.

[0145] In certain embodiments, the polynucleotide comprises a nucleotide sequence encoding a VH domain, wherein the VH domain comprises: (i) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 1, CDR-H2 of SEQ ID NO: 17, and CDR-H3 of SEQ ID NO: 33; (ii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 34; (iii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 3, CDR-H2 of SEQ ID NO: 19, and CDR-H3 of SEQ ID NO: 35; (iv) a CDR-H1 of SEQ ID NO: 4, (v) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO:5, a CDR-H2 of SEQ ID NO:21, and a CDR-H3 of SEQ ID NO:37; (vi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO:6, a CDR-H2 of SEQ ID NO:22, and a CDR-H3 of SEQ ID NO:38; (vii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO:7, a CDR-H2 of SEQ ID NO:23, and a CDR-H3 of SEQ ID NO:39; (viii) a CDR-H1 of SEQ ID NO:8, a CDR-H2 of SEQ ID NO:24, and (ix) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 9, a CDR-H2 of SEQ ID NO: 25, and a CDR-H3 of SEQ ID NO: 41; (x) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 10, a CDR-H2 of SEQ ID NO: 26, and a CDR-H3 of SEQ ID NO: 42; (xi) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 11, a CDR-H2 of SEQ ID NO: 27, and a CDR-H3 of SEQ ID NO: 43; (xii) a heavy chain variable domain comprising a CDR-H1 of SEQ ID NO: 12, a CDR-H2 of SEQ ID NO: 28, and a CDR-H3 of SEQ ID NO: 44. (xiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 13, CDR-H2 of SEQ ID NO: 29, and CDR-H3 of SEQ ID NO: 45; (xiv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 14, CDR-H2 of SEQ ID NO: 30, and CDR-H3 of SEQ ID NO: 46; (xv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 15, CDR-H2 of SEQ ID NO: 31, and CDR-H3 of SEQ ID NO: 47; (xvi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 16, CDR-H2 of SEQ ID NO: 32, and CDR-H3 of SEQ ID NO: 48;(xvii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 139; and (xviii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 136, CDR-H2 of SEQ ID NO: 138, and CDR-H3 of SEQ ID NO: 140.

[0146] In some embodiments, the polynucleotide comprises a nucleotide sequence encoding a VH domain, wherein the VH domain comprises: (ii.a) a heavy chain variable domain (X2m1) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 240; (ii.b) a heavy chain variable domain (X2m2) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 241; (ii.c) a heavy chain variable domain (X2m3) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: 242; (ii.d) a heavy chain variable domain (X2m4) comprising CDR-H1 of SEQ ID NO: 2, CDR-H2 of SEQ ID NO: 18, and CDR-H3 of SEQ ID NO: (ii.e) a heavy chain variable domain (X2m5) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 and a CDR-H3 of SEQ ID NO: 244; (ii.f) a heavy chain variable domain (X2m6) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 and a CDR-H3 of SEQ ID NO: 245; (ii.g) a heavy chain variable domain (X2m7) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 and a CDR-H3 of SEQ ID NO: 246; (ii.h) a heavy chain variable domain (X2m8) comprising a CDR-H1 of SEQ ID NO: 2, a CDR-H2 of SEQ ID NO: 18 and a CDR-H3 of SEQ ID NO: 247.

[0147] In some embodiments, the polynucleotide comprises a nucleotide sequence encoding a VH domain, wherein the VH domain comprises: (xvii.a) a heavy chain variable domain (X17m1) comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 248; (xvii.b) a heavy chain variable domain (X17m2) comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 249; (xvii.c) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 250; (xvii.d) a heavy chain variable domain (X17m3) comprising CDR-H1 of SEQ ID NO: 135, CDR-H2 of SEQ ID NO: 137, and CDR-H3 of SEQ ID NO: 251 (X17m6).

[0148] In one embodiment, the polynucleotide comprises a nucleotide sequence encoding a VH domain, wherein the VH domain comprises: (xix) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156; (xx) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 148, CDR-H2 of SEQ ID NO: 151, and CDR-H3 of SEQ ID NO: 157; (xxi) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 149, CDR-H2 of SEQ ID NO: 152, and CDR-H3 of SEQ ID NO: 158.

[0149] In one embodiment, the polynucleotide comprises a nucleotide sequence encoding a VH domain, wherein the VH domain comprises: (xxii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 153, and CDR-H3 of SEQ ID NO: 156; (xxiii) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 154, and CDR-H3 of SEQ ID NO: 156; (xxiv) a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 155, and CDR-H3 of SEQ ID NO: 156.

[0150] In certain embodiments, the polynucleotide comprises a nucleotide sequence encoding a VL domain, wherein the VL domain comprises: (i) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 49, CDR-L2 of SEQ ID NO: 65, and CDR-L3 of SEQ ID NO: 81; (ii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 50, CDR-L2 of SEQ ID NO: 66, and CDR-L3 of SEQ ID NO: 82; (iii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 51, CDR-L2 of SEQ ID NO: 67, and CDR-L3 of SEQ ID NO: 83; (v) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 52, CDR-L2 of SEQ ID NO: 68, and CDR-L3 of SEQ ID NO: 84; (v) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 53, CDR-L2 of SEQ ID NO: 69, and CDR-L3 of SEQ ID NO: 85; (vi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 54, CDR-L2 of SEQ ID NO: 70, and CDR-L3 of SEQ ID NO: 86; (vii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 55, CDR-L2 of SEQ ID NO: 71, and CDR-L3 of SEQ ID NO: 87; (vi (ii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 56, CDR-L2 of SEQ ID NO: 72, and CDR-L3 of SEQ ID NO: 88; (ix) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 57, CDR-L2 of SEQ ID NO: 73, and CDR-L3 of SEQ ID NO: 89; (x) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 58, CDR-L2 of SEQ ID NO: 74, and CDR-L3 of SEQ ID NO: 90; (xi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 59, CDR-L2 of SEQ ID NO: 75, and CDR-L3 of SEQ ID NO: 91; (xi i) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 60, CDR-L2 of SEQ ID NO: 76, and CDR-L3 of SEQ ID NO: 92; (xiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 61, CDR-L2 of SEQ ID NO: 77, and CDR-L3 of SEQ ID NO: 93; (xiv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 62, CDR-L2 of SEQ ID NO: 78, and CDR-L3 of SEQ ID NO: 94; (xv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 63, CDR-L2 of SEQ ID NO: 79, and CDR-L3 of SEQ ID NO: 95;(xvi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 64, CDR-L2 of SEQ ID NO: 80, and CDR-L3 of SEQ ID NO: 96; (xvii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 141, CDR-L2 of SEQ ID NO: 143, and CDR-L3 of SEQ ID NO: 145; and (xviii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 142, CDR-L2 of SEQ ID NO: 144, and CDR-L3 of SEQ ID NO: 146.

[0151] In certain embodiments, the polynucleotide comprises a nucleotide sequence encoding a VL domain, wherein the VL domain is: (xix) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 170, and CDR-L3 of SEQ ID NO: 180; (xx) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 181; (xxi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 160, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 182. (xxii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183; (xxiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 171, and CDR-L3 of SEQ ID NO: 184; (xxiv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 173, and CDR-L3 of SEQ ID NO: 185; (xxv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 173, and (xxvi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 161, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 187; (xxvii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 162, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 188; (xxviii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 163, CDR-L2 of SEQ ID NO: 174, and CDR-L3 of SEQ ID NO: 188; (xxix) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 164, -A light chain variable domain comprising CDR-L1 of SEQ ID NO: 165, CDR-L2 of SEQ ID NO: 175, and CDR-L3 of SEQ ID NO: 190; (xxxi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 166, CDR-L2 of SEQ ID NO: 176, and CDR-L3 of SEQ ID NO: 191; (xxxi) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 167, CDR-L2 of SEQ ID NO: 177, and CDR-L3 of SEQ ID NO: 192;(xxxii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 168, CDR-L2 of SEQ ID NO: 178, and CDR-L3 of SEQ ID NO: 193; (xxxiii) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 169, CDR-L2 of SEQ ID NO: 179, and CDR-L3 of SEQ ID NO: 194;

[0152] In one embodiment, the polynucleotide comprises a nucleotide sequence encoding a VL domain, wherein the VL domain comprises: (xxxiv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183; (xxxv) a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 195.

[0153] In certain embodiments, an isolated polynucleotide encoding a MuSK antibody-based molecule encodes any one of the VH and / or VL domain sequences provided below in Table 3. The nucleic acid molecules described herein include isolated polynucleotides, portions of expression vectors, or portions of linear DNA sequences, including linear DNA sequences used for in vitro transcription / translation, as well as vectors suitable for prokaryotic, eukaryotic, or filamentous phage expression, secretion, and / or display of the antibodies or binding fragments thereof described herein.

[0154] In a preferred embodiment, the polynucleotide comprises a nucleotide sequence encoding a VH comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234. In another preferred embodiment, the polynucleotide comprises a nucleotide sequence encoding a VL comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO:235.

[0155] In a preferred embodiment, the polynucleotide comprises a nucleotide sequence encoding an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), the molecule comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0156] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0157] In a more preferred embodiment, the polynucleotide comprises a nucleotide sequence encoding an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0158] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0159] In a more preferred embodiment, the polynucleotide comprises or consists of a nucleotide sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 264. In an even more preferred embodiment, the polynucleotide comprises or consists of SEQ ID NO:264.

[0160] In another more preferred embodiment, the polynucleotide comprises or consists of a nucleotide sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 265. In an even more preferred embodiment, the polynucleotide comprises or consists of SEQ ID NO:265.

[0161] In a more preferred embodiment, the polynucleotide comprises a nucleotide sequence encoding an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain is encoded by a nucleotide sequence that is at least 80% identical to SEQ ID NO: 264, and the light chain variable domain is encoded by a nucleotide sequence that is at least 80% identical to SEQ ID NO: 265. In certain embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0162] In one embodiment, the polynucleotide comprises: a) a nucleotide sequence that is at least 80% identical to SEQ ID NO: 276, which encodes a full-length heavy chain; and b) a nucleotide sequence that is at least 80% identical to SEQ ID NO: 278, which encodes a full-length light chain Contains :.

