Anti-TDP-43 binding molecules and uses thereof

Abstract

The present invention belongs to the field of transactivation-responsive DNA binding protein with a molecular weight of 43 kDa (TARDB or also known as TDP-43). The present invention relates to TDP-43 specific binding molecules, in particular anti-TDP-43 antibodies or antigen-binding fragments or derivatives thereof and their uses. The present invention provides means and methods for diagnosing, preventing, alleviating and / or treating diseases, disorders and / or abnormalities associated with TDP-43 aggregates, including but not limited to frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE) and limbic-dominant age-related TDP-43 encephalopathy (LATE).

Description

Technical Field

[0001] The present invention belongs to the field of transactivation responsive DNA binding protein with a molecular weight of 43 kDa (TARDB or also TDP-43). The present invention relates to TDP-43 specific binding molecules, in particular anti-TDP-43 antibodies or antigen-binding fragments or derivatives thereof and their uses. The present invention provides means and methods for diagnosing, preventing, alleviating and / or treating diseases, disorders and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathies, including but not limited to frontotemporaldementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE) and limbic-predominant age-related TDP-43 encephalopathy (LATE). Background Art

[0002] Age-related encephalopathy, characterized by the pathological aggregation of proteins in the central nervous system (CNS) and peripheral organs (proteinopathies), is one of the leading causes of disability and mortality worldwide. The best-characterized protein that forms aggregates is amyloid-β in Alzheimer's disease and related disorders. Other aggregation-prone proteins associated with other diseases that lead to neurodegeneration include, but are not limited to, Tau, α-synuclein (aSyn, a-syn), huntingtin, fused in sarcoma (FUS), dipeptide repeat proteins (DPRs) generated by unconventional translation of C9orf72 repeat expansions, superoxide dismutase 1 (SOD1), and TDP-43. Diseases involving TDP-43 aggregates are generally classified as TDP-43 proteinopathies and include, but are not limited to, ALS and FTD.

[0003] I. Introduction to TDP-43

[0004] Transactivation response (transactive response, TAR) DNA binding protein 43kDa (TDP-43) is a 414 amino acid protein encoded by the TARDBP gene on chromosome 1p36.2 (ALS10). TARDBP consists of six exons (exon 1 is non-coding; exons 2 to 6 are protein-coding). TDP-43 belongs to the heterogeneous ribonucleoprotein (hnRNP) RNA binding protein family (Wang et al., Trends in Molecular Medicine Vol.14 No.11, 2008, 479-485; Lagier-Tourenne et al., Human Molecular Genetics, 2010, Vol.19, Review Issue 1 R46-R64). TDP-43 contains five functional domains (Warraich et al., TheInternational Journal of Biochemistry&Cell Biology 42 (2010) 1606-1609, Figure 1): two RNA recognition motifs (RRM1 and RRM2), which have two highly conserved hexameric ribonucleoprotein 2 (RNP2) and octameric ribonucleoprotein 1 (RNP1) regions; nuclear export signal (NES) and nuclear localization signal (NLS), which enable it to shuttle between the nucleus and cytoplasm and transport bound mRNA; and a glycine-rich domain at the C-terminus that mediates protein-protein interactions. TDP-43 is involved in multiple aspects of RNA processing, including transcription, splicing, transport, and stabilization (Buratti and Baralle, FEBS Journal 277 (2010) 2268-2281). TDP-43 is a highly conserved, ubiquitously expressed protein with strictly self-regulated expression levels. It constantly shuttles between the nucleus and cytoplasm, but is primarily localized in the nucleus. In 2006, TDP-43 was identified as a protein that accumulates in the vast majority of cases of frontotemporal lobar degeneration (FTLD) with tau-negative, ubiquitin-positive inclusions (hereafter referred to as FTLD-TDP), as well as in most cases of amyotrophic lateral sclerosis (ALS) (Arai et al., Biochemical and Biophysical Research Communications 351 (2006) 602-611; Neumann et al., Science 314, (2006), 130-133).

[0005] Thirty-eight dominant-negative mutations of TDP-43, primarily located in the glycine-rich domain, have been identified in patients with sporadic and familial ALS and in patients with hereditary FTD (Lagier-Tourenne and Cleveland, Cell 136, 2009, 1001-1004, Figure 1 TDP-43 is intrinsically prone to aggregation, as shown by sedimentation assays, and this tendency is further enhanced by some ALS-associated TARDBP mutations (Ticozzi et al., CNS Neurol. Disord. Drug Targets. 2010, 9(3), 285-296.), linking TDP-43 aggregation to clinical disease manifestations.

[0006] II. TDP-43 in Neurodegeneration

[0007] TDP-43 aggregates have been identified in a growing number of neurodegenerative disorders (Lagier-Tourenne et al., Human Molecular Genetics, 2010, Vol.19, Review Issue 1 R46-R64), including but not limited to: frontotemporal dementia (FTD, such as sporadic or familial, with or without motor neuron disease (MND), progranulin (GRN) mutations, C9orf72 mutations, TARDBP mutations, valosine-containing protein (VCP) mutations, linkage to chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), nonfluent variant primary progressive aphasia (NSP Progressive Aphasia (nfvPPA), amyotrophic lateral sclerosis (ALS, such as sporadic ALS, with TARDBP mutations, with angiogenin (ANG) mutations), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD, including sporadic and familial forms of AD), Down syndrome, familial British dementia, polyglutamine diseases (Huntington disease and spinocerebellar ataxia type 3 (SCA3; also known as Machado-Joseph Disease)), hippocampal sclerosis dementia, and myopathies (sporadic inclusion body myositis; inclusion body myopathy with valosin-containing protein mutations (VCP; and Paget disease of bone). bone and frontotemporal dementia; oculopharyngeal muscular dystrophy with rimmed vacuoles; myofibrillar myopathy with mutations in the myotilin (MYOT) gene or the gene encoding desmin (DES), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD).

[0008] Aggregated TDP-43 from patient brains shows numerous abnormal modifications, including hyperphosphorylation, ubiquitination, acetylation, and proteolytic cleavage of a C-terminal fragment (Arai et al., Biochemical and Biophysical Research Communications 351 (2006) 602-611; Neumann et al., Science 314, (2006), 130-133; Neumann et al., Acta Neuropathol. (2009) 117: 137-149; Hasegawa et al., (2008) Annals of Neurology Vol 64 No 1, 60-70; Cohen et al., Nat Commun. 6: 5845, 2015). Another characteristic feature of TDP-43 pathology is the redistribution and accumulation of TDP-43 from the nucleus to the cytoplasm. The hallmark lesions of FTLD-TDP are neuronal cytoplasmic inclusions and glial cytoplasmic inclusions (NCIs and GCIs, respectively) and dystrophic neurites (DNs), which are immunoreactive for TDP-43 as well as ubiquitin and p62, but negative for other neurodegenerative disease-associated proteins. Differences in inclusion morphology and their tissue distribution are associated with specific mutations and / or clinical manifestations. To date, four types of TDP-43 pathology have been described by histological classification (Mackenzie and Neumann, J. Neurochem. (2016) 138 (Suppl 1), 54-70). FTLD-TDP type A cases are characterized by numerous short dystrophic neuritis (DN) and compact oval or crescent-shaped NCIs, predominantly in layer II of the neocortex (Mackenzie et al., 2016 J. Neurochem. 138(Suppl 1), 54-70, Figure 2f). This pathological condition is often clinically seen in the context of behavioral variant frontotemporal dementia (bvFTD) or nonfluent / grammatic variant primary progressive aphasia (nfvPPA) and is associated with mutations in progranulin (GRN). Type B cases show a moderate number of compact or granular NCIs in both superficial and deep cortical layers, with relatively few DNs and NIIs (neuronal intranuclear inclusions; Mackenzie et al., 2016 J. Neurochem. 138 (Suppl 1), 54-70, Figure 2 g). Most cases with concurrent FTD and ALS symptoms were found to have FTLD-TDP type B pathology. Type C cases had numerous long, tortuous neurites, primarily in the superficial cortical layers, with few or no NCIs (Mackenzie et al., 2016 J. Neurochem. 138(Suppl 1), 54-70, Figure 2 j). This pathology is particularly found in cases with semantic variant primary progressive aphasia (svPPA). FTLD-TDP type D shows abundant lenticular intranuclear inclusions (NIIs) and short DNs in the neocortex, with only rare NCIs (Mackenzie et al., 2016 J. Neurochem. 138(Suppl 1), 54-70, Figure 2 k). Type E is characterized by the presence of granulofilamentous neuronal inclusions (GFNI) and very fine punctate neuropil aggregates in addition to curvilinear oligodendrocyte inclusions in the white matter, affecting all neocortical layers (Edward B. Lee et al., Acta Neuropathol. 2017 July; 134(1): 65-78.). This pathological pattern has only been found in cases of VCP associated with inclusion body myositis.

[0009] III. TDP-43 in FTD

[0010] Frontotemporal dementia (FTD) is a clinical term that covers a broad spectrum of disorders characterized by pathological features based on the degeneration of the frontal and temporal lobes, called frontotemporal lobar degeneration (FTLD). FTD is the second most common cause of early degenerative dementia in the age group under 65 years old (Le Ber, Revue Neurologique 169 (2013) 811-819). FTD manifests as several syndromes, including bvFTD, characterized by personality and behavioral changes; semantic dementia (SD) and progressive nonfluent aphasia (PNFA), characterized by changes in language function; corticobasal syndrome (CBS), progressive supranuclear palsy syndrome, and motor neuron disease (FTD-MND), characterized by motor dysfunction. The clinical diagnosis of these syndromes is complex and a final conclusion can only be drawn by performing postmortem histopathological analysis to detect aggregated proteins and determine the affected brain areas. Regarding pathological protein inclusions, approximately 45% of cases showed pathological accumulation of misfolded Tau, 45% had pathological TDP-43, and smaller subsets had aggregates of FUS and other proteins.

[0011] IV. TDP-43 in ALS

[0012] Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by the premature loss of upper and lower motor neurons. The progression of ALS is characterized by fatal paralysis and respiratory failure, with a course of 1 to 5 years from diagnosis to death. In most cases of sporadic ALS, the neuropathology is characterized by abnormal cytoplasmic accumulation of TDP-43 in neurons and glial cells of the primary motor cortex, brainstem motor nuclei, spinal cord and associated white matter tracts. ALS with dementia involves accumulation of TDP-43 in the extramotor neocortex and hippocampus. The role of TDP-43 phosphorylation in ALS patients has been explored with the help of antibodies that specifically bind to phosphorylated TDP-43 in the nucleus and cytoplasmic inclusions, among which amino acids S379, S403, S404, S409, and S410 are the main sites of TDP-43 phosphorylation (Hasegawa et al., Ann Neurol 2008; 64: 60-70; Neumann et al., Acta Neuropathol (2009) 117: 137-149).

[0013] V. TDP-43 in AD and other diseases

[0014] TDP-43 pathology occurs in the brains of up to 57% of patients with Alzheimer's disease (Josephs KA et al., Acta Neuropathol. 2014; 127(6): 811-824; Josephs KA et al., Acta Neuropathol. 2014; 127(3): 441-450; McAleese et al., Brain Pathol. 2017 Jul; 27(4): 472-479). TDP-43 aggregation is associated with patient age and is associated with cognitive decline, memory loss, and mesial temporal atrophy in AD. Apparently, in AD, TDP-43 represents a second or independent pathology that shares overlapping brain distribution with beta-amyloid and tau pathology in the mesial temporal lobe. Pathological TDP-43 follows a general progressive deposition pattern that has been described by the so-called TDP-43 in AD (TAD) staging scheme: TDP-43 is first deposited in the amygdala (stage I), followed by deposition in the hippocampus, limbic, temporal, and finally frontostriatum (stage V) (Josephs KA et al., Acta Neuropathol. 2014; 127(6): 811-824; Josephs KA et al., Acta Neuropathol. 2014; 127(3): 441-450).

[0015] VI. TDP-43 Diffusion

[0016] Although the onset and initial symptoms of ALS and FTD vary significantly between patients, a common feature of disease progression is the spread of pathology from the initial lesion area to most neurons. The continued worsening of symptoms can be explained by the progressive spread of TDP-43 pathology. TDP-43 pathology in the brains of ALS patients has been shown to spread in a four-stage process and is believed to spread through synapses using anterograde axonal transport via axonal transmission from the cortex (Brettschneider et al., Ann Neurol. 2013 July; 74 (1): 20-38.). Recent experimental evidence supports the hypothesis that beta-amyloid, Tau, α-synuclein and TDP-43 spread proteins in neuronal tissue through a prion-like mechanism (Hasegawa et al., 2017), in which the starting point and topographical diffusion pattern are different for the four proteins (Brettschneider J et al., Nature Rev. Neuroscience, 2015, 109). It is believed that the common unifying mechanism of the disease is based on the intercellular spread of pathological protein aggregates. The mechanism consists of aggregate release from diseased cells, uptake by naive cells, and seeding of pathological protein conformations through templated conformational changes of endogenous proteins.

[0017] TDP-43 intercellular spread has been studied at the molecular level in a few in vitro models, in which insoluble TDP-43 preparations from patient brains were able to induce the formation of intracellular aggregates in recipient cells (Nonaka et al., Cell Reports 4 (2013), 124-134; Feiler et al., 2015; Porta et al., Nat. Comm., 2018). In addition, intracellular TDP-43 aggregates have been observed to be released in conjunction with exosomes before spreading to the next cell (Nonaka et al., Cell Reports 4 (2013, 124-134)). Similarly, adenoviral transduction of TDP-43 expression leads to cytoplasmic aggregates that are phosphorylated, ubiquitinated, and more importantly, act as seeds for initiating intercellular spread (Ishii et al., PLoS ONE 12 (6): e0179375, 2017). Patient-derived pathological TDP-43 can lead to widespread deposition of endogenous TDP-43 after intracranial inoculation into transgenic and wild-type mice (Porta et al., Nat. Comm., 2018).

[0018] VII. Prevention and Treatment of TDP-43 Proteinopathy

[0019] TDP-43 aggregation and pathological spread are the hallmarks of ALS and FTD – currently incurable and fatal diseases. Mutations in TDP-43 are associated with familial cases of ALS and FTD, providing a causal link between TDP-43 misfolding and disease progression.

[0020] VIII. Diagnosis of TDP-43 Proteinopathy

[0021] The diagnosis of FTD based on clinical presentation is inadequate because clinical manifestations can overlap with other disorders, especially in the early stages.

[0022] Many approaches are aimed at developing biochemical biomarkers to distinguish different types of FTD pathology. The development of antibodies against different conformations of TDP-43 could allow for the creation of more sensitive and specific diagnostic tools. In parallel with biochemical biomarkers, the development of imaging biomarkers could enable early and specific detection of pathology in TDP-43 proteinopathies. The ability to image TDP-43 deposition in the brain could be a major advance in the diagnosis and drug development of TDP-43 proteinopathies. Such detection could be achieved using cell-permeable antibody fragments.

[0023] The earliest event in neurodegenerative diseases based on misfolding of various proteins is the acquisition of an alternative conformation that renders the protein toxic. Furthermore, this misfolded conformation can self-propagate by recruiting endogenous normal proteins into the misfolded conformation, serving as the mechanistic basis for the observed spread through affected tissues.

[0024] In order to develop antibodies against different conformational states of a given protein, supramolecular antigen constructs have been designed in which the conformation of the presented antigen is controlled to produce conformation-specific antibodies against a given target in a specific conformational state (WO2012 / 055933 and WO2012 / 020124). Conformation-specific antibodies provide many advantages because they can distinguish between disease-associated conformations of these proteins and functional endogenous conformations. This approach provides many advantages in therapeutic applications because such antibodies are less likely to be attracted to their normal conformations while targeting misfolded disease-associated isoforms of the protein. Similarly, in diagnostic applications, such antibodies only recognize the disease-associated structural states of the protein, which is crucial for the development of sensitive and specific diagnostics.

[0025] The utility of TDP-43-based biomarkers for TDP-43 proteinopathies remains to be established. Such evaluation is hampered, in part, by the lack of high-affinity antibodies that can be used in suitable immunoassays to quantify pathological TDP-43 in biological fluids (Feneberg et al., Molecular Neurobiology, 2018).

[0026] Therefore, there is a clear need for biomarkers capable of detecting misfolded, aggregated TDP-43 and non-aggregated physiological TDP-43, in particular in human samples, for use in diagnosing different types of TDP-43 proteinopathies and / or for monitoring the efficacy of therapeutic agents for the treatment of diseases, disorders and abnormalities associated with TDP-43, in particular associated with TDP-43 aggregates, or TDP-43 proteinopathies.

[0027] TDP-43 proteinopathies are defined as a group of neurodegenerative diseases characterized by pathological TDP-43.

[0028] IX. Prior Art

[0029] Patent application WO 2008 / 151055 discloses methods and materials for determining whether a mammal has a neurodegenerative disease using the levels of TDP-43 polypeptide and / or TDP-43 polypeptide cleavage products (eg, 25kD and 35kD TDP-43 polypeptide cleavage products) in a biological fluid.

[0030] Patent application WO 2013 / 061163 discloses TDP-43-specific binding molecules, which include polypeptides such as human antibodies and fragments, derivatives and variants thereof. Summary of the Invention

[0031] In view of the foregoing, there is a need for anti-TDP-43 binding molecules that bind to misfolded aggregated TDP-43 and non-aggregated physiological TDP-43, in particular human TDP-43. Furthermore, the development of sensitive and specific biomarkers that allow differentiation of pathological types in the FTD spectrum is an urgent task.

[0032] This technical problem is solved by the embodiments provided herein.

[0033] Therefore, the present invention relates to binding molecules, in particular antibodies or antigen-binding fragments thereof, that specifically recognize misfolded aggregated TDP-43 and non-aggregated physiological TDP-43. In the present invention, misfolded TDP-43 includes misfolded monomeric TDP-43 and / or misfolded oligomeric TDP-43 and / or misfolded aggregated and / or post-translationally modified TDP-43 and / or misfolded truncated TDP-43. Post-translationally modified TDP-43 includes phosphorylated, ubiquitinated, acetylated, ubiquitinated and / or methylated TDP-43. Physiological TDP-43 includes soluble nuclear TDP-43. It is shown herein that the binding molecules of the present invention are capable of binding to pathological TDP-43, including TDP-43 aggregates and phosphorylated TDP-43 (see Example 13). Therefore, the present invention provides binding molecules, in particular antibodies or antigen-binding fragments thereof, that specifically recognize misfolded aggregated TDP-43 and non-aggregated physiological TDP-43. Such binding molecules are referred to herein as "pan-TDP-43" binding molecules, in particular pan-TDP-43 antibodies. As described herein, the TDP-43 binding molecules of the present invention may bind equally to misfolded aggregated TDP-43 and non-aggregated physiological TDP-43, or, when specifically binding to both types of TDP-43, may preferentially bind to one relative to the other. The present invention also provides binding molecules, in particular antibodies or antigen-binding fragments thereof, for preventing, alleviating, treating and / or diagnosing diseases, disorders and abnormalities associated with TDP-43, in particular TDP-43 aggregates, or TDP-43 proteinopathies. The present invention also provides binding molecules, in particular antibodies or antigen-binding fragments thereof, for detecting and / or understanding (i.e., identifying) specific pathological types that cause neurodegeneration. Use as diagnostic biomarkers is envisioned to enable more efficient and accurate subject selection for longitudinal monitoring in clinical studies, supporting the development of new treatments for TDP-43 proteinopathies.

[0034] The present invention also provides TDP-43 binding molecules, particularly antibodies or antigen-binding fragments thereof, as drugs (therapeutic agents).

[0035] Without wishing to be bound by theory, the present invention was developed based on the following hypothesis: modified conformation-specific antigenic peptides and peptide fragments derived from TDP-43 protein or the entire TDP-43 protein and antibodies obtainable by or through said peptides or fragments or the entire TDP-43 protein block TDP-43 cell-to-cell spread and / or disaggregate TDP-43 aggregates and / or block TDP-43 seeding and / or inhibit aggregation of TDP-43 protein or its fragments. The binding molecules, in particular polypeptides, more particularly antibodies or antigen-binding fragments thereof of the present invention bind to misfolded aggregated TDP-43, in particular to cytoplasmic and extracellular misfolded TDP-43. The binding molecules, in particular polypeptides, more particularly antibodies or antigen-binding fragments thereof of the present invention bind to full-length TDP-43 and / or truncated TDP-43. In one embodiment, the binding molecules, in particular polypeptides, more particularly antibodies or antigen-binding fragments thereof of the present invention specifically bind to cytoplasmic misfolded TDP-43.

[0036] Misfolded, aggregated TDP-43 or pathologically associated TDP-43 consists of TDP-43 protein that has lost its normal folding (i.e., is misfolded) and localization. Misfolded, aggregated TDP-43 can be found in the following: preinclusions, as well as neuronal and glial cytoplasmic inclusions (NCIs and GCIs, respectively), neuronal nuclear inclusions (NIIs), and dystrophic neurites (DNs), which are immunoreactive for TDP-43.

[0037] Non-aggregated physiological TDP-43 is a physiologically functional TDP-43 protein that is primarily localized in the cell nucleus and shuttled to the cytoplasm, in a state where it can exhibit its desired function in the in vivo cellular environment.

[0038] The binding molecules of the present invention, in particular antibodies or antigen-binding fragments thereof, unexpectedly have at least one, preferably two, more preferably three, and even more preferably all four of the following characteristics:

[0039] - Blocks the cell-to-cell spread of TDP-43;

[0040] - disaggregate TDP-43 aggregates;

[0041] - Inhibit the aggregation of TDP-43 protein or its fragments;

[0042] - Blocking TDP-43 seeding.

[0043] Independent of the combination of one, two, three or four of the above-listed features, the binding molecules of the present invention, preferably antibodies or antigen-binding fragments thereof, can improve / inhibit / reduce the development of TDP-43 pathology in in vivo models of TDP-43 proteinopathy and, more importantly, in patients suffering from TDP-43 pathology.

[0044] The TDP-43 binding molecules of the present invention, in particular antibodies or antigen-binding fragments thereof, can recruit and / or activate microglia. More particularly, it is shown herein (see Examples 10 and Figure 5 ) The TDP-43 binding molecules of the present invention can affect microglial morphology in terms of cell size and activation state. This may contribute to the reduction of TDP-43 pathology demonstrated by the TDP-43 binding molecules of the present invention.

[0045] In the present invention, binding molecules, in particular antibodies or antigen-binding fragments thereof, specifically recognize TDP-43. The binding molecules of the present invention include polypeptides and / or antibodies and / or antigen-binding fragments thereof that are specific for the TDP-43 protein. "Specifically recognizing TDP-43" means that the binding molecules of the present invention specifically, generally, and collectively bind with greater affinity to TDP-43, in particular to certain epitopes in TDP-43, in particular to epitopes of the TDP-43 protein that are exposed / accessible in one or more pathological conformations, compared to other epitopes. The binding molecules of the present invention, in particular polypeptides, more particularly antibodies or antigen-binding fragments thereof, that specifically bind to TDP-43 specifically recognize misfolded aggregated TDP-43 and non-aggregated physiological TDP-43. In a preferred embodiment, the full-length human TDP-43 comprises, preferably has, the sequence of SEQ ID NO: 1. In another preferred embodiment of the present invention, the binding molecule, in particular the antibody or antigen-binding fragment thereof, specifically binds to a defined binding region in full-length and / or truncated TDP-43, wherein the binding region is preferably comprised within amino acids 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411 or 140 to 200 of the full-length human TDP-43 having the sequence of SEQ ID NO: 1, more preferably, the binding region is comprised within amino acids 183 to 188, 203 to 213, 204 to 208, 204 to 211, 205 to 210, 316 to 323, 358 to 361, 400 to 405, 400 to 406 or 400 to 412 thereof. Therefore, the binding molecule, in particular the antibody or antigen-binding fragment thereof, preferably specifically binds to a peptide comprising, preferably consisting of, a binding region consisting of amino acids 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411 or 140 to 200 of the full-length human TDP-43 having the sequence of SEQ ID NO: 1. In another preferred embodiment of the present invention, the binding molecule, in particular an antibody or antigen-binding fragment thereof, preferably specifically binds to a peptide comprising, preferably consisting of, a binding region consisting of amino acids 183 to 188, 203 to 213, 204 to 208, 204 to 211, 205 to 210, 316 to 323, 358 to 361, 400 to 405, 400 to 406, or 400 to 412 of human TDP-43 (SEQ ID NO: 1). In some embodiments, the TDP-43 binding molecule, in particular an antibody or antigen-binding fragment thereof, binds in the C-terminal region of TDP-43.This can be advantageous, for example, because the C-terminal fragment of TDP-43 is found in the insoluble fraction and may therefore be pathologically relevant. More particularly, the TDP-43 binding molecule, in particular an antibody or antigen-binding fragment thereof, can bind to an epitope within amino acid residues 400 to 405, 400 to 406, or 400 to 412 of human TDP-43 (SEQ ID NO: 1). In some embodiments of the invention, the antibody is a monoclonal antibody. In some embodiments, the antibody is a murine antibody, a murinized antibody, a human antibody, a humanized antibody, or a chimeric antibody. It will be appreciated that equivalent binding regions exist in non-human TDP-43. Thus, for example, the mouse TDP-43 amino acid sequence (see Uniprot Accession No. Q921F2) is also 414 amino acids in length and is 96% (398 / 414 residues) identical to the human sequence. The present invention encompasses binding molecules, particularly antibodies or antigen-binding fragments thereof, that bind to regions / peptides of non-human TDP-43, particularly murine TDP-43, equivalent to those specified above with reference to SEQ ID NO: 1.

[0046] In particular, the present invention is summarized in the following embodiments:

[0047] 1. A TDP-43 binding molecule that binds to misfolded aggregated TDP-43 and non-aggregated physiological TDP-43, in particular human TDP-43.

[0048] 2. The TDP-43 binding molecule of the preceding embodiment, which binds to an epitope within amino acid residues 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411, or 140 to 200 of human TDP-43 (SEQ ID NO: 1).

[0049] 3. The TDP-43 binding molecule of the preceding embodiment, which binds to an epitope within amino acid residues 183 to 188, 203 to 213, 204 to 208, 204 to 211, 205 to 210, 316 to 323, 358 to 361, 400 to 405, 400 to 406, or 400 to 412 of human TDP-43 (SEQ ID NO: 1).

[0050] 4. The binding molecule of any preceding embodiment, which is an antibody or an antigen-binding fragment thereof.

[0051] 5. The binding molecule or TDP-43 binding molecule of any preceding embodiment, comprising:

[0052] a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 15, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 17; or

[0053] b) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 25, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 27; or

[0054] c) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 31, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 32, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 33, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 37; or

[0055] d) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 41, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 42, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 43, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 45, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 46, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 47; or

[0056] e) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 61, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 62, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 63, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 65, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 66, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 67; or

[0057] f) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 71, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 72, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 73, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 75, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 77; or

[0058] g) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 81, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 82, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 83, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 85, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 86, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 87; or

[0059] h) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 101, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 102, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 103, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; or

[0060] i) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 121, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 122, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 123, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 125, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 127; or

[0061] j) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 145, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 146, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 147; or

[0062] k) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 151, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 152, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 153, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 155, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 156, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 157.

[0063] 6. The binding molecule or TDP-43 binding molecule of any one of the preceding embodiments, which is an antibody or antigen-binding fragment thereof comprising:

[0064] a. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 10, or a heavy chain variable region (VH) having at least 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 10, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 14, or a light chain variable region (VL) having at least 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 14; or

[0065] b. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 20, or a heavy chain variable region (VH) having at least 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 20, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 24, or a light chain variable region (VL) having at least 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 24; or

[0066] c. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 30, or a heavy chain variable region (VH) having at least 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 30, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 34, or a light chain variable region (VL) having at least 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 34; or

[0067] d. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 40, or a heavy chain variable region (VH) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 40, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 44; or

[0068] e. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 60, or a heavy chain variable region (VH) having at least 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 60, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 64, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 64; or

[0069] f. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 70, or a heavy chain variable region (VH) having at least 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 70, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 74, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 74; or

[0070] g. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 80, or a heavy chain variable region (VH) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 80, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 84, or a light chain variable region (VL) having at least 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 84; or

[0071] h. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 100, or a heavy chain variable region (VH) having at least 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 100, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 104, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 104; or

[0072] i. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 120, or a heavy chain variable region (VH) having at least 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 120, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 124, or a light chain variable region (VL) having at least 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 124; or

[0073] j. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 140, or a heavy chain variable region (VH) having at least 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 140, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 144; or

[0074] k. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 150, or a heavy chain variable region (VH) having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 150, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 154, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 154.

[0075] 7. The binding molecule or TDP-43 binding molecule of any of the preceding embodiments, which is an antibody or antigen-binding fragment thereof comprising:

[0076] a. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 10 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 14; or

[0077] c. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 20 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 24; or

[0078] d. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 30 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 34; or

[0079] e. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 40 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 44; or

[0080] f a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 60 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 64; or

[0081] g. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 70 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 74; or

[0082] h. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 80 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 84; or

[0083] i. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 100 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 104; or

[0084] j. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 120 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 124; or

[0085] k. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 140 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 144; or

[0086] 1. A heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 150 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 154.

[0087] In some embodiments, the antibody comprises:

[0088] a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 15, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 17; or

[0089] b) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 25, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 27; or

[0090] c) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 31, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 32, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 33, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 37; or

[0091] d) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 41, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 42, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 43, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 45, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 46, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 47; or

[0092] e) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 61, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 62, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 63, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 65, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 66, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 67; or

[0093] f) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 71, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 72, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 73, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 75, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 77; or

[0094] g) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 81, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 82, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 83, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 85, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 86, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 87; or

[0095] h) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 101, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 102, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 103, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; or

[0096] i) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 121, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 122, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 123, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 125, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 127; or

[0097] j) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 145, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 146, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 147; or

[0098] k) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 151, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 152, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 153, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 155, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 156, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 157.

[0099] In some embodiments, the antibody comprises:

[0100] a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a VH-CDR3 comprising the amino acid sequence ES (Glu-Ser); or

[0101] b) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a vH-CDR3 comprising the amino acid sequence ES (Glu-Ser); or

[0102] c) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 31, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 32, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 33; or

[0103] d) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 41, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 42, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 43; or

[0104] e) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 61, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 62, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 63; or

[0105] f) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 71, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 72, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 73; or

[0106] g) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 81, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 82, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 83; or

[0107] h) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 101, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 102, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 103; or

[0108] i) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 121, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 122, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 123; or

[0109] j) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143; or

[0110] k) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 151, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 152, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 153.

[0111] In some embodiments, the antibody comprises:

[0112] a) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 15, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 17; or

[0113] b) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 25, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 27; or

[0114] c) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 37; or

[0115] d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 65, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 66, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 67; or

[0116] e) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 75, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 77; or

[0117] f) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 85, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 86, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 87; or

[0118] g) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; or

[0119] h) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 125, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 127; or

[0120] i) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 155, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 156, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 157.

[0121] In some embodiments, the antibody comprises:

[0122] a) a VH-CDR1 comprising an amino acid sequence having at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 11, a VH-CDR2 comprising an amino acid sequence having at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 12, a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 15, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 17; or

[0123] b) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 21, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 22, a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 25, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 27; or

[0124] c) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 31, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 32, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 33, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 37; or

[0125] d) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:41, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:42, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:43, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO:45, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO:46, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO:47; or

[0126] e) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:61, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:62, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:63, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO:65, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO:66, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO:67; or

[0127] f) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 71, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 72, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 73, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 75, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 77; or

[0128] g) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 81, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 82, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 83, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 85, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 86, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 87; or

[0129] h) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 101, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 102, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 103, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and a vL-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; or

[0130] i) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 121, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 122, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 123, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 125, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 127; or

[0131] j) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 141, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 142, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 143, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 145, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 146, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 147; or

[0132] k) a VH-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 151, a VH-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 152, a VH-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 153, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 155, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 156, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 157; or

[0133] In some embodiments, the antibody comprises:

[0134] a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 12, a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 15, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 16, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 17; or

[0135] b) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO:21, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO:22, a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:25, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:16, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:27; or

[0136] c) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO:31, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO:32, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO:33, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:35, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:36, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:37; or

[0137] d) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO:41, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO:42, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO:43, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:45, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:46, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:47; or

[0138] e) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO:61, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO:62, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO:63, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:65, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:66, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:67; or

[0139] f) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO:71, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO:72, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO:73, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:75, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:16, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:77; or

[0140] g) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO:81, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO:82, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO:83, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:85, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:86, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO:87; or

[0141] h) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 101, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 102, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 103, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 105, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 106, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 107; or

[0142] i) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 121, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 122, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 123, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 125, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 16, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 127; or

[0143] j) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 145, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 146, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 147; or

[0144] k) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 151, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 152, a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 153, a VL-CDR1 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 155, a VL-CDR2 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 156, and a VL-CDR3 comprising an amino acid sequence that has at least 80%, 90%, 95% or 100% sequence identity to SEQ ID NO: 157.