[0163] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0164] In one embodiment, the polynucleotide comprises: a) a nucleotide sequence at least 80% identical to SEQ ID NO: 277 encoding a heavy chain variable domain, and b) a nucleotide sequence at least 80% identical to SEQ ID NO: 279 encoding a light chain variable domain Contains :.

[0165] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0166] In one embodiment, the polynucleotide comprises: a) a nucleotide sequence at least 80% identical to SEQ ID NO:276 encoding a full-length heavy chain, which nucleotide sequence comprises a nucleotide sequence at least 80% identical to SEQ ID NO:277 encoding a heavy chain variable domain, and b) a nucleotide sequence at least 80% identical to SEQ ID NO: 278 encoding a full-length light chain, which nucleotide sequence comprises a nucleotide sequence at least 80% identical to SEQ ID NO: 279 encoding a light chain variable domain. Contains :.

[0167] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0168] In one embodiment, the polynucleotide comprises: a) the nucleotide sequence SEQ ID NO: 276; and b) Nucleotide sequence SEQ ID NO: 278 Contains :.

[0169] The polynucleotides of the present invention can be produced by chemical synthesis, for example, solid-phase polynucleotide synthesis on an automated polynucleotide synthesizer, and assembled into complete single-stranded or double-stranded molecules. Alternatively, polynucleotides can be produced by other techniques, such as PCR, and then routinely cloned. Techniques for producing or obtaining polynucleotides of a given sequence are well known in the art.

[0170] The polynucleotide may include at least one non-coding sequence, such as, for example, a promoter or enhancer sequence, an intron, a polyadenylation signal, a cis sequence facilitating RepA binding, etc. The polynucleotide sequence may also include additional sequences encoding, for example, a linker sequence, a marker or tag sequence facilitating protein purification or detection, such as a histidine tag or an HA tag, a signal sequence, a fusion protein partner, such as RepA, an Fc protein, or a bacteriophage coat protein, such as pIX or pIII.

[0171] (vector) In another aspect, there is provided a vector (preferably an expression vector) for use in treating a neuromuscular disorder in a human subject comprising a polynucleotide encoding a MuSK antibody-based molecule (or an anti-MuSK antibody or antigen-binding fragment thereof) described herein.

[0172] Such vectors include, but are not limited to, plasmid vectors, viral vectors, including but not limited to vaccinia vectors, lentiviral vectors, adenoviral vectors, adeno-associated viral vectors, vectors for baculovirus expression, transposon-based vectors, or any other vector polypeptide suitable for the introduction of the polynucleotides described herein into a given organism or genetic background by any means to facilitate expression of the encoded antibody. In one embodiment, the polynucleotides encoding the heavy chain variable domains, alone or together with the polynucleotides encoding the light chain variable domains described herein, are combined with a promoter, a translation initiation segment (e.g., a ribosome binding sequence and a start codon), a 3' untranslated region, a polyadenylation signal, a stop codon, and a sequence for transcription termination to form one or more expression vector constructs.

[0173] In one embodiment, the vector is an adenovirus-associated virus (AAV) vector. Many therapeutic AAV vectors suitable for delivery of the polynucleotide encoding the antibody described herein to the central nervous system are known in the art. See, for example, Deverman et al., "Gene Therapy for Neurological Disorders: Progress and Prospects," Nature Rev. 17:641-659 (2018), which is incorporated herein by reference in its entirety. Suitable AAV vectors include serotypes AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, or AAV11 in their native form or modified for enhanced tropism. AAV vectors known to have tropism for the CNS that are particularly suitable for therapeutic expression of the MuSK antibodies described herein include AAV1, AAV2, AAV4, AAV5, AAV8, and AAV9 in their native form or modified for enhanced tropism. In one embodiment, the AAV vector is an AAV2 vector. In another embodiment, the AAV vector is an AAV5 vector (Vitale et al., "Anti-tau Conformational scFv MC1 Antibody Efficiently Reduces Pathological Tau Species in Adult JNPL3 Mice," Acta Neuropathol. Commun. 6:82 (2018), incorporated herein by reference in its entirety).In another embodiment, the AAV vector is an AAV9 vector (Haiyan et al., "Targeting Root Cause by Systemic scAAV9-hIDS Gene Delivery: Functional Correction and Reversal of Severe MPSII in Mice," Mol. Ther. Methods Clin. Dev. 10:327-340 (2018), incorporated herein by reference in its entirety). In another embodiment, the AAV vector is an AAVrhlO vector (Liu et al., "Vectored Intracerebral Immunizations with the Anti-Tau Monoclonal Antibody PHF1 Markedly Reduces Tau Pathology in Mutant Transgenic Mice," J. Neurosci. 36(49): 12425-35(2016), incorporated herein by reference in its entirety).

[0174] In another embodiment, the AAV vector is a hybrid vector that contains the genome of one serotype, e.g., AAV2, and the capsid proteins of another serotype, e.g., AAV1 or AAV3-9, to control tropism. See, e.g., Broekman et al., "Adeno-associated Virus Vectors Serotyped with AAV8 Capsid are More Efficient than AAV-1 or -2 Serotypes for Widespread Gene Delivery to the Neonatal Mouse Brain," Neuroscience 138:501-510 (2006), incorporated herein by reference in its entirety. In one embodiment, the AAV vector is an AAV2 / 8 hybrid vector (Ising et al., AAV-mediated Expression of Anti-Tau ScFv Decreases Tau Accumulation in a Mouse Model of Tauopathy, J. Exp. Med. 214(5):1227(2017), which is incorporated by reference in its entirety). In another embodiment, the AAV vector is an AAV2 / 9 hybrid vector (Simon et al., A Rapid Gene Delivery-Based Mouse Model for Early-Stage Alzheimer Disease-Type Tauopathy, J. Neuropath. Exp. Neurol. 72(11):1062-71(2013), which is incorporated by reference in its entirety).

[0175] In another embodiment, the AAV vector is capable of enhancing its CNS delivery following intraparenchymal administration, e.g., AAV-DJ (Grimm et al., J. Viol. 82:5887-5911 (2008) which is incorporated by reference in its entirety); increasing delivery of neural stem and progenitor cells, e.g., SCH9 and AAV4.18 (Murlidharan et al., J. Virol. 89: 3976-3987 (2015) and Ojala et al., Mol. Ther. 26:304-319 (2018) which are incorporated by reference in their entireties); enhancing retrograde delivery, e.g., rAAV2-retro (Muller et al., Nat. Biotechnol. 26:304-319 (2018) which are incorporated by reference in their entireties). 21:1040-1046(2003)); selective delivery to brain endothelial cells, e.g., AAV-BRI (Korbelin et al., EMBO Mol. Med. 8: 609-625(2016) which are incorporated by reference in their entireties); or enhanced delivery of the adult CNS following IV administration, e.g., AAV-PHP.B and AAVPHP.eB (Deverman et al., Nat. Biotechnol. 34: 204-209(2016) and Chan et al., Nat. Neurosci. 20: 1172-1179(2017), which are incorporated by reference in their entireties.

[0176] According to this embodiment, the expression vector construct encoding the MuSK antibody-based molecule comprises a polynucleotide encoding a heavy chain polypeptide, a functional fragment thereof, a variant thereof, or a combination thereof. The expression construct may alternatively comprise a nucleic acid sequence encoding a light chain polypeptide, a functional fragment thereof, a variant thereof, or a combination thereof. In one embodiment, the expression vector construct comprises a nucleic acid sequence encoding a heavy chain polypeptide, a functional fragment thereof, or a variant thereof, and a light chain polypeptide, a functional fragment thereof, or a variant thereof.

[0177] In some embodiments, the expression construct further comprises a promoter sequence suitable for driving the expression of MuSK antibody-based molecules.Suitable promoter sequences include, but are not limited to, elongation factor 1-alpha promoter (EF1a) promoter, phosphoglycerate kinase-1 promoter (PGK) promoter, cytomegalovirus immediate early gene promoter (CMV), chimeric liver-specific promoter (LSP), cytomegalovirus enhancer / chicken β-actin promoter (CAG), tetracycline-responsive promoter (TRE), transthyretin promoter (TTR), simian virus 40 promoter (SV40), and CK6 promoter.Other promoters suitable for driving gene expression in mammalian cells known in the art are also suitable for incorporation into the expression constructs disclosed herein.

[0178] In certain embodiments, the expression construct (or expression vector) further encodes a linker sequence, which can encode an amino acid sequence that spatially separates and / or links one or more components of the expression construct (such as the heavy and light chain components of the encoded antibody).

[0179] In a preferred embodiment, the expression vector comprises a polynucleotide encoding an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) and comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235; wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0180] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0181] In a more preferred embodiment, the expression vector comprises nucleotides encoding an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) and comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235; wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0182] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0183] In one embodiment, the expression vector encodes an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), and a) a nucleotide sequence that is at least 80% identical to SEQ ID NO: 276, which encodes a full-length heavy chain; and b) a nucleotide sequence that is at least 80% identical to SEQ ID NO: 278, which encodes a full-length light chain :

[0184] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0185] In one embodiment, the expression vector encodes an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), and a) a nucleotide sequence at least 80% identical to SEQ ID NO: 277 encoding a heavy chain variable domain, and b) a nucleotide sequence at least 80% identical to SEQ ID NO: 279 encoding a light chain variable domain :

[0186] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0187] In one embodiment, the expression vector encodes an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), c) a nucleotide sequence at least 80% identical to SEQ ID NO:276 encoding a full-length heavy chain, which nucleotide sequence comprises a nucleotide sequence at least 80% identical to SEQ ID NO:277 encoding a heavy chain variable domain, and d) a nucleotide sequence at least 80% identical to SEQ ID NO: 278 encoding a full-length light chain, wherein the nucleotide sequence comprises a nucleotide sequence at least 80% identical to SEQ ID NO: 279 encoding a light chain variable domain. and includes a polynucleotide comprising:

[0188] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0189] In one embodiment, the expression vector encodes an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK) and: a) the nucleotide sequence of SEQ ID NO: 276; and b) the nucleotide sequence of SEQ ID NO: 278 The polynucleotide comprises:

[0190] (host cell) Another aspect of the invention is a host cell or cell-free expression system for use in treating a neuromuscular disease in a human subject, wherein the cell contains an expression vector encoding a MuSK antibody (or an antigen-binding fragment thereof) and, optionally, producing the MuSK antibody described herein.