[0145] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 11; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 12; (c) a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser); (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 15; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 17.

[0146] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 21; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 22; (c) a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser); (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 25; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 27.

[0147] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 31; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 32; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 33; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 35; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 36; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 37.

[0148] In some embodiments, the TDP-43 antibody comprises at least one, two, or three CDRs selected from the group consisting of: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 41; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 42; and (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 43.

[0149] In some embodiments, the TDP-43 antibody comprises at least four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 41; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 42; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 43; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 45; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 46; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 47.

[0150] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 61; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 62; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 63; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 65; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 66; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 67.

[0151] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 71; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 72; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 73; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 75; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 77.

[0152] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 81; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 82; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 83; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 85; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 86; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 87.

[0153] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 101; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 102; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 103; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 105; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 106; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 107.

[0154] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 121; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 122; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 123; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 125; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 127.

[0155] In some embodiments, the TDP-43 antibody comprises at least one, two, or three CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143.

[0156] In some embodiments, the TDP-43 antibody comprises at least four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 145; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 146; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 147.

[0157] In some embodiments, the TDP-43 antibody comprises at least one, two, three, four, five, or six CDRs selected from: (a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 151; (b) a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 152; (c) a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 153; (d) a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 155; (e) a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 156; and (f) a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 157.

[0158] In another embodiment, the TDP-43 antibody comprises a heavy chain variable domain (VH) selected from the group consisting of SEQ ID NO: 10, 20, 30, 40, 60, 70, 80, 100, 120, 140, 150, including post-translational modifications of said sequences. In a specific embodiment, the heavy chain variable domain (VH) comprises at least one, two or three CDRs selected from the following: (a) a VH-CDR1 comprising an amino acid sequence selected from SEQ ID NO: 11, 21, 31, 41, 61, 71, 81, 101, 121, 141, 151; (b) a VH-CDR2 comprising an amino acid sequence selected from SEQ ID NO: 12, 22, 32, 42, 62, 72, 82, 102, 122, 142, 152; (c) a VH-CDR3 comprising an amino acid sequence selected from SEQ ID NO: 33, 43, 63, 73, 89, 103, 123, 143, 153 and ES (Glu-Ser).

[0159] In another embodiment, the TDP-43 antibody comprises a light chain variable domain (VL) selected from the group consisting of SEQ ID NOs: 14, 24, 34, 64, 74, 84, 104, 124, 154, including post-translational modifications of said sequences. In a specific embodiment, the light chain variable domain (VL) comprises at least one, two, or three CDRs selected from the group consisting of: (a) a VL-CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 15, 25, 35, 65, 75, 85, 105, 125, 155; and (b) a VL-CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 16, 36, 66, 86, 106, 156; and (c) a VL-CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 17, 27, 37, 67, 77, 87, 107, 127, 157, and ES (Glu-Ser).

[0160] In some embodiments, the TDP-43 antibody comprises at least one, two, or three CDRs selected from the group consisting of: (a) a VH-CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 11, 21, 31, 41, 61, 71, 81, 101, 111, 121, 141, 151; (b) a VH-CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 12, 22, 32, 42, 62, 72, 82, 102, 122, 142, 152; (c) a VH-CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 33, 43, 63, 73, 83, 103, 123, 143, 153, and ES (Glu-Ser).

[0161] In some embodiments, the TDP-43 antibody comprises at least one, two, or three CDRs selected from the group consisting of: (a) a VL-CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 15, 25, 35, 65, 75, 85, 105, 125, 155; (b) a VL-CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 16, 36, 66, 86, 106, 156; (c) a VL-CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 17, 27, 37, 67, 77, 87, 107, 127, 157.

[0162] In some embodiments, the light chain variable domain (VL) comprises at least one, two or three CDRs selected from the following: (a) a VL-CDR1 comprising an amino acid sequence selected from SEQ ID NO: 15, 25, 35, 45, 65, 75, 85, 105, 125, 145, 155; and (b) a VL-CDR2 comprising an amino acid sequence selected from SEQ ID NO: 16, 36, 66, 86, 106, 156; (c) a VL-CDR3 comprising an amino acid sequence selected from SEQ ID NO: 17, 27, 37, 67, 77, 87, 107, 127, 157.

[0163] In some embodiments, the invention relates to antibodies derived from hybridoma clones 631B2A2, 633B12C8, 634H10H7, 636E5B8, 641H1E7, 642A10B11, 642D12B4, 646B7F7, 712A6B10, 809D9C2, or 809F12D8.

[0164] In some embodiments, the invention relates to an antibody selected from the group consisting of: ACI-7069-631B2-Abl, ACI-7069-633B12-Abl, ACI-7069-634H10-Ab2, ACI-7069-636E5-Abl, ACI-7069-641H1-Ab2, ACI-7069-642A10-Abl, ACI-7069-642D12-Abl, ACI-7069-646B7-Abl, ACI-7071-712A6-Abl, ACI-7071-809D9-Ab2, and ACI-7071-809F12-Abl.

[0165] In certain embodiments, the dissociation constant (KD) of a binding molecule or antibody provided herein is ≤1 μM, ≤100 nM, ≤10 nM, ≤1 nM, ≤0.1 nM, ≤0.01 nM, or ≤0.001 nM (e.g., 10-8 M or smaller, such as 10 -8 M to 10 -13 M, for example 10 -3 M to 10 -13 M), particularly with respect to binding to TDP-43, particularly soluble TDP-43, aggregated TDP-43, and / or oligomeric TDP-43. In some embodiments, the TDP-43 binding molecules of the present invention, particularly antibodies or antigen-binding fragments thereof, may have a lower KD for aggregated TDP-43 compared to soluble TDP-43. For example, the TDP-43 binding molecules of the present invention may have a KD of 30 nM or less, in some specific embodiments, 1 nM or less for aggregated TDP-43, and a KD of 500 nM or less for soluble TDP-43. This is demonstrated in Example 8A with reference to Table 8 for the TDP-43 binding molecules of the present invention.

[0166] In one embodiment, the binding affinity to soluble or aggregated FL TDP-43 can be assessed by determining the dissociation constant (KD) using surface plasmon resonance (SPR; Biacore T200, GE Healthcare Life Sciences). Detailed descriptions of suitable SPR methods that can be used are provided in Examples 8A and 8B.

[0167] The TDP-43 binding molecules of the present invention, in particular antibodies or antigen-binding fragments thereof, typically bind to TDP-43 with high affinity. For example, they may exhibit an EC50 value of 200 pM or less, more preferably 20 pM or less, and even more preferably 10 pM or less, as determined by a Luminex assay. Further details of suitable assays can be found in Example 3. Similarly, they may exhibit an EC50 value of 1600 ng / ml or less, more preferably 120 ng / ml or less, and even more preferably 60 ng / ml or less, as determined by an indirect ELISA. Further details of suitable assays can be found in Example 4.

[0168] The TDP-43 binding molecules of the present invention, particularly antibodies or antigen-binding fragments thereof, bind to both non-aggregated physiological TDP-43 and aggregated TDP-43. Thus, the TDP-43 binding molecules of the present invention, particularly antibodies or antigen-binding fragments thereof, can bind approximately equally well to soluble TDP-43 and aggregated TDP-43. The TDP-43 binding molecules of the present invention, particularly antibodies or antigen-binding fragments thereof, can bind approximately equally well to aggregated TDP-43 compared to non-aggregated TDP-43. More particularly, the TDP-43 binding molecules of the present invention, particularly antibodies or antigen-binding fragments thereof, can bind approximately equally well to aggregated TDP-43 compared to non-aggregated TDP-43 in the nucleus. In other embodiments, the TDP-43 binding molecules of the present invention, particularly antibodies or antigen-binding fragments thereof, can preferentially bind to aggregated TDP-43 compared to non-aggregated TDP-43 when binding to both substances. More specifically, the TDP-43 binding molecules, particularly antibodies or antigen-binding fragments thereof, of the present invention may preferentially bind to aggregated TDP-43 in the cytoplasm compared to non-aggregated TDP-43 in the nucleus when bound to the two substances. Alternatively, in other embodiments, the TDP-43 binding molecules, particularly antibodies or antigen-binding fragments thereof, of the present invention may preferentially bind to non-aggregated TDP-43 when bound to the two substances compared to aggregated TDP-43. More specifically, the TDP-43 binding molecules, particularly antibodies or antigen-binding fragments thereof, of the present invention may preferentially bind to non-aggregated TDP-43 in the nucleus compared to aggregated TDP-43 in the cytoplasm when bound to the two substances. These binding properties can be demonstrated, for example, using immunohistochemistry. Suitable methods are described herein with reference to Example 6, for which relevant controls are provided. The results are shown in Table 7.

[0169] The present invention also relates to compositions comprising the binding molecules described herein, in particular the antibodies or antigen-binding fragments thereof (including TDP-43 binding antibody fragments and derivatives) of the present invention. The present invention also relates to immunotherapeutic and / or immunodiagnostic methods using such compositions in the prevention, diagnosis and / or treatment of TDP-43 proteinopathies, wherein an effective amount of the composition is administered to a subject in need thereof.

[0170] In some embodiments, the present invention encompasses binding molecules described herein that specifically bind to TDP-43, particularly antibodies and antigen-binding fragments thereof, and the use of these binding molecules for diagnosing, preventing, alleviating and / or treating diseases, disorders and / or abnormalities associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathies, including but not limited to frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE), and limbic-dominant age-related TDP-43 encephalopathy (LATE). The methods and compositions disclosed herein can be used to diagnose, prevent, alleviating and / or treat diseases, disorders and / or abnormalities associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathies, including but not limited to frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). Preferably, the use of these binding molecules for diagnosing, preventing, alleviating and / or treating diseases, disorders and / or abnormalities associated with TDP-43, in particular TDP-43 aggregates, or TDP-43 proteinopathies is directed to amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD) or frontotemporal dementia (FTD). More preferably, the use is directed to amyotrophic lateral sclerosis (ALS). More preferably, the use is directed to Alzheimer's disease (AD). More preferably, the use is directed to frontotemporal dementia (FTD).

[0171] In another embodiment, a binding molecule as described herein, in particular an antibody or antigen-binding fragment thereof, specific for TDP-43 is contacted with a sample to detect, diagnose and / or monitor a disease, disorder and / or abnormality associated with TDP-43, in particular associated with TDP-43 aggregates, or a TDP-43 proteinopathy selected from frontotemporal dementia (FTD, e.g., sporadic or familial, with or without motor neuron disease (MND), with a progranulin (GRN) mutation, with a C9orf72 mutation, with a TARDBP mutation, with a valosin-containing protein (VCP) mutation, linked to chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), nonfluent variant primary progressive aphasia (NF vPPA), amyotrophic lateral sclerosis (ALS, e.g., sporadic ALS, with TARDBP mutations, with angiogenic protein (ANG) mutations), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy, Perry syndrome, Alzheimer's disease (AD, including sporadic and familial forms of AD), Down syndrome, familial British dementia, polyglutamine diseases (Huntington disease and spinocerebellar ataxia type 3 (SCA3; also known as Mahy-Joh disease)), hippocampal sclerosis dementia and myopathies (sporadic inclusion body myositis, inclusion body myopathy, with valosin-containing protein (VCP) mutations; and Paget's disease of bone and frontotemporal dementia), oculopharyngeal muscular dystrophy with rimmed vacuoles, myofibrillar myopathies with mutations in the myotilin (MYOT) gene or mutations in the gene encoding desmin (DES), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD).

[0172] In one embodiment, the present invention encompasses binding molecules described herein that specifically bind to TDP-43, in particular antibodies of the present invention or antigen-binding fragments thereof, and the use of these molecules, in particular these antibodies, to detect the presence of TDP-43 in a sample. Thus, the TDP-43 binding molecules of the present invention, such as the anti-TDP43 antibodies described herein, can be particularly useful for screening clinical samples, in particular human blood, CSF, interstitial fluid (ISF) and / or urine, for the presence of TDP-43 in the sample, for example, by using an ELISA-based assay or a surface-adapted assay. In some cases, tissue samples, such as brain tissue samples, can be used. The methods and compositions of the present invention can also be applied to diagnose presymptomatic disease and / or monitor disease progression and / or treatment efficacy. According to some embodiments, an antibody specific for TDP-43 (e.g., a full-length antibody or a TDP-43 binding fragment or derivative of the antibody) is contacted with a sample (e.g., blood, cerebrospinal fluid (CSF), interstitial fluid (ISF), or brain tissue) to detect, diagnose, and / or monitor frontotemporal dementia (FTD, e.g., sporadic or familial, with or without motor neuron disease (MND), with progranulin (GRN) mutations, with C9orf72 mutations, with TARDBP mutations, with valosin-containing protein (VCP) mutations, linked to chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), nonfluent variant primary progressive aphasia (nfvPPA), etc.), amyotrophic lateral sclerosis (AMS), myasthenia gravis, leukemia, ... Alzheimer's disease (AD), including sporadic and familial forms of AD, Down syndrome, familial British dementia, polyglutamine diseases (Huntington disease and spinocerebellar ataxia type 3 (SCA3; also known as Mayo-Joseph disease)), hippocampal sclerosis dementia and myopathies (sporadic inclusion body myositis, inclusion body myopathy, with valosin-containing protein (VCP) mutations, and Paget's disease of bone and frontotemporal dementia), oculopharyngeal muscular dystrophy with rimmed vacuoles, myofibrillar myopathy with mutations in the myotilin (MYOT) gene or mutations in the gene encoding desmin (DES), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD).The TDP-43 binding molecules of the present invention can be used to quantify TDP-43 in suitable samples, particularly clinical samples, such as blood, CSF, ISF, or urine, wherein relatively high levels of TDP-43 compared to a suitable control indicate disease and / or more advanced disease. Many suitable immunoassay formats are known. Thus, the method (e.g., ELISA, MSD (Meso Scale Discovery), HTRF (Homogeneous Time Resolved Fluorescence), and AlphaLISA) can be performed for diagnostic purposes, wherein high TDP-43 levels indicate disease. Alternatively, the method can be performed for monitoring purposes. Increasing levels over time can indicate disease progression. Decreasing levels over time can indicate disease regression. The method can also be used to monitor treatment, particularly to monitor the efficacy of a particular treatment. Successful treatment can be measured by reference to stable or declining TDP-43 levels following treatment. It is shown herein (Example 12) that, when measured using the antibodies of the invention, TDP-43 levels in CSF samples from patients with TDP-43 proteinopathies are higher than in control samples taken from healthy subjects (healthy controls). The control samples may or may not be run in parallel with the test samples. In some embodiments, the control levels are determined from a series of control samples taken from healthy subjects under similar or identical experimental conditions and used as a comparison level for the levels determined in the test samples. Methods for quantifying TDP-43 in suitable samples using the binding molecules of the invention can also be used to select treatments (for further treatment of the subject). Thus, personalized treatment methods are contemplated. Samples are taken before and after treatment. If treatment with a treatment results in stable or preferably decreased TDP-43 levels after treatment, the treatment can be selected for the subject. If the treatment does not result in stable or preferably decreased TDP-43 levels after treatment, the treatment is not selected for the subject. The treatment can be any suitable candidate therapeutic agent for treating TDP-43 proteinopathies. In some preferred embodiments, the treatment comprises a TDP-43 binding molecule of the invention, typically in the form of a pharmaceutical composition as described herein.