[0191] The MuSK antibody-based molecules described herein can optionally be produced by cell lines, mixed cell lines, immortalized cells, or clonal populations of immortalized cells, as is known in the art (see, e.g., Ausubel et al., eds., Current Protocols in Molecular Biology, John Wiley & Sons, Inc., NY, NY (1987-2001); Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Edition, Cold Spring Harbor, NY (1989); Harlow and Lane, Antibodies, a Laboratory Manual, Cold Spring Harbor, NY (1989); Colligan et al., eds., Current Protocols in Immunology, John Wiley & Sons, Inc., NY (1994-2001); Colligan et al., Current Protocols in Protein Science, John Wiley & Sons, Inc., NY (2002), all of which are incorporated by reference in their entireties. (See New York, NY, (1997-2001)).

[0192] In some embodiments, the host cells selected for expression may be of mammalian origin. Suitable mammalian host cells include, but are not limited to, COS-1 cells, COS-7 cells, HEK293 cells, BHK21 cells, CHO cells, BSC-1 cells, HeG2 cells, SP2 / 0 cells, HeLa cells, mammalian myeloma cells, mammalian lymphoma cells, or any derivative, immortalized, or transformed cells thereof. Other suitable host cells include, but are not limited to, yeast cells, insect cells, and plant cells. Alternatively, the host cell can be selected from a species or organism that is unable to glycosylate polypeptides, e.g., a prokaryotic cell or organism, such as BL21, BL21(DE3), BL21-GOLD(DE3), XL1-Blue, JM109, HMS174, HMS174(DE3), and any of the natural or artificial strains of E. coli species, Klebsiella species, or Pseudomonas species.

[0193] The MuSK antibody-based molecules described herein can be prepared by any of a variety of techniques using the isolated polynucleotides, vectors, and host cells described above. In general, antibodies can be produced by cell culture techniques, including the generation of monoclonal antibodies by conventional techniques, or by transfection of antibody genes, heavy chains, and / or light chains into a suitable bacterial or mammalian cell host to allow the production of antibodies, which may be recombinant. In some embodiments, the MuSK antibody-based molecules described herein are monoclonal antibodies or functional binding fragments thereof. Standard molecular biology techniques are used to prepare recombinant expression vectors, transfect host cells, select for transformants, culture host cells, and recover antibodies from the culture medium. Transfecting host cells can be performed using a variety of techniques commonly used for the introduction of foreign DNA into prokaryotic or eukaryotic host cells, such as electroporation, calcium phosphate precipitation, DEAE-dextran transfection, etc. Although it is possible to express the antibodies described herein in either prokaryotic or eukaryotic host cells, expression of the antibodies in eukaryotic cells, particularly mammalian cells, may be preferred because such eukaryotic cells, particularly mammalian cells, are more likely than prokaryotic cells to assemble and secrete a properly folded and immunologically active antibody.

[0194] As mentioned above, exemplary mammalian host cells for expressing the recombinant antibodies of the invention include Chinese hamster ovary (CHO cells) (including dhfr-CHO cells described in Urlaub and Chasin, Proc. Natl. Acad. Sci. USA, 77: 4216-4220 (1980), which is incorporated by reference in its entirety). Other suitable mammalian host cells include, but are not limited to, NS0 myeloma cells, COS cells, and SP2 cells. When a recombinant expression vector encoding an antibody gene is introduced into a mammalian host cell, the antibody is produced by culturing the host cell for a period of time sufficient to allow expression of the antibody in the host cell, or more preferably, secretion of the antibody into the culture medium in which the host cell is grown.

[0195] Host cells can also be used to produce functional antibody fragments, such as Fab fragments or scFv molecules. It is understood that variations on the above procedure are within the scope of the invention. For example, it may be desirable to transfect host cells with DNA encoding functional fragments of either the light and / or heavy chains of the antibodies described herein. Recombinant DNA technology can also be used to remove some or all of the DNA encoding either or both of the light and heavy chains that is not necessary for binding to the antigen of interest. Molecules expressed from such truncated DNA molecules are also encompassed by the antibodies described herein.

[0196] Antibodies and antibody binding fragments are recovered and purified from recombinant cell cultures by known methods, including, but not limited to, Protein A purification, ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxylapatite chromatography, and lectin chromatography. High performance liquid chromatography (HPLC) can also be used for purification.

[0197] In a preferred embodiment, the host cell expresses an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0198] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0199] In a more preferred embodiment, the host cell expresses an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0200] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0201] In one embodiment, the host cell comprises: a) a nucleotide sequence that is at least 80% identical to SEQ ID NO: 276, which encodes a full-length heavy chain; and b) a nucleotide sequence that is at least 80% identical to SEQ ID NO: 278, which encodes a full-length light chain The present invention expresses an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), including:

[0202] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0203] In one embodiment, the host cell comprises: a) a nucleotide sequence at least 80% identical to SEQ ID NO: 277 encoding a heavy chain variable domain, and b) a nucleotide sequence at least 80% identical to SEQ ID NO: 279 encoding a light chain variable domain The present invention expresses an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), including:

[0204] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0205] In one embodiment, the host cell comprises: a) a nucleotide sequence at least 80% identical to SEQ ID NO:276 encoding a full-length heavy chain, which nucleotide sequence comprises a nucleotide sequence at least 80% identical to SEQ ID NO:277 encoding a heavy chain variable domain, and b) a nucleotide sequence at least 80% identical to SEQ ID NO: 278 encoding a full-length light chain, which nucleotide sequence comprises a nucleotide sequence at least 80% identical to SEQ ID NO: 279 encoding a light chain variable domain. The present invention expresses an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), including:

[0206] In some embodiments, the identity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0207] In one embodiment, the host cell comprises: a) the nucleotide sequence of SEQ ID NO: 276, and b) the nucleotide sequence of SEQ ID NO: 278 The present invention expresses an antibody-based molecule that binds to human muscle-specific tyrosine protein kinase (MuSK), including:

[0208] Compositions Comprising MuSK Antibody-Based Molecules The MuSK antibody-based molecules or polynucleotides encoding the MuSK antibody-based molecules of the present invention can be advantageously administered as a composition.

[0209] Therefore, in a further aspect, there is provided a composition for use in treating a neuromuscular disorder in a human subject, the composition comprising an antibody or antigen-binding fragment, a polynucleotide, an expression vector, or a host cell or cell-free expression system as defined herein.

[0210] In certain embodiments, the composition is a pharmaceutical composition. In certain embodiments, the pharmaceutical composition comprises at least one pharma- ceutically acceptable carrier or excipient.

[0211] In certain embodiments, such compositions are pharmaceutical compositions that include an active therapeutic agent (i.e., a MuSK antibody) and one or more of a variety of other pharma- ceutically acceptable ingredients. See REMINGTON: THE SCIENCE AND PRACTICE OF PHARMACY (21st ed.) (2005) (Troy, DB et al. (eds.) Lippincott Williams & Wilkins (Publs.), Baltimore MD), which is incorporated herein by reference in its entirety. The preferred form depends on the intended mode of administration and therapeutic application. Depending on the desired formulation, the composition may also include pharma- ceutically acceptable non-toxic carriers, excipients, diluents, fillers, salts, buffers, detergents (e.g., non-ionic detergents such as Tween-20 or Tween-80), stabilizers (e.g., sugars or protein-free amino acids), preservatives, tissue fixatives, solubilizers, and / or other materials that are suitable for inclusion in pharmaceutical compositions and are vehicles commonly used to formulate pharmaceutical compositions for animal or human administration. The diluents are selected so as not to affect the biological activity of the combination. Examples of such diluents include distilled water, physiological phosphate-buffered saline, Ringer's solution, dextrose solution, and Hank's solution. In addition, the pharmaceutical composition or formulation may also include other carriers, or non-toxic, non-therapeutic, non-immunogenic stabilizers, and the like. Examples of suitable aqueous and non-aqueous carriers that may be utilized in the pharmaceutical compositions of the present invention include water, saline, phosphate buffered saline, ethanol, dextrose, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, and the like) and suitable mixtures thereof, vegetable oils such as olive oil, corn oil, peanut oil, cottonseed oil, and sesame oil, carboxymethylcellulose colloidal solutions, tragacanth gum, and injectable organic esters such as ethyl oleate, and / or various buffers. Other carriers are well known in the pharmaceutical art.

[0212] Pharmaceutically acceptable carriers include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions.The use of such media and agents for pharmaceutical active substances is known in the art.Except insofar as any conventional media or agent is incompatible with the active compound, its use in the pharmaceutical compositions of the present invention is contemplated.

[0213] The compositions may also include large, slowly metabolized macromolecules such as proteins, polysaccharides such as chitosan, polylactic acid, polyglycolic acid, and copolymers (e.g., latex-functionalized sepharose, agarose, cellulose, etc.), polymeric amino acids, amino acid copolymers, and lipid aggregates (e.g., oil droplets or liposomes). Suitability for carriers and other components of pharmaceutical compositions is determined based on the lack of significant negative effect on the desired biological properties of the active antibody-based molecules of the invention (e.g., less than substantially affecting antigen binding (e.g., 10% or less relative inhibition, 5% or less relative inhibition, etc.)).

[0214] The pharmaceutical compositions of the present invention may also contain pharma- ceutically acceptable antioxidants, such as, for example, (1) water-soluble antioxidants, such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite, and the like; (2) oil-soluble antioxidants, such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, α-tocopherol, and the like; and (3) metal chelators, such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like.

[0215] The pharmaceutical compositions of the present invention may also contain isotonic agents, such as sugars, polyalcohols such as mannitol, sorbitol, glycerol, or sodium chloride in the composition.