[0173] The TDP-43 binding molecules of the present invention can also be used to classify diseases into specific types or subtypes. Thus, methods are provided for classifying diseases, disorders, and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or for classifying TDP-43 proteinopathies, comprising:

[0174] a. performing the method of the invention, wherein the level of TDP-43 is quantified compared to a suitable control;

[0175] b. optionally identifying a mutation in a sample from a subject, including but not limited to a progranulin (GRN) mutation, a C9orf72 mutation, a TARDBP mutation, a valosin-containing protein (VCP) mutation, a TARDBP mutation, an angiogenic protein (ANG) mutation, a valosin-containing protein (VCP) mutation, a myotilin (MYOT) gene mutation, or a mutation in the gene encoding desmin (DES); and

[0176] c. Classify diseases, disorders and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathies.

[0177] Similarly, a method for classifying a disease, disorder and / or abnormality associated with TDP-43, particularly associated with TDP-43 aggregates, or a TDP-43 proteinopathy is provided, comprising: performing a method of the invention, wherein the level of TDP-43 in a sample obtained from a subject suffering from a disease, disorder and / or abnormality associated with TDP-43, or a TDP-43 proteinopathy is quantified, wherein the level is compared to a control sample obtained from a subject suffering from a different type or subtype of a disease, disorder and / or abnormality associated with TDP-43, particularly associated with TDP-43 aggregates, or a TDP-43 proteinopathy (i.e., a set of representative control levels is determined for the type or subtype of interest); and classifying the disease, disorder and / or abnormality associated with TDP-43, particularly associated with TDP-43 aggregates, or a TDP-43 proteinopathy based on the comparison. Thus, the classification is based on determining the closest match between the test sample and one or more control samples. These methods may also include identifying mutations in the sample, including but not limited to progranulin (GRN) mutations, C9orf72 mutations, TADBP mutations, valosin-containing protein (VCP) mutations, TARDBP mutations, angiogenic protein (ANG) mutations, valosin-containing protein (VCP) mutations, myotilin (MYOT) gene mutations, or mutations in the gene encoding desmin (DES), wherein the identified mutations are also used to classify diseases, disorders, and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathy. For the avoidance of doubt, identifying mutations in the sample can be performed by any suitable method; for example, based on nucleic acid sequencing of nucleic acid molecules within the sample. The sample can be separate and distinct from the sample in which the TDP-43 level was determined, but from the same subject.

[0178] In other embodiments, the present invention provides a method for preventing, alleviating and / or treating diseases, disorders and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathy. According to one embodiment, the method of the present invention comprises administering to a subject an effective concentration of a binding molecule as described herein, particularly an antibody of the present invention that is specific for TDP-43 (e.g., a full-length antibody or a TDP-43 binding fragment or derivative of the antibody). In another embodiment, the present invention provides a method for preventing, alleviating and / or treating TDP-43 proteinopathy. According to some embodiments, a binding molecule as described herein that is specific for TDP-43, particularly an antibody of the present invention or an antigen-binding fragment thereof, is administered to treat, alleviate and / or prevent frontotemporal degeneration (FTD) or amyotrophic lateral sclerosis (ALS). In another embodiment, a binding molecule described herein that is specific for TDP-43, in particular an antibody or antigen-binding fragment thereof of the invention, is administered to prevent, alleviate and / or treat a neurodegenerative disease selected from the group consisting of frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD, including sporadic and familial forms of AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE), and limbic-dominant age-related TDP-43 encephalopathy (LATE).

[0179] In another embodiment, a binding molecule as described herein, in particular an antibody or antigen-binding fragment thereof, that is specific for TDP-43 is administered to prevent, alleviate and / or treat a disease selected from the group consisting of frontotemporal dementia (FTD, e.g., sporadic or familial, with or without motor neuron disease (MND), with progranulin (GRN) mutation, with C9orf72 mutation, with TARDBP mutation, with valosin-containing protein (VCP) mutation, linked to chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), non-fluent variant primary progressive aphasia (nfvPPA), etc.), amyotrophic lateral sclerosis (ALS ... The patient may have a history of ALS (including sporadic and familial forms of ALS, with TARDBP mutations, with angiogenic protein (ANG) mutations), Alexander disease (AxD), limbic-predominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy, Perry syndrome, Alzheimer's disease (AD, including sporadic and familial forms of AD), Down syndrome, familial British dementia, polyglutamine diseases (Huntington disease and spinocerebellar ataxia type 3 (SCA3; also known as Mahy-Joh disease)), hippocampal sclerosis dementia and myopathies (sporadic inclusion body myositis, inclusion body myopathy with valosin-containing protein (VCP) mutations, and Paget's disease of bone and frontotemporal dementia), oculopharyngeal muscular dystrophy with rimmed vacuoles, myofibrillar myopathies with mutations in the myotilin (MYOT) gene or mutations in the gene encoding desmin (DES), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD). DETAILED DESCRIPTION

[0180] X. Definitions

[0181] As used herein, an "antigen binding molecule" is any molecule that can specifically or selectively bind to an antigen, particularly TDP-43. The binding molecule may include or may be an antibody or a fragment thereof. An anti-TDP-43 binding molecule is a molecule that binds to a TDP-43 protein at a specific recognition site (epitope), such as an anti-TDP-43 antibody or a fragment thereof. That is, the antigen binding molecule of the present invention binds to an epitope in the amino acid sequence of SEQ ID NO: 1. The antigen binding molecules provided herein, particularly antibodies or antigen binding fragments thereof, recognize full-length TDP-43. Other anti-TDP-43 binding molecules may also include multivalent molecules, multispecific molecules (e.g., diabodies), fusion molecules, aptamers, affimers, or other naturally occurring or recombinantly produced molecules. Exemplary antigen binding molecules that can be used in the present invention include antibody-like molecules. Antibody-like molecules are molecules that can exert a function by binding to a target molecule (see, for example, Current Opinion in Biotechnology 2006, 17: 653-658; Current Opinion in Biotechnology 2007, 18: 1-10; Current Opinion in Structural Biology 1997, 7: 463-469; Protein Science 2006, 15: 14-27), and include, for example, DARPin (WO 2002 / 020565), Affibody (WO 1995 / 001937), Affibody (WO 2004 / 044011; WO 2005 / 040229), Adnectin (WO 2002 / 032925) and fynomer (WO 2013 / 135588).

[0182] As used herein, the terms "anti-TDP-43 antibody" and "antibody that binds to TDP-43" or simply "antibody" refer to antibodies that are capable of binding to TDP-43 with sufficient affinity so that the antibody can be used as a diagnostic and / or therapeutic agent targeting TDP-43. In general, the term "antibody" is used in the broadest sense herein and covers a variety of antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific or biparatopic antibodies), fully human antibodies, and antibody fragments, as long as they exhibit the desired antigen binding activity. The antibodies within the present invention may also be chimeric antibodies, recombinant antibodies, antigen-binding fragments of recombinant antibodies, humanized antibodies, or antibodies displayed on the surface of phage or on the surface of chimeric antigen receptor (CAR) T cells.

[0183] An "antigen-binding fragment" of an antibody refers to a molecule that comprises a portion of an intact antibody and binds to an antigen that is bound by the intact antibody, other than an intact antibody. Some examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2; diabodies; linear antibodies; single-chain antibody molecules (e.g., scFv); and multispecific antibodies formed from antibody fragments.

[0184] An "antibody that binds to an epitope in a defined region of a protein" is an antibody that requires the presence of one or more amino acids in that region in order to bind to the protein.

[0185] In certain embodiments, an "antibody that binds to an epitope in a defined region of a protein" is identified by mutation analysis, wherein the amino acids of the protein are mutated, and the binding of the antibody to the resulting altered protein (e.g., an altered protein comprising the epitope) is determined to be at least 20% of the binding to the unaltered protein. In some embodiments, an "antibody that binds to an epitope in a defined region of a protein" is identified by mutation analysis, wherein the amino acids of the protein are mutated, and the binding of the antibody to the resulting altered protein (e.g., an altered protein comprising the epitope) is determined to be at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the binding to the unaltered protein. In certain embodiments, the binding of the antibody is determined by FACS, WB, or by a suitable binding assay such as ELISA.

[0186] The term "binding to" as used in the context of the present invention defines the binding (interaction) of at least two "antigen interaction sites" to each other. According to the present invention, the term "antigen interaction site" defines a motif of a polypeptide, i.e. a part of an antibody or antigen binding fragment of the present invention, which shows the ability to specifically interact with a specific antigen or a specific group of TDP-43 antigens. Said binding / interaction should also be understood as defining "specific recognition". According to the present invention, the term "specific recognition" means that the antibody is able to specifically interact and / or bind to at least two amino acids of TDP-43 as defined herein, in particular to at least two amino acids of amino acid residues 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411 and 140 to 200 of human TDP-43 (SEQ ID NO: 1), and even more in particular to interact / bind to at least two amino acids of amino acid residues 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411 and 140 to 200 of human TDP-43 (SEQ ID NO: 1). At least two of amino acid residues 183 to 188, 203 to 213, 204 to 208, 204 to 211, 205 to 210, 316 to 323, 358 to 361, 400 to 405, 400 to 406, or 400 to 412 of NO: 1) interact / bind.

[0187] The term "pan-TDP-43 antibody" refers to an antibody that binds to misfolded aggregated TDP-43 and non-aggregated physiological TDP-43, including monomeric TDP-43, oligomeric TDP-43, post-translationally modified TDP-43 (e.g., phosphorylated, ubiquitinated, acetylated, paraubiquitinated, and / or methylated), aggregated TDP-43, and truncated TDP-43.

[0188] The term "specifically interacts" as used according to the present invention means that the antibody or antigen-binding fragment thereof of the present invention does not or does not substantially cross-react with (poly)peptides having a similar structure. Thus, the antibodies or antigen-binding fragments thereof of the present invention specifically bind to / interact with a TDP-43 structure formed by a specific amino acid sequence among amino acid residues 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411, and 140 to 200 of human TDP-43 (SEQ ID NO: 1), and more particularly, bind to / interact with a TDP-43 structure formed by a specific amino acid sequence among amino acid residues 183 to 188, 203 to 213, 204 to 208, 204 to 211, 205 to 210, 316 to 323, 358 to 361, 400 to 405, 400 to 406, or 400 to 412 of human TDP-43 (SEQ ID NO: 1).

[0189] The cross-reactivity of the panel of antigen-binding molecules, in particular antibodies or antigen-binding fragments thereof, under investigation can be tested, for example, by evaluating the binding of said panel of antibodies or antigen-binding fragments thereof to the (poly)peptide of interest and to a number of more or less closely related (poly)peptides (structurally and / or functionally) under conventional conditions (see, for example, Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, (1988) and Using Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, (1999)). Only those constructs (i.e., antibodies, antigen-binding fragments thereof, etc.) that bind to a certain TDP-43 structure as defined herein, e.g., a specific epitope or (poly)peptide / protein of TDP-43 as defined herein, but not or not substantially to any other epitope or (poly)peptide of the same TDP-43, are considered specific for the epitope or (poly)peptide / protein of interest and are selected for further investigation according to the methods provided herein. These methods may include, inter alia, binding studies, blocking and competition studies with structurally and / or functionally closely related molecules. These binding studies may also include FACS analysis, surface plasmon resonance (SPR, e.g., using BIACORE TM ), analytical ultracentrifugation, isothermal titration calorimetry, fluorescence anisotropy, fluorescence spectroscopy, or by binding assays with radiolabeled ligands.

[0190] Thus, specificity can be determined experimentally by methods known in the art and as described herein. Such methods include, but are not limited to, Western blots, ELISA-, RIA-, ECL-, IRMA-tests and peptide scanning.

[0191] The term "monoclonal antibody" as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, that is, the individual antibodies constituting the population are identical except for possible naturally occurring mutations that may exist in small amounts. Monoclonal antibodies are highly specific and are directed against a single antigenic site. The advantage of monoclonal antibodies is that they can be synthesized by hybridoma cultures and are not substantially contaminated by other immunoglobulins. The modifier "monoclonal" indicates the characteristics of the antibody in a substantially homogeneous antibody population and should not be construed as requiring the antibody to be produced by any ad hoc method. As mentioned above, the monoclonal antibody used according to the present invention can be prepared by the hybridoma method described by Kohler, Nature 256 (1975), 495.

[0192] As used herein, the term "polyclonal antibody" refers to an antibody produced in the presence of one or more other different antibodies. Generally, polyclonal antibodies are produced by B lymphocytes in the presence of several other B lymphocytes that produce different antibodies. Typically, polyclonal antibodies are obtained directly from immunized animals.

[0193] As used herein, the term "fully human antibody" refers to an antibody that contains only human immunoglobulin protein sequences. If produced in a mouse, in a mouse cell, or in a hybridoma derived from a mouse cell, a fully human antibody may contain mouse sugar chains. Similarly, a "mouse antibody" or "rat antibody" refers to an antibody that contains only mouse / rat immunoglobulin protein sequences. Alternatively, if produced in a rat, in a rat cell, or in a hybridoma derived from a rat cell, a "fully human antibody" may contain rat sugar chains. Similarly, the term "rat antibody" refers to an antibody that contains only rat immunoglobulin sequences. Fully human antibodies can also be produced, for example, by phage display, which is a widely used screening technology that can produce and screen fully human antibodies. Phage antibodies can also be used in the context of the present invention. Phage display methods are described, for example, in US 5,403,484, US 5,969,108, and US 5,885,793. Another technology that can develop fully human antibodies involves improvements to mouse hybridoma technology. Mice are transgenic to contain human immunoglobulin loci in exchange for their own mouse genes (see, eg, US 5,877,397).

[0194] The term "chimeric antibody" refers to antibodies comprising variable regions of the invention fused or chimeric to antibody regions (e.g., constant regions) from another, human or non-human species (e.g., mouse, horse, rabbit, dog, cow, chicken).

[0195] The term antibody also relates to recombinant human antibodies, heterologous antibodies and heterohybrid antibodies. The term "recombinant (human) antibody" includes all human sequence antibodies prepared, expressed, produced or separated by recombinant means, such as antibodies isolated from animals (e.g., mice) that are genetically modified for human immunoglobulin genes; antibodies expressed using recombinant expression vectors transfected into host cells; antibodies isolated from recombinant, combinatorial human antibody libraries; or antibodies prepared, expressed, produced or separated by any other means involving splicing human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies have variable regions and constant regions (if present) derived from human germline immunoglobulin sequences. However, such antibodies can be subjected to in vitro mutagenesis (or, when using animals genetically modified for human Ig sequences, somatic mutagenesis in vivo is performed), and therefore the amino acid sequences of the VH and VL regions of the recombinant antibodies are sequences that, although derived from and related to human germline VH and VL sequences, may not be naturally present in the human antibody germline library in vivo.

[0196] "Heterologous antibody" is defined relative to the transgenic non-human organism in which such antibody is produced. The term refers to an antibody having an amino acid sequence or encoding nucleic acid sequence that corresponds to an amino acid sequence or encoding nucleic acid sequence present in an organism that does not consist of the transgenic non-human animal, and which organism is typically from a species other than the species of the transgenic non-human animal.

[0197] The term "heterohybrid antibody" refers to an antibody having light and heavy chains derived from different organisms. For example, an antibody having human heavy chains associated with mouse light chains is a heterohybrid antibody. Some examples of heterohybrid antibodies include chimeric antibodies and humanized antibodies.

[0198] The term antibody also relates to humanized antibodies. The non-human (for example, mouse or rabbit) antibody of " humanization " form is a chimeric immunoglobulin, an immunoglobulin chain, or its fragment (for example Fv, Fab, Fab', F(ab')2 or other antigen binding subsequences of antibody) comprising the minimum sequence deriving from non-human immunoglobulin. Usually, humanized antibodies are human immunoglobulins (acceptor antibodies), wherein the residues from the complementary determining region (complementary determining region, CDR) of the acceptor are replaced by the residues of the CDR from non-human species (donor antibody) (for example mouse, rat or rabbit) with desired specificity, affinity and ability. In some cases, the Fv framework residues of human immunoglobulin are replaced by corresponding non-human residues. In addition, humanized antibodies may be included in residues not found in the acceptor antibody or in the CDR or framework sequences imported. Carrying out these modifications is in order to further improve and optimize antibody performance. In general, a humanized antibody will comprise at least one, and typically substantially all, of two variable domains, wherein all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin, and all or substantially all of the FR regions are those of a human immunoglobulin consensus sequence. A humanized antibody may also comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. For further details, see: Jones et al., Nature 321 (1986), 522-525; Reichmann Nature 332 (1998), 323-327 and Presta Curr Op Struct Biol 2 (1992), 593-596.