[0216] The pharmaceutical composition of the present invention may also contain one or more adjuvants appropriate for the selected route of administration, such as preservatives, wetting agents, emulsifying agents, dispersing agents, preservatives, or buffers, which may enhance the shelf life or effectiveness of the pharmaceutical composition. The antibodies of the present invention may be prepared with a carrier that will protect the antibody against rapid release, such as a controlled release formulation, including implants, transdermal patches, and microencapsulated delivery systems. Such carriers may include gelatin, glyceryl monostearate, glyceryl distearate, biodegradable biocompatible polymers, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid, alone or in combination with waxes, or other materials known in the art. Methods for preparing such formulations are generally known to those skilled in the art. See, for example, SUSTAINED AND CONTROLLED RELEASE DRUG DELIVERY SYSTEMS, edited by JR Robinson, Marcel Dekker, Inc., New York, 1978.

[0217] In one embodiment, the antibody of the present invention can be formulated to ensure proper distribution in vivo. Pharmaceutically acceptable carriers for parenteral administration include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. The use of such media and agents for pharmaceutical active substances is known in the art.

[0218] Injectable pharmaceutical compositions must be generally sterile and stable under the conditions of manufacture and storage. The composition can be formulated as a solution, microemulsion, liposome, or other ordered structure suitable for achieving high drug concentration. The carrier can be, for example, an aqueous or non-aqueous solvent or dispersion medium containing water, ethanol, polyol (for example, glycerol, propylene glycol, polyethylene glycol, etc.), and suitable mixtures thereof, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. In many cases, it is preferable to include an isotonic agent, for example, sugar, glycerol, polyalcohol such as mannitol, sorbitol, or sodium chloride in the composition. Prolonged absorption of injectable compositions can be achieved by including an agent that delays absorption, such as monostearate salts and gelatin in the composition. Sterile injectable solutions can be prepared by incorporating the required amount of active compound in a suitable solvent, with one or a combination of ingredients as listed above, as needed, and then performing sterilization microfiltration. Usually, dispersions are prepared by incorporating the active compound into a sterile vehicle that contains a basic dispersion medium and other necessary ingredients. In the case of sterile powders for preparing sterile injectable solutions, examples of preparation methods are vacuum drying and freeze-drying (lyophilization), which produce a powder of the active ingredient and any additional desired ingredients from the solution that has been previously sterile-filtered.

[0219] For parenteral administration, the agent of the present invention is usually formulated as a solution or suspension of an injectable dose of the substance in a physiologically acceptable diluent with a pharmaceutical carrier, which may be a sterile liquid such as water, oil, saline, glycerol, or ethanol. In addition, auxiliary substances, such as wetting or emulsifying agents, surfactants, pH buffering substances, and the like, may be present in the composition. Other components of the pharmaceutical composition are of petroleum, animal, vegetable, or synthetic origin. Peanut oil, soybean oil, and mineral oil are all examples of useful materials. In general, glycols, such as propylene glycol or polyethylene glycol, are preferred liquid carriers, particularly for injection solutions. The agent of the present invention can be administered in the form of a depot injection or implant preparation, which can be formulated in such a way as to allow sustained release of the active ingredient.

[0220] Usually, compositions are prepared as injections, either as liquid solutions or suspensions; solid forms suitable for dissolving or suspending in liquid vehicles before injection can also be prepared.Preparations can also be emulsified or encapsulated in liposomes or microparticles, such as polylactides, polyglycolides, or copolymers, for enhanced adjuvant effect (Langer et al., Science 249:1527 (1990); Hanes et al., Advanced Drug Delivery Reviews 28:97-119 (1997), the entirety of which is incorporated herein by reference).Additional formulations suitable for other modes of administration include oral, intranasal, and pulmonary formulations, suppositories, and transdermal applications.

[0221] In one embodiment, the composition comprises an anti-MuSK antibody (or antigen-binding fragment) comprising a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 153, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 195 (3B2g2m1).

[0222] In one embodiment, the composition comprises an anti-MuSK antibody (or antigen-binding fragment) comprising a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 153, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183 (3B2g1m1).

[0223] In one embodiment, an anti-MuSK antibody (or antigen-binding fragment) comprising a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 154, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183 (3B2g1m2).

[0224] In one embodiment, an anti-MuSK antibody (or antigen-binding fragment) comprising a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 154, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 195 (3B2g2m2).

[0225] In one embodiment, an anti-MuSK antibody (or antigen-binding fragment) comprising a heavy chain variable domain comprising CDR-H1 of SEQ ID NO: 147, CDR-H2 of SEQ ID NO: 150, and CDR-H3 of SEQ ID NO: 156, and a light chain variable domain (3B2) comprising CDR-L1 of SEQ ID NO: 159, CDR-L2 of SEQ ID NO: 172, and CDR-L3 of SEQ ID NO: 183.

[0226] In a preferred embodiment, the composition comprises an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK), comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0227] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0228] In a more preferred embodiment, the composition comprises an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK), comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0229] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0230] In a preferred embodiment, the composition comprises an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK) comprising a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0231] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0232] In a more preferred embodiment, the composition comprises an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK) comprising a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0233] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0234] In a preferred embodiment, the composition comprises an antibody or antigen-binding fragment that binds human muscle-specific tyrosine protein kinase (MuSK) comprising a wild-type human IgG constant Fc region (wherein a L234A and / or L235A substitution, numbered according to the EU numbering system, has been introduced into the Fc region), a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0235] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0236] In a more preferred embodiment, the composition comprises an antibody or antigen-binding fragment that binds human muscle-specific tyrosine protein kinase (MuSK) comprising a wild-type human IgG constant Fc region (wherein a L234A and / or L235A substitution, numbered according to the EU numbering system, has been introduced into the Fc region), a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0237] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0238] In one embodiment, the composition comprises: a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 268, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 269. Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; one or more of a G236R substitution; a G236K substitution; a G237A substitution; a P238A substitution; a F243L substitution; a D265A substitution; a S267E substitution; a H268A substitution; a R292P substitution; a Y300L substitution; a K322A substitution; a K322Q substitution; a A327Q substitution; a L328F substitution; a L328R substitution; a P329A substitution; a P329G substitution; a A330L substitution; a A330S substitution; a P331S substitution; a I332E substitution; a P396L substitution or each of the combinations of mutations previously described in the fourth embodiment of the present application have been introduced into the full length heavy chain, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A, The present invention includes an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK).

[0239] In one embodiment, the composition comprises: a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains : and c) wherein the full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system. The present invention includes an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK).

[0240] In certain embodiments, binding to an effector ligand is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% reduced, or no longer detectable, compared to binding to the same ligand by an antibody that does not have any amino acid substitutions in its human IgG constant Fc region.

[0241] In one embodiment, the composition comprises: a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 270, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 271 and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; one or more of a G236R substitution; a G236K substitution; a G237A substitution; a P238A substitution; a F243L substitution; a D265A substitution; a S267E substitution; a H268A substitution; a R292P substitution; a Y300L substitution; a K322A substitution; a K322Q substitution; a A327Q substitution; a L328F substitution; a L328R substitution; a P329A substitution; a P329G substitution; a A330L substitution; a A330S substitution; a P331S substitution; a I332E substitution; a P396L substitution or each of the combinations of mutations previously described in the fourth embodiment of the present application have been introduced into the full length heavy chain, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A, The present invention includes an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK).

[0242] In one embodiment, the composition comprises: a) a full length heavy chain comprising SEQ ID NO: 270, and b) a full-length light chain comprising SEQ ID NO: 271 Contains : and c) wherein the full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system. The present invention includes an antibody or antigen-binding fragment that binds to human muscle-specific tyrosine protein kinase (MuSK).

[0243] In certain embodiments, binding to an effector ligand is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% reduced, or no longer detectable, compared to binding to the same ligand by an antibody that does not have any amino acid substitutions in its human IgG constant Fc region.

[0244] The MuSK antibody-based molecules of the invention can be administered by parenteral, topical, oral, or intranasal means for therapeutic treatment. Intramuscular injection (e.g., into the arm or leg muscle) and intravenous infusion are preferred methods of administration of the molecules of the invention. In some methods, such molecules are administered as a sustained release composition or device, such as the Medipad™ device (Elan Pharm. Technologies, Dublin, Ireland). In some methods, the antibodies disclosed herein are injected directly into a particular tissue (e.g., intracranial injection).

[0245] In one embodiment, the pharmaceutical composition of the present invention is administered parenterally.The phrases "parenteral administration" and "administered parenterally" as used herein refer to modes of administration other than enteral and topical administration, usually by injection, and include epidermal, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intracranial, intraorbital, intracardiac, intradermal, intraperitoneal, intratendinous, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, intracranial, intrathoracic, epidural, and intrasternal injection, subcutaneous injection, and infusion.In one embodiment, the pharmaceutical composition is administered by intravenous or subcutaneous injection or infusion.

[0246] In some embodiments, anti-MuSK antibody or its antigen-binding fragment (or polynucleotide, expression vector, host cell, or composition) for use according to the present invention is administered in combination with an anticholinergic compound.Anticholinergic compounds are compounds that can inhibit the effect of the neurotransmitter acetylcholine at synapses or at neuroeffector junctions such as neuromuscular junctions.Preferably, anticholinergic compounds are compounds that can attenuate the activity of muscarinic acetylcholine receptors.

[0247] The anticholinergic compound can also be formulated in a composition as the anti-MuSK antibody or antigen-binding fragment. The composition of the type disclosed herein for the anti-MuSK antibody or antigen-binding fragment can also be used in a composition that includes an anticholinergic compound. The two compounds can be present in a single composition. Alternatively, they can be formulated in separate compositions.

[0248] The use of a compound that results in activating and inducing the mechanism that promotes the stability and / or repair of NMJ (neuromuscular junction) is particularly attractive for the treatment of any neuromuscular disease that affects such NMJ.In a preferred embodiment, the use of two different compounds that each activate and induce the mechanism that promotes the stability and / or repair of NMJ is even more attractive, because such combination treatment has been demonstrated to be synergistic.Therefore, this combination is highly beneficial for the treatment of neuromuscular disease, particularly neuromuscular disease or disorder that affects NMJ, such as ALS.

[0249] In one embodiment, an anti-MuSK antibody or antigen-binding fragment thereof, polynucleotide, expression vector, host cell, or composition for use according to any of the preceding claims, wherein the neuromuscular disorder is characterized by impaired neuromuscular transmission and / or NMJ denervation.

[0250] Disorders of neuromuscular transmission include: A muscarinic hyperexcitability b motor neuron death, c. NMJ denervation, and D. Impairment of neurotransmission It can be characterized by at least one of:

[0251] In certain embodiments, the impaired neuromuscular transmission or impaired neurotransmission can be characterized by a defect in MuSK signaling, a defect in MuSK dimerization, a defect in MuSK phosphorylation, a defect in MuSK signaling, and / or a defect in acetylcholine receptor clustering.

[0252] In certain embodiments, impaired neuromuscular transmission or impaired neurotransmission can be characterized by impaired motor performance, reduced grip strength, impaired contractile properties of muscle at the NMJ, impaired resistance to muscle fatigue, or reduced muscle mass.

[0253] In certain embodiments, the neuromuscular disorder is analyzed or assessed or diagnosed by electrophysiological assessment; pharmacodynamic assessment; levels of neurofilament (e.g., neurofilament light chain (NFL)) in blood serum, plasma, and / or cerebrospinal fluid (CSF); or a NMJ biopsy.

[0254] The neuromuscular disorder may be selected from the group consisting of amyotrophic lateral sclerosis (ALS), spinal muscular atrophy (SMA), myasthenia gravis (MG), congenital myasthenia, Lambert-Eaton myasthenic syndrome (LEMS), Lyme disease, polio, post-polio, heavy metal poisoning, Kennedy syndrome, adult-onset Tay-Sachs disease, hereditary spastic paraplegia, multifocal neuropathy, cervical spondylosis, extramedullary tumors with compressive radiculopathy and myelopathy, inclusion body myositis, progressive bulbar palsy, progressive muscular atrophy, motor neuron syndrome, and thyrotoxic myopathy. A preferred neuromuscular disorder is ALS.

[0255] In some embodiments, the anti-MuSK antibody or antigen-binding fragment thereof defined herein can be administered to asymptomatic ALS subjects. That is, such an antibody or antigen-binding fragment can be administered before the onset of ALS in the subject. The same is true for other neuromuscular diseases.

[0256] In this context, an asymptomatic ALS subject may be a subject who has been diagnosed as having a predisposition to developing a neuromuscular disorder or disease, such as ALS. Identifying an individual (or subject) with a neuromuscular disorder may mean performing the identification using a diagnostic method. Such a subject may be a symptomatic subject diagnosed at or after disease onset, or a subject predisposed to developing a neuromuscular disorder or disease (i.e., an asymptomatic subject diagnosed before disease onset, which is synonymous with presymptomatic disease).

[0257] Neuromuscular disorders can be caused by genetic defects. Genetic defects are caused in whole or in part by changes in genomic DNA sequence compared with the genomic DNA sequence of corresponding individuals or subjects who do not suffer from the genetic defect. Genetic defects can be caused by mutations in one gene (simple disorders), by mutations in multiple genes (multifactorial genetic disorders), by a combination of genetic mutations and environmental factors, or by chromosomal damage (changes in the overall number or structure of chromosomes, the structures that carry genes). Types of genetic mutations include base substitutions, deletions, and insertions.

[0258] In an embodiment, a human subject is identified as having (or being predisposed to develop) a neuromuscular disease caused by a genetic defect. In one embodiment, the neuromuscular disease is ALS, and the genetic defect is of the SOD1 gene. Individuals or human subjects predisposed to develop ALS include those who have one or more risk factors for developing ALS, including aging, medical or family history, or having a genetic predisposition to one or more SOD-1 related diseases. One of the underlying genetic causes or predispositions for ALS is a mutation in the human SOD1 gene. Therefore, identification of subjects suffering from or susceptible to ALS or predisposed to developing ALS can be performed by genetic testing of the subject's SOD1 gene using assays known in the art, such as, for example, gene sequencing. It is known in the art that at least 180 mutations in human SOD1 are associated with ALS. In certain embodiments, the SOD1 mutations are A4V, H46R, G93S, A4T, G141X, D133A, V148G, N139K, G85R, G93A, V14G, C6S, I113T, D49K, G37R, A89V, E100G, D90A, T137A, E100K, G41A, G41D, G41S, G13R, G72S, L8V, F20C, Q22L, H48R, T54R, S591, V87A, T88deltaTAD, A89T, V97M, S105deltaSL, VI 18L, D124G, LI The mutation is one or more selected from the group consisting of: 14F, D90A, G12R, G147R, and G37R. In one embodiment, the mutation in the SOD1 gene is G37R.

[0259] Thus, an asymptomatic individual or subject may be one having any of the following HIV-infected individuals or subjects: A4V, H46R, G93S, A4T, G141X, D133A, V148G, N139K, G85R, G93A, V14G, C6S, I113T, D49K, G37R, A89V, E100G, D90A, T137A, E100K, G41A, G41D, G41S, G13R, G72S, L8V, F20C, Q22L, H48R, T54R, S591, V87A, T88deltaTAD, A89T, V97M, S105deltaSL, VI 18L, D124G, LI It can be identified (prior to disease onset) if the individual has one SOD1 mutation selected from the group consisting of: 14F, D90A, G12R, G147R, and G37R.

[0260] Analysis of a subject's susceptibility to ALS (i.e., an asymptomatic subject who is prone to developing ALS) can be performed by analyzing the subject's family history for ALS. Family history analysis can include a three-generation pedigree documenting ALS, review of medical records and autopsy studies of family members, and identification of an autosomal dominant pattern of SOD1 mutations.

[0261] Identification of individuals or subjects who are asymptomatic for ALS (but predisposed to developing such disease) can also be analyzed by ALS markers. For example, ALS-specific markers include circulating micro-RNA, circular RNA (circRNA), or messenger RNA (mRNA), TDP-42 aggregates, 8-oxo-deoxyguanosine (8-oxodG), 15-F2t-isoprostane (IsoP), plasma TNF-a, IL-10, TRAIL, plasma IL-1b, CSF These may include TRAIL, proinflammatory T-helper (Th) 17 cells, Th1 cells, anti-inflammatory Th2, regulatory T cells (Treg), proinflammatory IL-1b, IL-6, IFN-g, anti-inflammatory IL-10, cholesterol, LDL-cholesterol, apolipoprotein B, HDL-cholesterol, apolipoprotein-AI, plasma creatinine (PCr), plasma ferritin, transferrin, hepcidin, chitotriosidase-1 (CHIT1), chitinase-3-like protein 2 (CHI3L2 / YKL39), total tau (tTau), phosphorylated tau (pTau), amyloid b (Ab), novel INHAT inhibitor (NIR), ubiquitin C-terminal hydrolase-L1 (UCHL1), microtubule-associated protein 2, capping actin protein, gelsolin-like (CAPG), or glycoprotein nonmetastatic melanoma protein B (GPNMB). A human subject can be considered to be susceptible to ALS if at least one of the measurements of such markers deviates from the measurements of normal people of the same age but without ALS by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%. The analysis of the subject's susceptibility to ALS disease can also be analyzed by imaging. For example, such imaging analysis can be MRI evaluation of skeletal muscle, muscle function score obtained from the image, or prediction of bulbar palsy progression by tongue ultrasound examination combined with or without MRI.

[0262] In certain embodiments, the anticholinergic compound is administered separately, sequentially, or simultaneously with the anti-MuSK antibody or antigen-binding fragment thereof, polynucleotide, expression vector, host cell, cell-free expression system, or composition.

[0263] In some embodiments, the anticholinergic compound is administered at the onset of disease or within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 days, or within 1, 2, 3, 4, 5, 6, 7 weeks; or within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months. In some embodiments, the anticholinergic compound is administered at the onset of disease or within 1 week after the onset of disease. Surprisingly, attractive results were obtained when the anticholinergic compound was administered at the onset of disease or as soon as possible after the onset of disease. It should be understood by those skilled in the art that the anticholinergic compound is preferably not used to relieve or reduce symptoms associated with neuromuscular disorders (e.g., ALS). In some embodiments, the anticholinergic compound is not used to relieve or reduce urinary urgency. In certain embodiments, anticholinergic compounds are not used to relieve or reduce urinary urgency in neuromuscular diseases such as ALS.

[0264] In certain embodiments, the anticholinergic compound is administered at the onset of the disease or within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 days, within 1, 2, 3, 4, 5, 6, or 7 weeks, or within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after onset of the disease, but prior to diagnosis of the disease. In preferred embodiments, administration is within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 days of diagnosis; within 1, 2, 3, 4, 5, 6, or 7 weeks prior to diagnosis; or within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months prior to diagnosis.

[0265] In some embodiments, anti-MuSK antibody or its antigen-binding fragment, polynucleotide, expression vector, host cell, or composition is administered before the onset of disease or at the onset of disease.Before the onset of disease can mean 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 days before the onset of disease, or 1, 2, 3, 4, 5, 6, 7, 8 weeks before the onset of disease, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 months before the onset of disease.In this context, pre-onset human subject can mean that human subject is asymptomatic for said neuromuscular disease, for example, ALS.

[0266] Therefore, in another aspect of the present invention, there is provided an anti-MuSK antibody or its antigen-binding fragment (polynucleotide, expression vector, host cell, cell-free expression system, or composition) for use in treating ALS in a human subject, wherein the antibody or antigen-binding fragment is administered before the onset of disease, preferably within 1, 2, 3, 4, 5, or 6 months before the onset of disease.In this context, a pre-symptomatic human subject may mean that the human subject is asymptomatic to ALS.In some embodiments, the subject is diagnosed as having a predisposition to develop a neuromuscular disorder or disease, such as ALS.