[0199] A popular method for antibody humanization involves CDR grafting, in which a functional antigen-binding site from a non-human "donor" antibody is grafted onto a human "acceptor" antibody. CDR grafting methods are known in the art and are described in, for example, US 5,225,539, US 5,693,761, and US 6,407,213. Another related approach is to produce humanized antibodies from transgenic animals that are genetically engineered to contain one or more humanized immunoglobulin loci capable of gene rearrangement and gene conversion (see, e.g., US 7,129,084).

[0200] Thus, in the context of the present invention, the term "antibody" refers to intact immunoglobulin molecules as well as portions of such immunoglobulin molecules (i.e., "antigen-binding fragments thereof"). Furthermore, as described above, the term relates to modified and / or altered antibody molecules. The term also relates to recombinantly or synthetically produced / synthesized antibodies. The term also relates to intact antibodies and antibody fragments thereof, such as isolated light and heavy chains, Fab, Fv, Fab', Fab'-SH, F(ab')2. The term antibody also includes, but is not limited to, fully human antibodies, chimeric antibodies, humanized antibodies, CDR-grafted antibodies, and antibody constructs, such as single-chain Fv (scFv) or antibody fusion proteins.

[0201] In the context of the present invention, a "single-chain Fv" or "scFv" antibody fragment has the V H and V L domains, wherein these domains are present in a single polypeptide chain. Typically, scFv polypeptides are also H With V L A polypeptide linker is included between the domains which enables the scFv to form the desired antigen binding structure. Techniques for producing single-chain antibodies are described, for example, in Plückthun, The Pharmacology of Monoclonal Antibodies, Rosenburg and Moore eds. Springer-Verlag, NY (1994), 269-315.

[0202] As used herein, a "Fab fragment" comprises a light chain and a heavy chain C H 1 and variable region. The heavy chain of a Fab molecule cannot form a disulfide bond with another heavy chain molecule.

[0203] The "Fc" region contains two C H 2 and C H The two heavy chain fragments are connected by two or more disulfide bonds and by C H The 3 domains are held together by hydrophobic interactions.

[0204] A "Fab' fragment" comprises a light chain and a portion of a heavy chain comprising V H domain and C H 1 domain and also has a C H 1 and C H The region between the two Fab' domains allows for the formation of an interchain disulfide bond between the two heavy chains of the two Fab' fragments to form the F(ab')2 molecule.

[0205] A "F(ab')2 fragment" comprises two light chains and two heavy chains, the heavy chains being comprised of H 1 and C H The F(ab')2 fragment consists of two Fab' fragments held together by the disulfide bond between the two heavy chains.

[0206] The "Fv region" comprises the variable regions from both the heavy and light chains, but lacks the constant regions.

[0207] The antibodies, antibody constructs, antibody fragments, antibody derivatives (all of Ig origin) used according to the present invention, or their corresponding immunoglobulin chains can be further modified using conventional techniques known in the art, for example by using amino acid deletions, insertions, substitutions, additions and / or recombination and / or any other modifications known in the art, alone or in combination. Methods for introducing such modifications into DNA sequences based on the amino acid sequences of immunoglobulin chains are well known to those skilled in the art; see, for example, Sambrook et al., Molecular Cloning: A Laboratory Manual; Cold Spring Harbor Laboratory Press, 2nd edition (1989) and 3rd edition (2001). The term "Ig-derived domain" particularly relates to a (poly)peptide construct comprising at least one CDR. The listed fragments or derivatives of the Ig-derived domain define the following (polypeptide) peptides, which are part of the above antibody molecules and / or modified by chemical / biochemical or molecular biological methods. Corresponding methods are known in the art and are described, inter alia, in laboratory manuals (see, Sambrook et al., Molecular Cloning: A Laboratory Manual; Cold Spring Harbor Laboratory Press, 2nd edition (1989) and 3rd edition (2001); Gerhardt et al., Methods for General and Molecular Bacteriology ASM Press (1994); Lefkovits, Immunology Methods Manual: The Comprehensive Sourcebook of Techniques; Academic Press (1997); Golemis, Protein-Protein Interactions: A Molecular Cloning Manual Cold Spring Harbor Laboratory Press (2002)).

[0208] The term "CDR" as used herein refers to a "complementarity determining region," which is well known in the art. CDR is a portion of an immunoglobulin that determines the specificity of the molecule and contacts a specific ligand. CDR is the most variable part of the molecule and contributes to the diversity of these molecules. Three CDR regions are present in each V domain: CDR1, CDR2, and CDR3. CDR-H represents the CDR region of the variable heavy chain, while CDR-L represents the CDR region of the variable light chain. VH means variable heavy chain, and VL means variable light chain. The CDR regions of the Ig-derived regions can be determined as described in Kabat "Sequences of Proteins of Immunological Interest," 5th edit. NIH publication no. 91-3242 USDepartment of Health and Human Services (1991). The CDR sequences provided herein are defined according to Kabat. However, those skilled in the art will appreciate that the present invention is intended to encompass binding molecules in which CDR sequences are defined according to any useful identification / numbering scheme.For example, the following numbering scheme can be used to define CDRs: Chothia (Canonical structures for the hypervariable regions of immunoglobulins. Chothia C, Lesk AM. J Mol Biol. 1987 Aug 20; 196 (4): 901-17); IMGT (IMGT, the international ImMunoGeneTics database. Giudicelli V, Chaume D, Bodmer J, Müller W, Busin C, Marsh S, Bontrop R, Marc L, Malik A, Lefranc MP. Nucleic Acids Res. 1997 Jan 1; 25(1): 206-11 and Unique database numbering system for immunogenetic analysis. Lefranc MP. ImmunolToday. 1997 Nov; 18(11): 509); MacCallum (MacCallum RM, Martin AC, Thornton JM, J MolBiol. 1996 Oct 11;262(5):732-45) and Martin (Abhinandan KR, Martin ACR. Analysis and improvements to Kabat and structurally correct numbering of antibody variable domains. Mol Immunol. (2008) 45:3832-9.10.1016 / j.molimm.2008.05.022).

[0209] Therefore, in the context of the present invention, the antibody molecules described herein above are selected from the group consisting of whole antibodies (immunoglobulins, such as IgG1, IgG2, IgG2a, IgG2b, IgA1, IgGA2, IgG3, IgG4, IgA, IgM, IgD or IgE), F(ab)-, Fab'-SH-, FV-, Fab'-, F(ab')2-fragments, chimeric antibodies, CDR-grafted antibodies, fully human antibodies, divalent antibody constructs, antibody fusion proteins, synthetic antibodies, divalent single chain antibodies, trivalent single chain antibodies and multivalent single chain antibodies.

[0210] "Humanization method " is well known in the art, and is particularly directed to antibody molecules, such as molecules in Ig origin are described.Term " humanization " refers to the humanized form of non-human (for example, mouse) antibody or its fragment (for example Fv, Fab, Fab ', F (ab '), scFv or other antigen-binding portion sequences of antibody) comprising some parts of the sequence deriving from non-human antibody.Humanized antibody includes human immunoglobulin, wherein the residue from human immunoglobulin complementary determining region (CDR) is replaced by the residue from the CDR of non-human species (such as mouse, rat or rabbit) with desired binding specificity, affinity and ability.In general, humanized antibody will comprise at least one, and generally substantially all in two variable domains, wherein all or substantially all of CDR districts correspond to those of non-human immunoglobulin, and all or substantially all of FR districts are those of human immunoglobulin consensus sequence. Optimally, the humanized antibody will also comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin; see, inter alia, Jones et al., Nature 321 (1986), 522-525, Presta, Curr. Op. Struct. Biol. 2 (1992), 593-596. Methods for humanizing non-human antibodies are well known in the art. Typically, a humanized antibody has one or more amino acids introduced therein from a non-human source, while retaining the original binding activity of the antibody. Methods for humanizing antibodies / antibody molecules are also described in detail in Jones et al., Nature 321 (1986), 522-525; Reichmann et al., Nature 332 (1988), 323-327; and Verhoeyen et al., Science 239 (1988), 1534-1536. Specific examples of humanized antibodies, such as antibodies against EpCAM, are known in the art (see, for example, LoBuglio, Proceedings of the American Society of Clinical Oncology Abstract (1997), 1562 and Khor, Proceedings of the American Society of Clinical Oncology Abstract (1997), 847).

[0211] Thus, in the context of the present invention, antibody molecules or antigen-binding fragments thereof are provided, which are humanized and can be successfully used in pharmaceutical compositions.

[0212] The specificity of the antibodies or antigen-binding fragments of the present invention can be expressed not only by the properties of the amino acid sequence of the antibody or antigen-binding fragment as defined above, but also by the epitope to which the antibody is capable of binding. Therefore, in one embodiment, the present invention relates to anti-misfolded TDP-43 antibodies or antigen-binding fragments thereof that recognize the same epitope as the antibodies of the present invention.

[0213] Those skilled in the art will appreciate that an epitope can be contained within the TDP-43 protein, but can also be contained in its degradation products or can be a chemically synthesized peptide. Amino acid positions are indicated only to illustrate the location of the corresponding amino acid sequence within the TDP-43 protein sequence. The present invention encompasses all peptides comprising the epitope. The peptide can be a portion of a polypeptide greater than 100 amino acids in length, or it can be a small peptide of less than 100, preferably less than 50, more preferably less than 25, and even more preferably less than 16 amino acids. The amino acids of such a peptide can be natural amino acids or unnatural amino acids (e.g., β-amino acids, γ-amino acids, D-amino acids), or a combination thereof. Furthermore, the present invention encompasses retro-inverso peptides corresponding to the epitope. The peptide can be unconjugated or conjugated. It can be conjugated to, for example, a small molecule (e.g., a drug or fluorophore), a high molecular weight polymer (e.g., polyethylene glycol (PEG), polyethyleneimine (PEI), hydroxypropylmethacrylate (HPMA), etc.), or a protein, fatty acid, or sugar moiety, or can be incorporated into a membrane.

[0214] Whether identical epi-position is recognized in order to test the antibody in question and the antibody of the present invention, following competition study can be carried out: the Vero cell that will be infected with 3 kinds of MOI (multiplicity of infection) is hatched 1 hour with the antibody in question as competitor of different concentrations after 20h.In the second incubation step, antibody of the present invention is applied with the constant concentration of 100nM, and use the fluorescent-labeled antibody for the constant domain of antibody of the present invention, detect its combination by flow cytometry.To be inversely proportional (inversely proportional) with the concentration of the antibody in question, carry out in conjunction with indicating two kinds of antibody recognition identical epi-positions.Yet, many other mensurations as known in the art can be used.

[0215] The present invention also relates to the production of specific antibodies against native and recombinant TDP-43 polypeptides. This production is based, for example, on immunization of animals such as mice. However, other animals for producing antibodies / antisera are also contemplated in the present invention. For example, monoclonal and polyclonal antibodies can be produced from rabbits, mice, goats, donkeys, etc. A polynucleotide encoding a corresponding selected polypeptide of TDP-43 can be subcloned into a suitable vector, wherein the recombinant polypeptide is expressed in an organism capable of expression, such as bacteria. Thus, the expressed recombinant protein can be injected intraperitoneally into mice, and the resulting specific antibodies can be obtained, for example, from mouse serum provided by intracardiac blood puncture. The present invention also contemplates the production of specific antibodies against native and recombinant polypeptides using DNA vaccine strategies, as exemplified in the accompanying Examples. DNA vaccine strategies are well known in the art and encompass liposome-mediated delivery, injection by gene gun or jet, and intramuscular or intradermal injection. Therefore, antibodies against polypeptides or proteins or epitopes of TDP-43, in particular the antibody epitopes provided herein, can be obtained by directly immunizing animals with a vector expressing the desired polypeptide or protein or epitope of TDP-43, in particular the following antibody epitopes of the present invention, which are located at amino acid residues 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411 and 140 to 200 of SEQ ID NO: 1; more particularly the following antibody epitopes of the present invention, which are located at amino acid residues 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, 397 to 411 and 140 to 200 of SEQ ID NO: 1. The amount of specific antibody obtained can be quantified using ELISA, which is also described below. Additional methods for generating antibodies are known in the art, see, for example, Harlow and Lane, "Antibodies, A Laboratory Manual," CSH Press, Cold Spring Harbor, 1988.

[0216] Thus, under specified assay conditions, a specific antibody binds to the corresponding epitope of TDP-43 and to each other, without binding to other components present in the sample in significant amounts. Specific binding to a target analyte under such conditions may require a binding moiety selected for its specificity for the specific target analyte. A variety of immunoassay formats can be used to select antibodies that specifically react with a particular antigen. For example, solid-phase ELISA immunoassays are routinely used to select monoclonal antibodies that specifically immunoreact with the analyte. See Shepherd and Dean (2000), Monoclonal Antibodies: A Practical Approach, Oxford University Press and / or Howard and Bethell, for a description of immunoassay formats and conditions that can be used to determine specific immunoreactivity. Generally speaking, a specific or selective reaction will be at least twice the background signal-to-noise ratio, and more typically more than 10 to 100 times the background. One skilled in the art will be able to provide and generate specific binding molecules for a novel polypeptide. For specific binding assays, it is readily possible to avoid undesired cross-reactivity, for example, polyclonal antibodies can be readily purified and selected by known methods (see Shepherd and Dean, loc. cit.).

[0217] The "class" of an antibody refers to the type of constant domain or constant region possessed by its heavy chain. There are five major classes of antibodies: IgA, IgD, IgE, IgG, and IgM, and several of these can be further divided into subclasses (isotypes), such as IgG1, IgG2, IgG2a, IgG2b, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant domains corresponding to the different classes of immunoglobulins are called α, δ, ε, γ, and μ, respectively.

[0218] In certain embodiments, the amino acid sequence variants of the antibodies provided herein are contemplated. For example, it is desirable to improve the binding affinity and / or other biological properties of the antibody. The amino acid sequence variants of the antibody can be prepared by introducing suitable modifications into the nucleotide sequence encoding the antibody, or by peptide synthesis. Such modifications include, for example, the disappearance of residues in the antibody amino acid sequence and / or the insertion and / or its replacement therein. Any combination of disappearance, insertion and replacement can be performed to obtain the final construct, provided that the final construct has desired characteristics, such as antigen binding.

[0219] In certain embodiments, antibody variants with one or more amino acid replacements are provided. The target site of replacement mutagenesis includes CDR and FR. Conservative replacements are shown under the heading of "preferred replacements" in Table 1. More variations are provided under the heading of "exemplary replacements" in Table 1, and are further described below with reference to amino acid side chain categories. Amino acid replacements can be introduced into the target antibody, and for desired activity, for example, retained / improved antigen binding, reduced immunogenicity or improved ADCC or CDC screening products.

[0220] Table 1

[0221] Original residue Exemplary Replacements Preferred replacement Ala(A) Val; Leu; Ile Val Arg(R) Lys; Gln; Asn Lys Original residue Exemplary Replacements Preferred replacement Asn(N) Gln; His; Asp, Lys; Arg Gin Asp(D) Glu; Asn Glu Cys(C) Scr;Ala Scr Gin(Q) Asn;Glu Asn Glu(E) Asp; Gln Asp

[0222] Gly(G) Ala Ala His(H) Asn; Gln; Lys; Arg Arg Ile(I) Leu; Val; Met; Ala; Phe; norleucine Leu Lcu(L) Norleucine; Ilc; Val; Met; Ala; Phe Ile Lys(K) Arg; Gln; Asn Arg Met(M) Leu; Phe; Ile Leu Phc(F) Trp; Leu; Val; Ilc; Ala; Tyr Tyr Pro(P) Ala Ala Ser(S) Thr Thr Thr(T) Val; Ser Ser Trp(W) Tyr; Phe Tyr Tyr(Y) Trp; Phe; Thr, Ser Phe Val(V) Ilc; Leu; Met; Phe; Ala; norleucine Lcu

[0223] Amino acids can be grouped according to common side chain properties:

[0224] (1) Hydrophobicity: norleucine, Met, Ala, Val, Leu, Ile;

[0225] (2) Neutral hydrophilicity: Cys, Ser, Thr, Asn, Gln;

[0226] (3) Acidic: Asp, Glu;

[0227] (4) Basic: His, Lys, Arg;

[0228] (5) Residues that affect chain orientation: Gly, Pro;

[0229] (6) Aromatic: Trp, Tyr, Phe.