[0267] In certain embodiments, the antibody or antigen-binding fragment binds to the MuSK Frizzled (Fz)-like domain sequence of SEQ ID NO:129.

[0268] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a wild-type human IgG constant Fc region comprising at least 80% sequence identity to SEQ ID NO: 266 or 267.

[0269] In certain embodiments, the antibody or antigen-binding fragment is an agonist MuSK antibody and / or has reduced or eliminated effector function.

[0270] In certain embodiments, the reduced or eliminated effector function is due to the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; or P396L substitutions into the human IgG constant Fc region (SEQ ID NO: 266 or SEQ ID NO: 267) of the antibody-based molecule described herein.

[0271] In a preferred embodiment, the L234A or L235A substitution is introduced into the human IgG constant Fc region of the antibody-based molecules described herein. In a more preferred embodiment, the L234A and L235A substitutions are introduced into the human IgG constant Fc region of the antibody-based molecules described herein. This embodiment results in an antibody-based molecule having a heavy chain represented by SEQ ID NO: 268 or 270.

[0272] In one embodiment, the antibody or antigen-binding fragment comprises: a) a heavy chain variable domain (VH) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234; and b) a light chain variable domain (VL) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235 Contains :.

[0273] In certain embodiments, the antibody or antigen-binding fragment comprises a heavy chain variable domain (VH) and a light chain variable domain (VL): wherein the VH is - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the VL is - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0274] In certain embodiments, the antibody or antigen-binding fragment comprises a heavy chain variable domain (VH) and a light chain variable domain (VL): -wherein the VH comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the VL comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the VH is a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147, a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 153, and a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the VL is a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 172, and a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0275] In one embodiment, the antibody or antigen-binding fragment comprises: a) a heavy chain variable domain (VH) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234; and b) a light chain variable domain (VL) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235 Contains :.

[0276] In one embodiment, the antibody or antigen-binding fragment comprises: a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0277] Surprisingly, when anti-MuSK antibody (or antigen-binding fragment or host cell polynucleotide or expression vector) is administered before the onset of disease, attractive results are obtained. In some embodiments, the human subject is therefore asymptomatic for said neuromuscular disorder, for example, ALS. In such embodiments, the human subject has been diagnosed as susceptible to developing a neuromuscular disorder such as ALS, taking into account its family history, genetic background, or taking into account an increase in the level of neurofilament (e.g., neurofilament light chain (NFL)) determined in its blood serum or cerebrospinal fluid (CSF), or taking into account a positive genetic test for ALS-associated gene mutation, or taking into account changes in the level of ALS biomarkers, or from a combination thereof. However, the person has not yet developed visible symptoms; he is asymptomatic. In a preferred embodiment, the human subject is administered as early as possible after being diagnosed with a genetic defect or ALS-associated gene mutation, when he has not yet developed visible symptoms (i.e., asymptomatic subject). In a more preferred embodiment, the human subject is administered as soon as possible after being diagnosed with genetic defect or ALS-related gene mutation and before it develops visible symptoms, and the human subject has a family history of ALS.In this context, "immediately" can mean within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 hours, or within 1, 2, 3, 4, 5, 6, or 7 days, or within 1, 2, 3, or 4 weeks.In this context, such treatment shows only limited or no toxicity and / or side effects or adverse effects when the diagnosis of ALS is not confirmed.

[0278] In another aspect of the invention, there is provided a combination comprising an anti-MuSK antibody, or antigen-binding fragment thereof, as described herein and an anticholinergic compound, preferably for use in the treatment of a neuromuscular disease in a human subject.

[0279] The neuromuscular disorder is characterized by impaired neuromuscular transmission and / or denervation of the NMJ (neuromuscular junction). A muscarinic hyperexcitability b motor neuron death, c. Neuromuscular junction (NMJ) denervation, and D. Impairment of neurotransmission Characterized by at least one of:

[0280] In some embodiments, the neuromuscular disorder is selected from the group consisting of amyotrophic lateral sclerosis (ALS), spinal muscular atrophy (SMA), myasthenia gravis (MG), congenital myasthenia, Lambert-Eaton myasthenic syndrome (LEMS), Lyme disease, polio, post-polio, heavy metal poisoning, Kennedy syndrome, adult-onset Tay-Sachs disease, hereditary spastic paraplegia, multifocal neuropathy, cervical spondylosis, extramedullary tumors with compressive radiculopathy and myelopathy, inclusion body myositis, progressive bulbar palsy, progressive muscular atrophy, motor neuron syndrome, and thyrotoxic myopathy. In a preferred embodiment, the neuromuscular disease is ALS.

[0281] In this context, combination does not require that the anti-MuSK antibody, or antigen-binding fragment thereof, described herein and the anticholinergic compound be physically present together in one composition.

[0282] In some embodiments, the anticholinergic compounds are administered separately, sequentially, or simultaneously. In some embodiments, the antibody or antigen-binding fragment is administered before the onset of disease, or within 1, 2, 3, 4, 5, or 6 months before the onset of disease. In some embodiments, the antibody or antigen-binding fragment is administered before the onset of disease, or preferably within 1, 2, 3, 4, 5, or 6 months before the onset of disease, and / or wherein the anticholinergic compound is administered at the onset of disease, or within 1, 2, 3, 4, 5, 6, or 7 weeks after the onset of disease. In this context, a pre-symptomatic human subject may mean that the human subject is asymptomatic for the neuromuscular disorder. In some embodiments, the subject treated with the antibody was initially diagnosed as having a predisposition to develop a neuromuscular disorder or disease.

[0283] In certain embodiments, the anti-MuSK antibody, or antigen-binding fragment thereof, binds to the MuSK Frizzled (Fz)-like domain sequence of SEQ ID NO:129.

[0284] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a wild-type human IgG constant Fc region comprising at least 80% sequence identity to SEQ ID NO: 266 or 267.

[0285] In certain embodiments, the antibody or antigen-binding fragment is an agonist MuSK antibody and / or has reduced or eliminated effector function.

[0286] In certain embodiments, the reduced or eliminated effector function is due to the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; or P396L substitution into the human IgG constant Fc region of SEQ ID NO:266 or SEQ ID NO:267 of the antibody-based molecule described herein.

[0287] In a preferred embodiment, the L234A or L235A substitution is introduced into the human IgG constant Fc region of the antibody-based molecules described herein. In a more preferred embodiment, the L234A and L235A substitutions are introduced into the human IgG constant Fc region of the antibody-based molecules described herein. This embodiment results in an antibody-based molecule having a heavy chain represented by SEQ ID NO: 268 or 270.

[0288] In one embodiment, the antibody or antigen-binding fragment comprises: a) a heavy chain variable domain (VH) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234; and b) a light chain variable domain (VL) comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235 Contains :.

[0289] In certain embodiments, the antibody or antigen-binding fragment comprises a heavy chain variable domain (VH) and a light chain variable domain (VL): wherein the VH is - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the VL is - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0290] In certain embodiments, the antibody or antigen-binding fragment comprises a heavy chain variable domain (VH) and a light chain variable domain (VL): -wherein the VH comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the VL comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the VH is a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147, a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 153, and a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the VL is a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 172, and a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0291] In one embodiment, the antibody or antigen-binding fragment comprises: a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0292] In one embodiment, the antibody or antigen-binding fragment comprises: a) a full length heavy chain comprising SEQ ID NO: 270, and b) a full-length light chain comprising SEQ ID NO: 271 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0293] In certain embodiments, the anti-MuSK antibody or antigenic fragment thereof (polynucleotide, expression vector, host cell, cell-free expression system, or composition) is administered prior to the onset of disease (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 days prior to the onset of disease or 1, 2, 3, 4, 5, 6, or 7 weeks prior to the onset of disease) and the anticholinergic compound is administered at the time of disease onset (e.g., within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 days or within 1, 2, 3, 4, 5, 6, or 7 weeks after the onset of disease). In this context, a pre-symptomatic human subject can mean that the human subject is asymptomatic to the neuromuscular disorder, e.g., ALS.

[0294] In a preferred embodiment, the disease manifestations include at least one of the symptoms selected from the group consisting of: muscle contractions, muscle spasms, spasticity, muscle weakness, slurred speech and / or nasal voice, difficulty chewing or swallowing, dysphagia, dysarthria, and difficulty breathing. In a more preferred embodiment, the disease is ALS and the disease manifestations include at least one of the symptoms selected from the group consisting of: muscle contractions, muscle spasms, spasticity, muscle weakness, slurred speech and / or nasal voice, difficulty chewing or swallowing, dysphagia, dysarthria, and difficulty breathing.

[0295] Disease onset can be assessed by a physician or veterinarian, hi some embodiments, the onset of weight loss is considered disease onset.

[0296] In a preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0297] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0298] In a more preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0299] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0300] In a preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO:266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0301] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0302] In a more preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO:266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0303] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0304] In a preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region (wherein a L234A and / or a L235A substitution, numbered according to the EU numbering system, has been introduced into the Fc region), a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0305] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0306] In a more preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region (wherein a L234A and / or a L235A substitution, numbered according to the EU numbering system, has been introduced into the Fc region), a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0307] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0308] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 268, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 269. Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions are introduced into the full length heavy chain: G236R; G236K; G237A; P238A; F243L; D265A; S267E; H268A; R292P; Y300L; K322A; K322Q; A327Q; L328F; L328R; P329A; P329G; A330L; A330S; P331S; I332E; P396L or each of the combinations of mutations previously described in the fourth embodiment of the present application, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A.

[0309] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0310] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 270, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 271 Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions are introduced into the full length heavy chain: G236R; G236K; G237A; P238A; F243L; D265A; S267E; H268A; R292P; Y300L; K322A; K322Q; A327Q; L328F; L328R; P329A; P329G; A330L; A330S; P331S; I332E; P396L or each of the combinations of mutations previously described in the fourth embodiment of the present application, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A.