[0230] Non-conservative substitutions will entail exchanging a member of one of these classes for another class.

[0231] A type of replacement variant relates to one or more hypervariable region residues that replace a parent antibody (e.g., humanized or human antibody). Typically, the resulting variant selected for further study will have improvements (e.g., improvements) (e.g., improved affinity, reduced immunogenicity) and / or will substantially retain some biological properties of the parent antibody relative to the parent antibody in some biological properties. An exemplary replacement variant is an affinity matured antibody, which can, for example, be readily produced using affinity maturation techniques based on phage display, such as those described herein. In brief, one or more CDR residues are mutated, the variant antibody is displayed on phage and screened for specific biological activity (e.g., binding affinity).

[0232] Changes (e.g., substitutions) can be made in the CDRs, for example, to improve antibody affinity. Such changes can be made in CDR "hotspots," residues encoded by codons that mutate at high frequency during the somatic maturation process (see, e.g., Chowdhury, Methods Mol. Biol. 207: 179-196 (2008)), and / or SDRs (a-CDRs), and the resulting variant VH or VL tested for binding affinity. Affinity maturation by constructing and reselecting from secondary libraries has been described, for example, in Hoogenboom et al., Methods in Molecular Biology 178: 1-37 (O'Brienet al., ed., Human Press, Totowa, NJ, (2001)). In some embodiments of affinity maturation, diversity is introduced into the variable genes selected for maturation by any of a variety of methods (e.g., error-prone PCR, chain shuffling, or oligonucleotide-directed mutagenesis). A secondary library is then created. The library is then screened to identify any antibody variants with the desired affinity. Another method for introducing diversity involves a CDR-guided approach in which several CDR residues (e.g., 4 to 6 residues at a time) are randomized. CDR residues involved in antigen binding can be specifically identified, for example, using alanine scanning mutagenesis or modeling. In particular, CDR-H3 and CDR-L3 are typically targeted.

[0233] In certain embodiments, substitutions, insertions, or deletions may occur in one or more CDRs, as long as such changes do not substantially reduce the ability of the antibody to bind to the antigen. For example, conservative changes that do not substantially reduce binding affinity (e.g., conservative substitutions as provided herein) may be made in the CDRs. Such changes may be outside the CDR "hot spots" or SDRs. In certain embodiments of the variant VH and VL sequences provided above, each CDR is unaltered or comprises no more than one, two, or three amino acid substitutions.

[0234] A useful method for identifying residues or regions of an antibody that can be targeted for mutagenesis is called "alanine scanning mutagenesis," as described by Cunningham and Wells (1989) Science, 244: 1081-1085. In this method, residues or groups of target residues (e.g., charged residues such as Arg, Asp, His, Lys, and Glu) are identified and replaced with neutral or negatively charged amino acids (e.g., alanine or polyalanine) to determine whether the interaction of the antibody with the antigen is affected. Additional substitutions can be introduced at amino acid positions that demonstrate functional sensitivity to the initial substitutions. Alternatively or in addition, crystal structures of antigen-antibody complexes are used to identify contact points between the antibody and the antigen. Such contact residues and neighboring residues can be targeted or eliminated as candidates for substitution. Variants can be screened to determine whether they contain the desired properties.

[0235] Amino acid sequence insertions include amino and / or carboxyl terminal fusions of polypeptides ranging in length from one residue to 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Some examples of terminal insertions include antibodies with an N-terminal methionyl residue. Other insertion variants of the antibody molecule include fusions of the N- or C-terminus of the antibody with an enzyme (e.g., for ADEPT) or a polypeptide that increases the serum half-life of the antibody.

[0236] In certain embodiments, the antibodies provided herein are altered to increase or decrease the extent to which the antibodies are glycosylated. Addition or deletion of glycosylation sites to an antibody can be conveniently achieved by altering the amino acid sequence to create or remove one or more glycosylation sites.

[0237] When an antibody comprises an Fc region, the carbohydrates attached thereto may be altered. Natural antibodies produced by mammalian cells typically contain branched biantennary oligosaccharides, typically linked to Ash297 of the CH2 domain of the Fc region via an N-linkage. See, e.g., Wright et al., TIBTECH 15: 26-32 (1997). Oligosaccharides may include a variety of carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose linked to GlcNAc in the "stem" of the biantennary oligosaccharide structure. In some embodiments, the oligosaccharides in the antibodies of the present invention may be modified to produce antibody variants with certain improved properties.

[0238] In one embodiment, antibody variants are provided having carbohydrate structures lacking fucose attached to the Fc region (directly or indirectly). For example, the amount of fucose in such an antibody can be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is determined by calculating the average amount of fucose in the sugar chains of Ash297 relative to the sum of all sugar structures (e.g., complex, hybrid, and high mannose structures) attached to Asn 297, as measured by MALDI-TOF mass spectrometry, for example, as described in WO 2008 / 077546. Ash297 refers to the asparagine residue located at approximately position 297 in the Fc region (Eu numbering of Fc region residues; see Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)); however, due to minor sequence variations in antibodies, Ash297 may also be located approximately ±3 amino acids upstream or downstream of position 297, i.e., at positions 294 and 300. Such fucosylation variants may have improved ADCC function. See, for example, U.S. Patent Publication Nos. US 2003 / 0157108 (Presta, L.); US 2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd.). Some examples of publications related to "defucosylated" or "fucose-deficient" antibody variants include: US 2003 / 0157108; WO 2000 / 61739; WO 2001 / 29246; US 2003 / 0115614; US 2002 / 0164328; US 2004 / 0093621; US ​​2004 / 0132140; US 2004 / 0110704; US 2004 / 0110282; US 2004 / 0109865; WO 2003 / 085119; WO 2003 / 084570; WO 2005 / 035586; WO 2005 / 035778; WO2005 / 053742; WO2002 / 031140; Okazaki et al., J. Mol. Biol. 336: 1239-1249 (2004); Yamane-Ohnuki et al., Biotech. Bioeng. 87: 614 (2004).Some examples of cell lines capable of producing defucosylated antibodies include protein fucosylation-deficient Lec13 CHO cells (Ripka et al., Arch. Biochem. Biophys. 249: 533-545 (1986); U.S. Patent Application No US 2003 / 0157108 A1, Presta, L; and WO 2004 / 056312 A1, Adams et al., particularly in Example 11), and knockout cell lines, such as α-1,6-fucosyltransferase gene FUT8 knockout CHO cells (see, e.g., Yamane-Ohnuki et al., Bioteeh. Bioeng. 87: 614 (2004); Kanda, Y. et al., Bioteehnol. Bioeng., 94(4): 680-688 (2006); and WO 2003 / 085107).

[0239] Also provided are antibody variants having bisected oligosaccharides, for example, wherein the biantennary oligosaccharide attached to the Fc region of the antibody is bisected by GlcNAc. Such antibody variants may have reduced fucosylation and / or improved ADCC function. Some examples of such antibody variants are described in, for example, WO 2003 / 011878 (Jean-Mairet et al.); U.S. Patent No. 6,602,684 (Umana et al.); and US 2005 / 0123546 (Umana et al.). Also provided are antibody variants having at least one galactose residue in the oligosaccharide attached to the Fc region. Such antibody variants may have improved CDC function. Such antibody variants are described in, for example, WO 1997 / 30087 (Patel et al.); WO 1998 / 58964 (Raju, S.); and WO 1999 / 22764 (Raju, S.).

[0240] In certain embodiments, one or more amino acid modifications can be introduced into the Fc region of an antibody provided herein, thereby generating an Fc region variant. An Fc region variant can be included in a human Fc region sequence (e.g., human IgG1, IgG2, IgG3, or IgG Fc region) comprising an amino acid modification (e.g., replacement) at one or more amino acid positions.

[0241] In certain embodiments, the present invention contemplates antibody variants that possess some, but not all, effector functions, making them desirable candidates for applications where the in vivo half-life of the antibody is important but certain effector functions (e.g., complement activation and ADCC) are unnecessary or deleterious. In vitro and / or in vivo cytotoxicity assays can be performed to determine reduction / depletion of CDC and / or ADCC activity. For example, Fc receptor (FcR) binding assays can be performed to ensure that the antibody lacks FcγR binding (and therefore may lack ADCC activity), but retains FcRn binding ability. NK cells, the primary cells mediating ADCC, express only FcγRIII, while monocytes and microglia express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol. 9: 457-492 (1991). Some non-limiting examples of in vitro assays for assessing ADCC activity of a molecule of interest are described in U.S. Pat. Nos. 5,500,362 (see, e.g., Hellstrom, I. et al., Proc. Nat'l Acad. Sci. USA 83:7059-7063 (1986)) and Hellstrom, I et al., Proc. Nat'l Acad. Sci. USA 82:1499-1502 (1985); 5,821,337 (see Bruggemann, M. et al., J. Exp. Med. 166:1351-1361 (1987)).

[0242] Alternatively, non-radioactive assays can be used (see, for example, the ACTI™ non-radioactive cytotoxicity assay for flow cytometry (Cell Technology, Inc. Mountain View, CA); and the CytoTox Non-radioactive cytotoxicity assay (Promega, Madison, WI). Useful effector cells for such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells.

[0243] Alternatively or additionally, ADCC activity of the molecule of interest can be assessed in vivo, eg, in a animal model such as that disclosed in Clynes et al., Proc. Nat'l Acad. sci. USA 95:652-656 (1998).

[0244] C1q binding assays can also be performed to determine that the antibody cannot bind to C1q and, therefore, lacks CDC activity. See, e.g., C1q and C3c binding ELISAs in WO 2006 / 029879 and WO 2005 / 100402. To assess complement activation, a CDC assay can be performed (see, e.g., Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996); Cragg, MS et al., Blood 101:1045-1052 (2003); and Cragg, MS and MJ Glennie, Blood 103:2738-2743 (2004)). Determination of FcRn binding and in vivo clearance / half-life can also be performed using methods known in the art (see, eg, Petkova, SB et al., Int'l. Immunol. 18(12): 1759-1769 (2006)).

[0245] Antibodies with reduced effector function include those with substitutions at one or more of residues 238, 265, 269, 270, 297, 327, and 329 in the Fc region ( U.S. Pat. No. 6,737,056 ). Such Fc mutants include those with substitutions at two or more of amino acids 265, 269, 270, 297, and 327, including the so-called "DANA" Fc mutant in which residues 265 and 297 are substituted with alanine ( U.S. Pat. No. 7,332,581 ). Alternatively, antibodies with reduced effector function include antibodies in which one or more of residues 234, 235, and 329 of the Fc region are substituted, i.e., residues 234 and 235 are substituted with alanine, and residue 329 is substituted with glycine, so-called "PG-LALA" Fc mutants (Lo, M. et al., Journal of Biochemistry, 292, 3900-3908).

[0246] Certain antibody variants with improved or diminished binding to FcRs have been described (see, e.g., U.S. Patent No. 6,737,056; WO 2004 / 056312, and Shields et al., J. Biol. Chem. 9(2): 6591-6604 (2001)).

[0247] In certain embodiments, the antibody variant comprises an Fc region with one or more amino acid substitutions that improve ADCC, eg, substitutions at positions 298, 333, and / or 334 of the Fc region (EU numbering of residues).

[0248] In some embodiments, alterations are made in the Fc region that result in altered (i.e., improved or reduced) C1q binding and / or complement-dependent cytotoxicity (CDC), e.g., as described in U.S. Pat. No. 6,194,551, WO 99 / 51642, and Idusogie et al., J. Immunol. 164:4178-4184 (2000).

[0249] Antibodies with increased half-life and improved binding to the neonatal Fc receptor (FcRn), which is responsible for the transfer of maternal IgG to the fetus (Guyer et al., J. Immunol. 117: 587 (1976) and Kim et al., J. Immunol. 24: 249 (1994)), are described in US 2005 / 0014934A1 (Hinton et al.). These antibodies comprise an Fc region having one or more substitutions therein that improve binding of the Fc region to FcRn. Such Fc variants include Fc variants in which one or more of the following Fc region residues are replaced: 238, 256, 265, 272, 286, 303, 305, 307, 311, 312, 317, 340, 356, 360, 362, 376, 378, 380, 382, ​​413, 424, or 434, e.g., an Fc variant in which Fc region residue position 434 is replaced ( U.S. Pat. No. 7,371,826 ). See also Duncan & Winter, Nature 322:738-40 (1988); U.S. Patent No. 5,648,260; U.S. Patent No. 5,624,821; and WO 94 / 29351 for further examples of Fc region variants.

[0250] In certain embodiments, it may be desirable to produce cysteine ​​engineered antibodies, such as "thioMAbs," in which one or more residues of an antibody are replaced by cysteine ​​residues. In some specific embodiments, the replaced residues occur at accessible sites of the antibody. By replacing these residues with cysteine, reactive sulfhydryl groups are thereby located at accessible sites of the antibody and can be used to conjugate the antibody to other moieties, such as drug moieties or linker-drug moieties, to produce immunoconjugates, as further described herein. In certain embodiments, any one or more of the following residues may be replaced by cysteine: V205 (Kabat numbering) of the light chain; A118 (EU numbering) of the heavy chain; and S400 (EU numbering) of the heavy chain Fc region. Cysteine ​​engineered antibodies can be produced as described, for example, in U.S. Patent No. 7,521,541.

[0251] In certain embodiments, the antibodies provided herein may also be modified to include additional non-proteinaceous moieties known in the art and readily available. Suitable moieties for antibody derivatization include, but are not limited to, water-soluble polymers. Some non-limiting examples of water-soluble polymers include, but are not limited to, polyethylene glycol (PEG), copolymers of ethylene glycol / propylene glycol, carboxymethyl cellulose, dextran, polyvinyl alcohol, polyvinyl pyrrolidone, poly-1,3-dioxolane, poly-1,3,6-tri ... In some embodiments, the present invention provides the polyols of formula (I) or (II) for example, polyols, e.g. ...

[0252] In another embodiment, a conjugate of an antibody with a nonproteinaceous moiety that can be selectively heated by exposure to radiation is provided. In one embodiment, the nonproteinaceous moiety is a carbon nanotube (Kam et al., Proc. Natl. Acad. Sci. USA 102: 11600-11605 (2005)). The radiation can be of any wavelength, including but not limited to, a wavelength that does not damage normal cells but heats the nonproteinaceous moiety to a temperature that kills cells adjacent to the antibody-nonproteinaceous moiety.

[0253] Antibodies can be produced using recombinant methods and compositions, for example, as described in U.S. Patent No. 4,816,567. In one embodiment, an isolated nucleic acid encoding an anti-misfolded TDP-43 antibody described herein is provided. Such a nucleic acid may encode an amino acid sequence comprising the VL of the antibody and / or an amino acid sequence comprising the VH of the antibody (e.g., the light chain and / or heavy chain of the antibody). In another embodiment, one or more vectors (e.g., expression vectors) comprising such a nucleic acid are provided. In another embodiment, a host cell comprising such a nucleic acid is provided. In one such embodiment, the host cell comprises (e.g., has been transformed with): (1) a vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and an amino acid sequence comprising the VH of the antibody; or (2) a first vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and a second vector comprising a nucleic acid encoding an amino acid sequence comprising the VH of the antibody. In one embodiment, the host cell is eukaryotic, for example, a Chinese hamster ovary (CHO) cell or a lymphoid cell (e.g., YO, NSO, Sp20). In one embodiment, a method for preparing an anti-misfolded TDP-43 antibody is provided, wherein the method comprises: culturing a host cell comprising a nucleic acid encoding the antibody as provided above under conditions suitable for expression of the antibody, and optionally recovering the antibody from the host cell (or host cell culture medium).

[0254] For recombinant production of anti-misfolded TDP-43 antibodies, nucleic acid encoding the antibody, e.g., as described above, is isolated and inserted into one or more vectors for further cloning and / or expression in a host cell or cell-free expression system. Such nucleic acid can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that specifically bind to genes encoding the heavy and light chains of the antibody).

[0255] Suitable host cells for cloning or expressing antibody encoding vectors include prokaryotic or eukaryotic cells as described herein. For example, antibodies can be produced in bacteria, particularly when glycosylation and Fc effector functions are not required. For expression of antibody fragments and polypeptides in bacteria, see, for example, U.S. Patent Nos. 5,648,237, 5,789,199, and 5,840,523. (See also Charlton, Methods in Molecular Biology, Val. 248 (BKCLo, ed., Humana Press, Totowa, NJ, 2003), pp. 245-254, describing expression of antibody fragments in Escherichia coli (E. coli). After expression, the antibodies in the soluble fraction can be isolated from the bacterial cell paste and can be further purified.

[0256] In addition to prokaryotes, eukaryotic microorganisms such as filamentous fungi or yeast are also suitable cloning or expression hosts for antibody encoding vectors, including fungi and yeast strains in which the glycosylation pathway has been "humanized" to produce antibodies with partially or fully human glycosylation patterns. See Gerngross, Nat. Biotech. 22: 1409-1414 (2004) and Li et al., Nat. Biotech. 24: 210-215 (2006).

[0257] Suitable host cells for expressing glycosylated antibodies also come from multicellular organisms (invertebrates and vertebrates). Some examples of invertebrate cells include plant and insect cells. Many baculovirus strains have been identified that can be used in conjunction with insect cells, particularly for transfecting Spodoptera frugiperda cells.

[0258] Plant cell cultures can also be used as hosts. See, for example, U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (describing plant cell cultures for producing antibodies in transgenic plants). TM technology).

[0259] Vertebrate cells can also be used as hosts. For example, mammalian cell lines suitable for suspension culture can be used. Other examples of useful mammalian host cell lines are macaque kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney cell line (293 or 293 cells, as described, e.g., in Graham et al., J. Gen Viral. 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (TM4 cells, as described, e.g., in Mather, Biol. Reprod. 23:243-251 (1980)); macaque kidney cells (CV 1); African green macaque kidney cells (VER0-76); human cervical carcinoma cells (HeLa); canine kidney cells (MDCK; Buffalo rat liver cells (BRL 3A); human lung cells (W138); human hepatocytes (Hep G2); mouse mammary tumor (MMT 060562); TRI cells, as described, e.g., in Mather et al., Annals N.Y. Aead. Sei. 383:44-68 (1982); MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include Chinese hamster ovary (CHO) cells, including DHFR CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)); and myeloma cell lines, such as YO, NSO, and Sp2 / 0. For a review of certain mammalian host cell lines suitable for antibody production, see, for example, Yazaki and Wu, Methods in Molecular Biology, Val. 248 (BKCLo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0260] The method for producing the TDP-43 binding molecules, in particular antibodies, of the present invention may comprise the following steps:

[0261] a. culturing suitable host cells or cell-free expression systems under conditions suitable for producing binding molecules, particularly antibodies; and

[0262] b. Isolation of binding molecules, particularly antibodies. Suitable culture and isolation techniques are available to the skilled person.

[0263] The anti-TDP-43 antibodies provided herein can be identified, screened for their physical / chemical properties and / or biological activities, or characterized by a variety of assays known in the art.

[0264] In one aspect, the antibodies of the invention are detected by known methods, such as ELISA, Its antigen binding activity was tested by FACS, immunofluorescence or immunohistochemistry.

[0265] In another aspect, competition assays can be used to identify antibodies that compete with any of the antibodies described herein for binding to TDP-43. In certain embodiments, such competing antibodies bind to the same epitope (e.g., a linear or conformational epitope) as the antibodies described herein. Detailed exemplary methods for mapping epitopes bound to antibodies are provided in Morris (1996) "Epitope Mapping Protocols," Methods in Molecular Biology vol. 66 (Humana Press, Totowa, NJ).

[0266] In one exemplary competition assay, immobilized TDP-43 is incubated in a solution containing a first labeled antibody (e.g., any of the antibodies described herein) that binds to TDP-43 and a second unlabeled antibody that is tested for its ability to compete with the first antibody for binding to TDP-43. As a control, immobilized TDP-43 is incubated in a solution containing the first labeled antibody but not the second unlabeled antibody. Following incubation under conditions permissive for binding of the first antibody to TDP-43, excess unbound antibody is removed, and the amount of label associated with immobilized TDP-43 is measured. If the amount of label associated with immobilized TDP-43 in the test sample is significantly reduced relative to the control sample, this indicates that the second antibody is competing with the first antibody for binding to TDP-43. See Harlow and Lane (1988) Antibodies: A Laboratory Manual ch. 14 (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

[0267] The present invention also provides immunoconjugates comprising an anti-TDP-43 antibody provided herein conjugated to one or more therapeutic agents, such as a chemotherapeutic agent or drug, a growth inhibitory agent, a toxin (e.g., a protein toxin, an enzymatically active toxin of bacterial, fungal, plant, or animal origin, or a fragment thereof), a radioactive isotope (i.e., a radioconjugate), a blood-brain barrier penetrating moiety, or a detectable label.

[0268] In another aspect of the present invention, an article of manufacture comprising materials useful for treating, preventing, and / or diagnosing the aforementioned diseases, disorders, abnormalities, or TDP-43 proteinopathies, particularly those associated with TDP-43 aggregates, is provided. The article of manufacture comprises a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, intravenous (IV) solution bags, and the like. The container can be formed from a variety of materials, such as glass or plastic. The container holds a composition that is effective for treating, preventing, and / or diagnosing a condition, either alone or in combination with another composition, and can have a sterile access port (e.g., the container can be an IV solution bag or a vial with a stopper pierceable by a hypodermic needle). At least one active agent in the composition is an antibody of the present invention. The label or package insert indicates that the composition is used to treat a selected condition. Furthermore, the article of manufacture may comprise: (a) a first container comprising a composition comprising an antibody of the present invention; and (b) a second container comprising a composition comprising an additional therapeutic agent. The article of manufacture in this embodiment of the invention may further comprise a package insert indicating that the composition is useful for treating a specific condition. Alternatively or additionally, the article of manufacture may further comprise a second (or third) container comprising a pharmaceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer's solution, and dextrose solution. It may also contain other materials desirable from a commercial and user perspective, including other buffers, diluents, filters, needles, and syringes.

[0269] It will be understood that any of the above articles of manufacture may comprise an immunoconjugate of the invention in place of or in addition to an anti-TDP-43 antibody.

[0270] XI. Exemplary TDP-43-Specific Binding Molecules or Antibodies

[0271] In some embodiments of the invention, the antibody comprises:

[0272] a) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 15, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 17; or

[0273] b) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VH-CDR3 comprising the amino acid sequence ES(Glu-Ser), a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 25, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 27; or

[0274] c) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 31, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 32, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 33, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 37; or

[0275] d) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 41, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 42, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 43, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 45, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 46, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 47; or

[0276] e) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 61, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 62, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 63, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 65, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 66, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 67; or

[0277] f) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 71, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 72, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 73, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 75, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 77; or

[0278] g) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 81, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 82, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 83, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 85, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 86, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 87; or

[0279] h) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 101, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 102, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 103, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 105, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 106, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 107; or

[0280] i) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 121, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 122, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 123, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 125, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 16, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 127; or

[0281] j) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 141, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 142, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 143, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 145, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 146, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 147; or

[0282] k) a VH-CDR1 comprising the amino acid sequence of SEQ ID NO: 151, a VH-CDR2 comprising the amino acid sequence of SEQ ID NO: 152, and a VH-CDR3 comprising the amino acid sequence of SEQ ID NO: 153, a VL-CDR1 comprising the amino acid sequence of SEQ ID NO: 155, a VL-CDR2 comprising the amino acid sequence of SEQ ID NO: 156, and a VL-CDR3 comprising the amino acid sequence of SEQ ID NO: 157.

[0283] In some embodiments, the antibody comprises:

[0284] a. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 10 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 14; or

[0285] b. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 20 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 24; or

[0286] c. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 30 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 34; or

[0287] d. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 40 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 44; or

[0288] e. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 60 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 64; or

[0289] f. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 70 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 74; or

[0290] g. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 80 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 84; or

[0291] h. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 100 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 104; or

[0292] i. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 120 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 124; or

[0293] j. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 140 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 144; or

[0294] k. A heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 150 and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 154.

[0295] In some embodiments, the antibody comprises:

[0296] a. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 10, or a heavy chain variable region (VH) having at least 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 10, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 14, or a light chain variable region (VL) having at least 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 14; or

[0297] b. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 20, or a heavy chain variable region (VH) having at least 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 20, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 24, or a light chain variable region (VL) having at least 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 24; or

[0298] c. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 30, or a heavy chain variable region (VH) having at least 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 30, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 34, or a light chain variable region (VL) having at least 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 34; or

[0299] d. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 40, or a heavy chain variable region (VH) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 40, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 44; or

[0300] e. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 60, or a heavy chain variable region (VH) having at least 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 60, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 64, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 64; or

[0301] f. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 70, or a heavy chain variable region (VH) having at least 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 70, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 74, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 74; or

[0302] g. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 80, or a heavy chain variable region (VH) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 80, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 84, or a light chain variable region (VL) having at least 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 84; or

[0303] h. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 100, or a heavy chain variable region (VH) having at least 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 100, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 104, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 104; or

[0304] i. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 120, or a heavy chain variable region (VH) having at least 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 120, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 124, or a light chain variable region (VL) having at least 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 124; or

[0305] j. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 140, or a heavy chain variable region (VH) having at least 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 140, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 144; or

[0306] k. a heavy chain variable region (VH) comprising the sequence of SEQ ID NO: 150, or a heavy chain variable region (VH) having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 150, and a light chain variable region (VL) comprising the sequence of SEQ ID NO: 154, or a light chain variable region (VL) having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 154.

[0307] In some embodiments, the invention relates to antibodies derived from hybridoma clones 631B2A2, 633B12C8, 634H10H7, 636E5B8, 641H1E7, 642A10B11, 642D12B4, 646B7F7, 712A6B10, 809D9C2, or 809F12D8, as further described herein.

[0308] In some embodiments, the invention relates to an antibody selected from the group consisting of ACI-7069-631B2-Abl, ACI-7069-633B12-Abl, ACI-7069-634H10-Ab2, ACI-7069-636E5-Abl, ACI-7069-641H1-Ab2, ACI-7069-642A10-Abl, ACI-7069-642D12-Abl, ACI-7069-646B7-Abl, ACI-7071-712A6-Abl, ACI-7071-809D9-Ab2, and ACI-7071-809F12-Abl, as further described herein.

[0309] In some embodiments, an (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid encodes an antibody described herein.

[0310] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 18 encoding an anti-TPD-43 antibody.

[0311] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 19 encoding an anti-TPD-43 antibody.

[0312] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 28 encoding an anti-TPD-43 antibody.

[0313] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 29 encoding an anti-TPD-43 antibody.

[0314] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 38 encoding an anti-TPD-43 antibody.

[0315] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 39 encoding an anti-TPD-43 antibody.

[0316] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 48 encoding an anti-TPD-43 antibody.

[0317] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 49 encoding an anti-TPD-43 antibody.

[0318] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 68 encoding an anti-TPD-43 antibody.

[0319] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 69 encoding an anti-TPD-43 antibody.

[0320] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 78 encoding an anti-TPD-43 antibody.

[0321] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 79 encoding an anti-TPD-43 antibody.

[0322] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 88 encoding an anti-TPD-43 antibody.

[0323] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 89 encoding an anti-TPD-43 antibody.

[0324] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 108 encoding an anti-TPD-43 antibody.

[0325] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 109 encoding an anti-TPD-43 antibody.

[0326] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 128 encoding an anti-TPD-43 antibody.

[0327] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 129 encoding an anti-TPD-43 antibody.

[0328] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 148 encoding an anti-TPD-43 antibody.

[0329] In some embodiments, an (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 149 encoding an anti-TPD-43 antibody.

[0330] In some embodiments, an (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 158 encoding an anti-TPD-43 antibody.

[0331] In some embodiments, a (isolated) nucleic acid is provided, wherein the (isolated) nucleic acid comprises SEQ ID NO: 159 encoding an anti-TPD-43 antibody.

[0332] XII. Compositions and Methods

[0333] In some embodiments, an immunoconjugate is provided, wherein the immunoconjugate comprises an (isolated) antibody as described herein and a therapeutic agent. In some embodiments, a labeled antibody is provided, comprising an antibody as described herein and a detectable label.

[0334] In some embodiments, a pharmaceutical composition is provided, comprising an (isolated) antibody described herein and a pharmaceutically acceptable carrier.

[0335] In some embodiments, the TDP-43-specific binding molecules of the invention are linked to a detectable label.

[0336] In some embodiments, the TDP-43-specific binding molecule is part of an immunoconjugate in which the TDP-43-specific binding molecule is covalently linked to another suitable therapeutic agent.

[0337] In some embodiments, the TDP-43 specific binding molecule or immunoconjugate comprising the same is present as a composition comprising the TDP-43 specific binding molecule and a TDP-43 agonist and a cognate molecule, or alternatively an antagonist thereof.

[0338] In some embodiments, the TDP-43 specific binding molecule is part of a pharmaceutical composition comprising the TDP-43 specific binding molecule, or an immunoconjugate in which the TDP-43 specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising the TDP-43 specific binding molecule and a TDP-43 agonist and cognate molecule, or alternatively, an antagonist thereof, in combination with a pharmaceutically acceptable carrier.

[0339] In some embodiments, the TDP-43 specific binding molecule is part of a detection and / or diagnostic kit comprising the TDP-43 specific binding molecule, or an immunoconjugate in which the TDP-43 specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising the TDP-43 specific binding molecule and a TDP-43 agonist and a cognate molecule, or alternatively, an antagonist thereof.

[0340] Also provided are kits comprising the binding molecules of the invention. In particular, such kits can be used to perform the diagnostic methods of the invention (including classification, monitoring, and treatment selection methods). Thus, kits for diagnosing diseases, disorders, and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathies, or for use in the methods of the invention, are provided, comprising the TDP-43-specific binding molecules of the invention. Such kits can contain all necessary components for performing the methods provided herein. Typically, each component is stored separately in a single, integrated package. Suitable additional components for inclusion in the kit are, for example, buffers, detectable dyes, laboratory equipment, reaction vessels, instructions, and the like. The instructions for use can be customized for the specific method in which the kit is to be used. Also provided are suitably labeled TDP-43 binding molecules of the invention, which can be included in such kits.

[0341] In some embodiments, the TDP-43-specific binding molecules are used in immunodiagnostic methods for preventing, diagnosing, or treating TDP-43 proteinopathies.

[0342] In some embodiments, the TDP-43 specific binding molecule is part of an immunotherapeutic approach for preventing or treating a TDP-43 proteinopathy, wherein an effective amount of the TDP-43 specific binding molecule, or an immunoconjugate in which the TDP-43 specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising a TDP-43 specific binding molecule and a TDP-43 agonist and a cognate molecule, or alternatively an antagonist thereof, is administered to a subject in need thereof.

[0343] In some embodiments, a TDP-43-specific binding molecule, or an immunoconjugate in which a TDP-43-specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising a TDP-43-specific binding molecule and a TDP-43 agonist and a cognate molecule, or alternatively an antagonist thereof, is administered to a subject in need thereof for the diagnosis, prevention, alleviation or treatment of diseases, disorders and / or abnormalities associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathies, including but not limited to frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE), limbic-dominant age-related TDP-43 encephalopathy (LATE).