[0311] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full length heavy chain comprising SEQ ID NO: 270, and b) a full-length light chain comprising SEQ ID NO: 271 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0312] In an embodiment, anticholinergic compound is a muscarinic receptor antagonist.Muscarinic receptor, also known as muscarinic acetylcholine receptor or mAchR, is an acetylcholine receptor that forms G-protein receptor complex in the cell membrane of certain neurons and cells.Muscarinic receptor plays several roles in mediating the effect of neurotransmitter acetylcholine.For example, muscarinic receptor is contained in the presynaptic membrane of somatic neurons of neuromuscular junction, where they are involved in the regulation of acetylcholine release.

[0313] Five subtypes of muscarinic receptors, M1-M5, are generally recognized. This classification is derived from their different selectivity for certain agonists and antagonists. M1, M3, and M5 receptors couple to Gq proteins in the cell membrane, whereas M2 and M4 receptors couple to Gi / o proteins in the cell membrane. Without being bound by this theory, the genes CHRM1-5 code for the M1-M5 receptors, respectively.

[0314] Basal or constitutive activity of a muscarinic receptor is defined as the physical, biological, and / or chemical activity of the receptor in the absence of acetylcholine, muscarinic receptor agonists, and muscarinic receptor antagonists.

[0315] Agonists of muscarinic receptors, also called muscarinic receptor agonists, are defined as compounds that increase the physical, biological, and / or chemical activity of the receptor when they contact the receptor, where increased activity means activity similar to that caused by contacting the receptor with acetylcholine.

[0316] Antagonists of muscarinic receptors, also called muscarinic receptor antagonists, are defined as neutral muscarinic receptor antagonists or negative muscarinic receptor antagonists.

[0317] A muscarinic receptor neutral antagonist is a compound that competes with a muscarinic receptor neutral agonist or a muscarinic receptor negative antagonist for binding to the receptor, thereby inhibiting the action of the agonist or negative antagonist (i.e., increasing or decreasing activity), but which does not significantly alter the basal activity of the receptor upon binding alone.

[0318] In an embodiment, the anticholinergic compound is a neutral muscarinic receptor antagonist.

[0319] A muscarinic receptor negative antagonist is a compound that, when it contacts a receptor, reduces the physical, biological, and / or chemical activity of the receptor, even in the absence of a muscarinic receptor agonist, where reduced activity means activity opposite to that caused by contacting the receptor with acetylcholine.

[0320] In an embodiment, the anticholinergic compound is a muscarinic receptor negative antagonist.

[0321] A muscarinic receptor antagonist is defined as selective for one or more muscarinic receptor subtypes M1, M2, M3, M4, and / or M5 if the effect of the antagonist (blocking an agonist, blocking a negative antagonist, or decreasing activity) is only noticeable when contacting one or more muscarinic receptor subtypes, while the effect is significantly less, or not at all, when contacting muscarinic receptors of another subtype. Thus, when a muscarinic receptor antagonist is said to be selective for muscarinic receptor M3, it is understood that contacting the antagonist with muscarinic receptors of subtypes M1, M2, M4, or M5 will result in a significantly lesser effect or no effect at all. In this context, significantly less means at least 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 10000, 100000, or 1,000,000 times less, or at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 120%, 140%, 160%, 180%, 200%, 220%, 240%, 260%, 280%, 300%, 320%, 340%, 360%, 380%, 400%, 420%, 440%, 460%, 480%, 500%, 600%, 700%, 800%, 900%, 1000%, 1500%, 2000%, 2500%, 3000%, 3500%, 4000%, 4500%, 5000%, 5500%, 6000%, 6500%, 7000%, 7500%, 8000%, 8500%, 9000%, 9500%, 10000%, 20000%, 30000%, 40000%, 50000%, 60000%, 70000%, 80000%, 90000%, 100000%, 1000000%, 10000000% or 100000000% less.

[0322] Preferably, the activity of muscarinic receptors of subtypes M1, M3, and M5 can be measured using dynamic Ca2+ imaging. These receptors regulate the levels of IP3, which controls the release of Ca2+ from internal stores [7].

[0323] In an embodiment, the anticholinergic compound is - selective for the muscarinic M1 receptor, or - selective for the muscarinic M3 receptor, or - selective for the muscarinic M5 receptor, or - selective for the muscarinic M1 and M3 receptors, or - selective for the muscarinic M1 and M5 receptors, or - selective for the muscarinic M3 and M5 receptors, or -Selective for muscarinic M1, M3, and M5 receptors :It is a muscarinic receptor antagonist.

[0324] In an embodiment, the anticholinergic compound is - selective for the muscarinic M3 receptor, or - selective for the muscarinic M1 and M3 receptors, or - selective for the muscarinic M3 and M5 receptors, or -Selective for muscarinic M1, M3, and M5 receptors :It is a muscarinic receptor antagonist.

[0325] In an embodiment, the anticholinergic compound is selected from the group consisting of darifenacin, ipratropium bromide, tiotropium bromide, trospium, glycopyrronium, aclidinium, umeclidinium, solifenacin, disilomine, fesoterodine, flavoxate, glycopyrrolate, propantheline, 1R,2R,4S,5S,7S)-7-[({4-fluoro-2-(thiophen-2-yl)phenyl}carbamoyl)oxy]-9,9-dimethyl-3-oxa-9-azatricyclo[3.3.1.02,4]nonan-9-ium formate (BS46 in

[38] ), N-(2-[3-([3R]-1-(cyclohexylmethyl)-3-piperidine)-2-carboxylate, N-(2-[3-([3R]-1-(cyclohexylmethyl)-3-piperidine)-2-carboxylate), ... lysinyl]methylamino)-3-oxopropyl]amino-2-oxoethyl)-3,3,3-triphenyl-propionamide (J-115311 in

[39] ), 3,3,3-triphenylpropionamide derivatives with one or two amino acid residues between the triphenylpropionic acid moiety and the piperidinylmethylamine moiety (

[40] ), OrM3 (

[41] ), or (3R)-3-[[[(3-fluorophenyl)[(3,4,5-trifluorophenyl)methyl]amino]carbonyl]oxy]-1-[2-oxo-2-(2-thienyl)ethyl]-1-azoniabicyclo[2.2.2]octane bromide (CHF 5407 in

[41] ). Without being bound by this theory, these compounds can be considered as muscarinic receptor antagonists.

[0326] In embodiments, the anticholinergic compound is darifenacin, ipratropium bromide, tiotropium bromide, trospium, glycopyrronium, aclidinium, umeclidinium, solifenacin, disilomine, fesoterodine, flavoxate, glycopyrrolate, or propantheline. Without being bound by this theory, darifenacin, ipratropium bromide, and tiotropium bromide can be considered muscarinic receptor antagonists.

[0327] In an embodiment, the anticholinergic compound is darifenacin, ipratropium bromide, tiotropium bromide or trospium. Without being bound by this theory, darifenacin, ipratropium bromide, and tiotropium bromide can be considered as muscarinic receptor antagonists.

[0328] In a preferred embodiment, the anticholinergic compound is darifenacin, which can be represented by the following structure: [ka]

[0329] Preferably, the darifenacin is darifenacin hydrobromide. Darifenacin hydrobromide can be represented by the following structure: [ka]

[0330] In embodiments, any of the anticholinergic compounds disclosed in the above embodiments may be present as a pharma- ceutically acceptable salt thereof. In particular, the anticholinergic compound is darifenacin or a pharma- ceutically acceptable salt thereof.

[0331] Examples of pharma- ceutically acceptable salts include, but are not limited to, alkali metal (e.g., sodium, potassium, or lithium) or alkaline earth metal (e.g., calcium) salts; however, any salt that is generally non-toxic and effective when administered to the subject being treated is acceptable. Further salts include, but are not limited to, (1) acid addition salts obtained by reaction of the free base of the parent compound with inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, phosphoric acid, sulfuric acid, and perchloric acid, or with organic acids such as acetic acid, oxalic acid, (D) or (L) malic acid, maleic acid, urethanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, tartaric acid, citric acid, succinic acid, or malonic acid; or (2) salts formed when an acidic proton present in the parent compound is either replaced with a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or an aluminum ion, or coordinated with an organic base such as ethanolamine, diethanolamine, triethanolamine, trimethamine, N-methylglucamine, etc. Pharmaceutically acceptable salts are well known to those skilled in the art, and any such pharma- ceutical acceptable salt may be contemplated in connection with the embodiments described herein.

[0332] Acceptable salts can be obtained using standard procedures known in the art, including, but not limited to, reacting sufficiently acidic compounds with a suitable base to yield physiologically acceptable anions. Suitable acid addition salts are formed from acids which form non-toxic salts. Illustrative, but non-limiting, examples include acetate, aspartate, benzoate, besylate, bicarbonate / carbonate, bisulfate / sulfate, borate, camsylate, citrate, edisylate, esylate, formate, fumarate, gluceptate, gluconate, glucuronate, hexafluorophosphate, hybenzate, hydrochloride / chloride, hydrobromide / bromide, hydroiodide / iodide, lactate, mayate, maleate, maloate, mesylate, methylsulfate, naphthylate, 2-napsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate / hydrogenphosphate / dihydrogenphosphate, saccharate, stearate, succinate, tartrate, tocyiate, and trifluoroacetate. Suitable base salts of the compounds described herein are formed from bases which form non-toxic salts, illustrative examples of which include, but are not limited to, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, sodium, tromethamine, and zinc salts. Hemisalts of acids and bases may also be formed, such as hemisulfate and hemicalcium salts.

[0333] The anticholinergic compounds disclosed in the above embodiments can be administered as a composition, preferably a therapeutic composition.In an embodiment, the composition is preferably formulated as a once-a-day sustained release tablet for oral use, comprising darifenacin as darifenacin hydrobromide.Preferably, the composition comprises one or more of the following excipients: anhydrous dibasic calcium phosphate, hypromellose, magnesium stearate, titanium dioxide, yellow iron oxide, red iron oxide, PEG 400, and / or talc.In some embodiments, the composition is known as ENABLEX™.ENABLEX™ is formulated as 7.5mg or 15mg darifenacin (as darifenacin hydrobromide).

[0334] In a preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) and an anticholinergic compound is used.

[0335] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0336] In a more preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) and an anticholinergic compound is used.

[0337] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0338] In a preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO:266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0339] In a more preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region, a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO:266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO:235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0340] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0341] In a preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region (wherein a L234A and / or a L235A substitution, numbered according to the EU numbering system, has been introduced into the Fc region), a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising SEQ ID NO: 147 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 147; - a CDR-H2 amino acid sequence comprising SEQ ID NO: 153 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 153, and - a CDR-H3 amino acid sequence comprising SEQ ID NO: 156 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising SEQ ID NO: 159 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 159; - a CDR-L2 amino acid sequence comprising SEQ ID NO: 172 or having 1, 2, 3, 4 or 5 amino acid modifications compared to SEQ ID NO: 172, and - a CDR-L3 amino acid sequence comprising SEQ ID NO: 195 or having 1, 2, 3, 4, or 5 amino acid modifications compared to SEQ ID NO: 195 (3B2g2m1) Contains :.

[0342] In a more preferred embodiment, the neuromuscular disorder is ALS, and the anti-MuSK antibody or antigen-binding fragment comprises a wild-type human IgG constant Fc region (wherein a L234A and / or L235A substitution, numbered according to the EU numbering system, is introduced into the Fc region), a heavy chain variable domain, and a light chain variable domain, wherein the wild-type human IgG constant Fc region comprises SEQ ID NO: 266 or 267, a heavy chain variable domain, and a light chain variable domain, wherein the heavy chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 234, and the light chain variable domain comprises an amino acid sequence at least 80% identical or similar to SEQ ID NO: 235, and wherein the heavy chain variable domain comprises - a CDR-H1 amino acid sequence comprising or consisting of SEQ ID NO: 147, - a CDR-H2 amino acid sequence comprising or consisting of SEQ ID NO: 153, and - CDR-H3 amino acid sequence comprising or consisting of SEQ ID NO: 156 (3B2g2m1) Contains : and wherein the light chain variable domain comprises - a CDR-L1 amino acid sequence comprising or consisting of SEQ ID NO: 159, - a CDR-L2 amino acid sequence comprising or consisting of SEQ ID NO: 172, and - CDR-L3 amino acid sequence comprising or consisting of SEQ ID NO: 195 (3B2g2m1) Contains :.

[0343] In some embodiments, the identity or similarity is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%.

[0344] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 268, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 269. Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A substitution; L234D substitution; L234E substitution; L234G substitution; L234H substitution; L234F substitution; L234K substitution; L234Q substitution; L234R substitution; L234S substitution; L234T substitution; L235A substitution; L235D substitution; L235E substitution; L235F substitution; L235G substitution; L235V substitution; L235H substitution; L235I substitution; L235K substitution; L235R substitution; L235S substitution; L235T substitution; L235Q substitution; L237A substitution; S239D substitution; E233P substitution; L234V substitution; C236 deletion; G236E substitution; One or more of the following substitutions are introduced into the full length heavy chain: G236R; G236K; G237A; P238A; F243L; D265A; S267E; H268A; R292P; Y300L; K322A; K322Q; A327Q; L328F; L328R; P329A; P329G; A330L; A330S; P331S; I332E; P396L or each of the combinations of mutations previously described in the fourth embodiment of the present application, preferably the mutation is L234A or L235A, more preferably the mutation is L234A and L235A.

[0345] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full length heavy chain comprising SEQ ID NO: 268, and b) a full-length light chain comprising SEQ ID NO: 269 Contains : and c) wherein said full length heavy chain comprises the L234A and L235A mutations numbered according to the EU numbering system.

[0346] In one embodiment, the neuromuscular disorder is ALS and the anti-MuSK antibody or antigen-binding fragment is a) a full-length heavy chain comprising an amino acid sequence that is at least 80% identical or similar to SEQ ID NO: 270, and b) a full-length light chain comprising an amino acid sequence at least 80% identical or similar to SEQ ID NO: 271 Contains : and c) wherein the following mutations (all numbered according to the EU numbering system): N297A substitution; N297Q substitution; L234A subst...

Claims

1. (a) a full-length heavy chain comprising SEQ ID NO: 270 and a full-length light chain comprising SEQ ID NO: 271, or (b) a full-length heavy chain comprising SEQ ID NO: 268 and a full-length light chain comprising SEQ ID NO: 269 : An anti-MuSK antibody or an antigen-binding fragment thereof comprising:

2. A polynucleotide comprising a nucleotide sequence encoding the antibody or antigen-binding fragment thereof of claim 1.

3. 3. An expression vector comprising the polynucleotide of claim 2, preferably operably linked to a regulatory region that allows expression of the antibody or antigen-binding fragment thereof in a host cell or a cell-free expression system.

4. A host cell or a cell-free expression system containing the expression vector of claim 3.

5. A composition comprising the antibody or antigen-binding fragment thereof of claim 1, the polynucleotide of claim 2, the expression vector of claim 3, the host cell of claim 4, or the cell-free expression system of claim 4.

6. 6. The composition of claim 5, which is a pharmaceutical composition comprising at least one pharmaceutically acceptable carrier or excipient.

7. A combination comprising the anti-MuSK antibody or antigen-binding fragment thereof of claim 1 and an anticholinergic compound.

8. 8. The combination according to claim 7, wherein the anticholinergic compound is a muscarinic receptor antagonist, preferably a muscarinic receptor antagonist selective for muscarinic receptors M1 and / or M3 and / or M5.

9. 9. The combination of claim 8, wherein the muscarinic receptor antagonist is selective for the muscarinic receptor M3, preferably the anticholinergic compound is darifenacin, ipratropium bromide, tiotropium bromide, or trospium.

10. 9. The combination of claim 8, wherein the muscarinic receptor antagonist is darifenacin.

11. 8. The combination of claim 7, wherein the anti-MuSK antibody or antigen-binding fragment thereof and the anticholinergic compound are present in a single composition.

12. 8. The combination of claim 7, wherein the anti-MuSK antibody or antigen-binding fragment thereof and the anticholinergic compound are present in separate compositions.

13. A pharmaceutical composition for treating a neuromuscular disorder, comprising the anti-MuSK antibody or antigen-binding fragment thereof of claim 1, or the combination of claim 7.

14. 14. The pharmaceutical composition of claim 13, wherein the treatment comprises the administration of the combination of claim 7 and the anticholinergic compounds administered separately, sequentially, or simultaneously.

15. 14. The pharmaceutical composition of claim 13, wherein the neuromuscular disorder is characterized by impaired neuromuscular transmission and / or denervation of the NMJ (neuromuscular junction).

16. The neuromuscular disorder is A. Muscarinic hyperexcitability b. motor neuron death, c. Neuromuscular junction (NMJ) denervation, and D. Impairment of synaptic transmission 14. The pharmaceutical composition of claim 13, characterized by at least one of:

17. 14. The pharmaceutical composition of claim 13, wherein the neuromuscular disorder is selected from the group consisting of amyotrophic lateral sclerosis (ALS), spinal muscular atrophy (SMA), myasthenia gravis (MG), congenital myasthenia, Lambert-Eaton myasthenic syndrome (LEMS), Lyme disease, polio, post-polio, heavy metal poisoning, Kennedy syndrome, adult-onset Tay-Sachs disease, hereditary spastic paraplegia, multifocal neuropathy, cervical spondylosis, extramedullary tumor with compressive radiculopathy and myelopathy, inclusion body myositis, progressive bulbar palsy, progressive muscular atrophy, motor neuron syndrome, and thyrotoxic myopathy.

18. 14. The pharmaceutical composition of claim 13, wherein the neuromuscular disorder is ALS.

19. The pharmaceutical composition of claim 13, wherein the anti-MuSK antibody or antigen-binding fragment thereof, or the combination, is administered to an asymptomatic human subject at the time of disease onset, preferably within 1, 2, 3, 4, 5, or 6 months before the onset of the disease.

20. 20. The pharmaceutical composition of claim 19, wherein the asymptomatic human subject is diagnosed as predisposed to developing a neuromuscular disorder or disease.

21. 20. The pharmaceutical composition of claim 19, wherein the disease manifestations include at least one symptom selected from the group consisting of muscle contractions, muscle cramps, spasticity, muscle weakness, slurred speech and / or nasal voice, difficulty chewing or swallowing, dysphagia, dysarthria, and difficulty breathing.

22. 14. The pharmaceutical composition of claim 13, wherein the neuromuscular disorder is analyzed by electrophysiological or pharmacodynamic assessment: in blood serum, plasma, and / or cerebrospinal fluid (CSF) neurofilaments (e.g., neurofilament light chain (NFL)); or in an NMJ biopsy.

23. administration of the anti-MuSK antibody or antigen-binding fragment thereof, or the combination to the human subject, to achieve the following therapeutic effect: - increasing the number or percentage of fully innervated NMJs in said subject, maintaining the number or percentage of fully innervated NMJs in said subject, decreasing the number or percentage of fully denervated NMJs in said subject, improving the reliability of synaptic transmission, or preventing, stabilizing, or reducing motor neuron death in said subject; and / or - improving the subject's athletic performance and / or grip strength; and / or - improving the contractile properties of muscles at the NMJ in said subject; and / or - improved resistance to muscle fatigue at the NMJ in said subject; and / or - inducing an increase in muscle mass at the NMJ in said subject; and / or - improving the subject's quality of life or delaying the onset of a deterioration in quality of life; and / or - reducing muscarinic activity (or reducing muscarinic hyperexcitability) of perisynaptic Schwann cells (PSCs) in said subject, or repairing NMJs in said subject.

15. The pharmaceutical composition of claim 14, wherein the composition provides one or more of:

24. 14. The pharmaceutical composition of claim 13, wherein said treatment results in stabilization of said disorder.

25. The pharmaceutical composition of claim 13, wherein treatment of the neuromuscular disorder results in improvement by electrophysiological or pharmacodynamic assessment; in neurofilaments (e.g., neurofilament light chain (NFL)) in blood serum, plasma, and / or cerebrospinal fluid (CSF); or in NMJ biopsies of the treated human subject compared to a human subject not treated with the anti-MuSK antibody or antigen-binding fragment thereof.

Images