[0344] In some embodiments, a TDP-43-specific binding molecule, or an immunoconjugate wherein the TDP-43-specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising a TDP-43-specific binding molecule and a TDP-43 agonist and a cognate molecule, or alternatively an antagonist thereof, is administered to a subject in need thereof for diagnosis or monitoring of a disease, disorder and / or abnormality associated with TDP-43, particularly associated with TDP-43 aggregates, or a TDP-43 proteinopathy selected from the group consisting of: The following are frontotemporal dementias (FTDs, such as sporadic or familial, with or without motor neuron disease (MND), progranulin (GRN) mutations, C9orf72 mutations, TARDBP mutations, valosin-containing protein (VCP) mutations, chromosome 9p-related, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral changes, Heteromorphic FTD (bvFTD), nonfluent variant primary progressive aphasia (nfvPPA), amyotrophic lateral sclerosis (ALS, such as sporadic ALS, TARDBP mutations, angiopoietin (ANG) mutations), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy, Perry syndrome, Alzheimer's disease (AD, including sporadic and familial forms of AD), Down syndrome, familial British dementia, polymyalgia Glutamine disorders (Huntington disease and spinocerebellar ataxia type 3 (SCA3; also called Mahy-Joh disease)), hippocampal sclerosis dementia and myopathies (sporadic inclusion body myositis; inclusion body myopathy with mutations in the valosin-containing protein (VCP; as well as Paget's disease of bone and frontotemporal dementia); oculopharyngeal muscular dystrophy with rimmed vacuoles; myofibrillar myopathy with mutations in the myotilin (MYOT) gene or mutations in the gene encoding desmin (DES)), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD).

[0345] In other embodiments, the present invention relates to any method for detecting, diagnosing or monitoring a disease, disorder and / or abnormality associated with TDP-43, in particular associated with TDP-43 aggregates, or a TDP-43 proteinopathy selected from the group consisting of frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE) and limbic-dominant age-related TDP-43 encephalopathy (LATE).

[0346] Preferably, the disease, disorder and / or abnormality associated with TDP-43, in particular associated with TDP-43 aggregates, or TDP-43 proteinopathy is selected from amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD) and frontotemporal dementia (FTD). More preferably, the disease, disorder and / or abnormality associated with TDP-43, in particular associated with TDP-43 aggregates, or TDP-43 proteinopathy is amyotrophic lateral sclerosis (ALS). More preferably, the disease, disorder and / or abnormality associated with TDP-43, in particular associated with TDP-43 aggregates, or TDP-43 proteinopathy is Alzheimer's disease (AD). More preferably, the disease, disorder and / or abnormality associated with TDP-43, in particular associated with TDP-43 aggregates, or TDP-43 proteinopathy is frontotemporal dementia (FTD).

[0347] In some embodiments, the TDP-43-specific binding molecules are used in methods for diagnosing presymptomatic disease or for monitoring disease progression and treatment efficacy, or for predicting responsiveness, or for selecting subjects likely to respond to treatment with a TDP-43-specific binding molecule. The methods are preferably performed using samples of human blood or urine. Most preferably, the methods involve ELISA-based assays or surface-adapted assays.

[0348] In some embodiments, the TDP-43-specific binding molecules are used in methods wherein a TDP-43-specific binding molecule of the invention is contacted with a sample (e.g., blood, cerebrospinal fluid, or brain tissue) to detect, diagnose, or monitor frontotemporal degeneration (FTD) or amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), chronic traumatic encephalopathy, Perry syndrome, limbic-dominant age-related TDP-43 encephalopathy (LATE), and / or Parkinson's disease (PD).

[0349] In some embodiments, the TDP-43-specific binding molecules are used in methods wherein a TDP-43-specific binding molecule of the invention is contacted with a sample (e.g., blood, cerebrospinal fluid, or brain tissue) to detect, diagnose, or select from the following diseases: frontotemporal dementia (FTD, e.g., sporadic or familial, with or without motor neuron disease (MND), with progranulin (GRN) mutations, with C9orf72 mutations, with TARDBP mutations, with valosin-containing protein (VCP) mutations, associated with chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), nonfluent variant primary progressive aphasia (nfvPPA), etc.), amyotrophic lateral sclerosis (ALS), S, such as sporadic ALS with TARDBP mutations, angiogenic protein (ANG) mutations), Alexander disease (AxD), limbic-predominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy, Perry syndrome, Alzheimer's disease (AD, including sporadic and familial forms of AD), Down syndrome, familial British dementia, polyglutamine diseases (Huntington disease and spinocerebellar ataxia type 3 (SCA3; also known as Mahy-Joh disease)), hippocampal sclerosis dementia and myopathies (sporadic inclusion body myositis; inclusion body myopathy with mutations in the valosin-containing protein (VCP; as well as Paget's disease of bone and frontotemporal dementia; oculopharyngeal muscular dystrophy with rimmed vacuoles; myofibrillar myopathy with mutations in the myotilin (MYOT) gene or mutations in the gene encoding desmin (DES))), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD).

[0350] In some embodiments, a TDP-43-specific binding molecule, or an immunoconjugate in which a TDP-43-specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising a TDP-43-specific binding molecule and a TDP-43 agonist and a homologous molecule, or alternatively an antagonist thereof, is administered to a subject in need thereof for the prevention, alleviation or treatment of diseases, disorders and / or abnormalities associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathy, or frontotemporal degeneration (FTD) or amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD, including sporadic and familial forms of AD), chronic traumatic encephalopathy, Perry syndrome and limbic-dominant age-related TDP-43 encephalopathy (LATE) and / or Parkinson's disease (PD).

[0351] In some embodiments, a TDP-43-specific binding molecule, or an immunoconjugate wherein the TDP-43-specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising a TDP-43-specific binding molecule and a TDP-43 agonist and a homologous molecule, or alternatively an antagonist thereof, is administered to a subject in need thereof for the treatment of a disease selected from the group consisting of frontotemporal dementia (FTD, e.g., sporadic or familial, with or without motor neuron disease (MND), with progranulin (GRN) mutations, with C9orf72 mutations, with TARDBP mutations, with valosin-containing protein (VCP) mutations, associated with chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), nonfluent variant primary progressive aphasia amyotrophic lateral sclerosis (ALS, such as sporadic ALS, TARDBP mutations, and angiogenic protein (ANG) mutations), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy, Perry syndrome, Alzheimer's disease (AD, including sporadic and familial forms of AD), Down syndrome, familial British dementia, polyglutamine diseases (Huntington disease and spinal cord microglia), The invention also includes ataxia cerebral ataxia type 3 (SCA3; also known as Mahy-Joh disease), hippocampal sclerosis dementia and myopathies (sporadic inclusion body myositis; inclusion body myopathy with mutations in the valosin-containing protein (VCP; as well as Paget's disease of bone and frontotemporal dementia); oculopharyngeal muscular dystrophy with rimmed vacuoles; myofibrillar myopathy with mutations in the myotilin (MYOT) gene or mutations in the gene encoding desmin (DES)), traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD). Preferably, the treatment of the disease helps maintain or improve mental cognition and / or reduces the level of TDP-43 aggregates in the brain.

[0352] In some embodiments, a TDP-43-specific binding molecule, or an immunoconjugate in which a TDP-43-specific binding molecule is covalently linked to another suitable therapeutic agent, or a composition comprising a TDP-43-specific binding molecule and a TDP-43 agonist and a cognate molecule, or alternatively an antagonist thereof, is administered to a subject in need thereof for the manufacture of a medicament for treating diseases, disorders and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or a TDP-43 proteinopathy, or frontotemporal degeneration (FTD) or amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD, including sporadic and familial forms of AD), chronic traumatic encephalopathy, Perry syndrome and limbic-dominant age-related TDP-43 encephalopathy (LATE), and / or Parkinson's disease (PD).

[0353] Pharmaceutical formulations of anti-TDP-43 antibodies (preferred types of TDP-43-specific binding molecules) or immunoconjugates as described herein are prepared by mixing such antibodies or immunoconjugates having the desired degree of purity with one or more optional pharmaceutically acceptable carriers (Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)) in the form of lyophilized formulations or aqueous solutions. Pharmaceutically acceptable carriers are generally nontoxic to recipients at the dosages and concentrations employed and include, but are not limited to, buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (e.g., octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride; benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol and m-cresol); low molecular weight (less than about 10 residues) ) polypeptides; proteins such as serum albumin, gelatin or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine or lysine; monosaccharides, disaccharides and other carbohydrates including glucose, mannose or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn protein complexes); and / or nonionic surfactants such as polyethylene glycol (PEG). Exemplary pharmaceutically acceptable carriers herein also include interstitial drug dispersants such as soluble neutral-active hyaluronidase glycoprotein (sHASEGP), such as human soluble PH-20 hyaluronidase glycoprotein, such as rHuPH20 ( Baxter International, Inc.). Certain exemplary sHASEGPs and methods of use, including rHuPH20, are described in U.S. Patent Publication Nos. 2005 / 0260186 and 2006 / 0104968. In one aspect, sHASEGP is combined with one or more additional glycosaminoglycanases, such as chondroitinase.

[0354] Exemplary lyophilized antibody or immunoconjugate formulations are described in U.S. Patent No. 6,267,958. Aqueous antibody or immunoconjugate formulations include those described in U.S. Patent No. 6,171,586 and WO2006 / 044908, the latter formulations comprising a histidine-acetate buffer.

[0355] The formulations herein may also contain more than one active ingredient, as necessary for the particular indication being treated, preferably ones with complementary activities that do not adversely affect each other.

[0356] The active ingredient can be encapsulated in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, such as hydroxymethylcellulose or gelatin microcapsules and poly(methyl methacrylate) microcapsules, respectively; in colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules); or in macroemulsions. Such techniques are disclosed in Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980).

[0357] Sustained-release formulations can be prepared. Some suitable examples of sustained-release formulations include semipermeable matrices of solid hydrophobic polymers comprising the antibody or immunoconjugate, which are in the form of shaped articles, e.g., films or microcapsules. Formulations for in vivo administration are typically sterile. Sterility can be easily achieved, for example, by filtration through a sterile filtration membrane.

[0358] Any of the antigen binding molecules, anti-TDP-43 antibodies, or immunoconjugates provided herein can be used in a method, such as a method of treatment.

[0359] In another aspect, an anti-TDP-43 antibody (preferred type of TDP-43 specific binding molecule) or immunoconjugate for use as a medicament is provided. In a further aspect, an anti-misfolded TDP-43 antibody (preferred type of TDP-43 specific binding molecule) or immunoconjugate for use in a therapeutic method is provided. In certain embodiments, an anti-TDP-43 antibody (preferred type of TDP-43 specific binding molecule) or immunoconjugate for use in preventing, diagnosing and / or treating TDP-43 proteinopathies is provided. In a preferred embodiment of the present invention, an anti-TDP-43 antibody (a preferred type of TDP-43 specific binding molecule) or immunoconjugate is provided for use in preventing, diagnosing and / or treating diseases, disorders and / or abnormalities associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathy, including but not limited to frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE) and / or limbic-dominant age-related TDP-43 encephalopathy (LATE).

[0360] In another aspect, the present invention provides for the use of an anti-TDP-43 antibody (a preferred type of TDP-43-specific binding molecule) or immunoconjugate in the manufacture or preparation of a medicament. In one such embodiment, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, e.g., as described below.

[0361] According to any of the above embodiments, a "subject" or "individual" may be an animal, a mammal, preferably a human.

[0362] In another aspect, the present invention provides a pharmaceutical formulation comprising any of the anti-TDP-43 antibodies (preferred types of TDP-43 specific binding molecules) or immunoconjugates provided herein, for example for use in any of the above treatment methods. In one embodiment, the pharmaceutical formulation comprises any of the anti-TDP-43 antibodies (preferred types of TDP-43 specific binding molecules) or immunoconjugates provided herein, and a pharmaceutically acceptable carrier. In another embodiment, the pharmaceutical formulation comprises any of the anti-TDP-43 antibodies (preferred types of TDP-43 specific binding molecules) or immunoconjugates provided herein, and at least one additional therapeutic agent, e.g., as described below.

[0363] The antibodies or immunoconjugates of the invention can be used alone or in combination with other agents in therapy. For example, the antibodies (preferred types of TDP-43-specific binding molecules) or immunoconjugates of the invention can be co-administered with at least one additional therapeutic agent.

[0364] Such combination therapies described above encompass both combined administration (wherein two or more therapeutic agents are contained in the same or separate formulations) and separate administration. In the case of separate administration, administration of the antibodies (preferred types of TDP-43-specific binding molecules) or immunoconjugates of the present invention can occur before, simultaneously with, and / or after administration of the additional therapeutic agent and / or adjuvant. The antibodies (preferred types of TDP-43-specific binding molecules) or immunoconjugates of the present invention can also be used in combination with radiation therapy.

[0365] The antibodies (preferred types of TDP-43-specific binding molecules) or immunoconjugates of the present invention (and any additional therapeutic agents) can be administered by any suitable means, including parenteral, intrapulmonary and intranasal, and, if desired, local treatment, intralesional, intrauterine or intravesical administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal or subcutaneous administration. Administration can be by any suitable route, for example, by injection, such as intravenous or subcutaneous injection, depending in part on whether the administration is short-term or long-term. Various dosing regimens are contemplated herein, including but not limited to single administration or multiple administrations at different time points, bolus administration, and pulse infusion.

[0366] The antibodies (preferred types of TDP-43-specific binding molecules) or immunoconjugates of the present invention can be formulated, dosed, and administered in a manner consistent with good medical practice. Factors to consider in this context include: the specific disease, disorder, and / or abnormality associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathy being treated; the specific mammal being treated; the clinical condition of the individual subject; the cause of the disease, disorder, and / or abnormality associated with TDP-43, particularly associated with TDP-43 aggregates, or TDP-43 proteinopathy; the delivery site of the agent; the method of administration; the administration regimen; and other factors known to medical practitioners. The antibody or immunoconjugate need not be, but is optionally, formulated with one or more agents currently used to prevent or treat the disease, disorder, and / or abnormality associated with TDP-43 aggregates, or TDP-43 proteinopathy in question. The effective amount of such other agents depends on the amount of antibody or immunoconjugate present in the formulation; the disease, disorder and / or abnormality associated with TDP-43, particularly TDP-43 aggregates, or the type of TDP-43 proteinopathy; or treatment, as well as other factors described above. These are generally used in the same dosages and administration routes as described herein, or at about 1% to 99% of the dosages described herein, or in any dosage and by any route determined empirically / clinically appropriate.

[0367] For the prevention or treatment of disease, the appropriate dosage of the antibody (preferred type of TDP-43 specific binding molecule) or immunoconjugate of the present invention (when used alone or in combination with one or more other additional therapeutic agents) will depend on the type of disease being treated, the type of antibody or immunoconjugate, the severity and cause of the disease, whether the antibody or immunoconjugate is being administered for preventive or therapeutic purposes, previous treatments, the subject's clinical history and response to the antibody or immunoconjugate, and the judgment of the attending physician. The antibody (preferred type of TDP-43 specific binding molecule) or immunoconjugate is appropriately administered to the subject at one time or over a series of treatments. Depending on the type and severity of the disease, about 1 μg / kg to 15 mg / kg (e.g., 0.1 mg / kg to 10 mg / kg) of the antibody (preferred type of TDP-43 specific binding molecule) or immunoconjugate can be an initial candidate dose for administration to the subject, whether, for example, by one or more separate administrations or by continuous infusion. Depending on the above factors, a typical daily dose can be about 1 μg / kg to 100 mg / kg or more. For repeated administration over several days or longer, depending on the disease, continued treatment will generally be used until desired disease symptoms are suppressed. An exemplary dosage of an antibody or immunoconjugate is about 0.05 mg / kg to about 10 mg / kg. Therefore, one or more dosages (or any combination thereof) of about 0.5 mg / kg, 2.0 mg / kg, 4.0 mg / kg or 10 mg / kg can be administered to the subject. Such dosage can be administered intermittently, for example, weekly or every three weeks (for example, so that the subject receives about 2 to about 20, or for example, about 6 dosages of the antibody). A higher initial loading dose can be administered, followed by one or more lower dosages. However, other dosage regimens can be used. The progress of the treatment is easily monitored by conventional techniques and assays.

[0368] It will be understood that any of the above formulations or treatment methods can be performed using both the immunoconjugates of the invention and anti-TDP-43 antibodies (a preferred type of TDP-43-specific binding molecule).

[0369] In another aspect of the present invention, an article of manufacture comprising the materials described above that can be used to treat, prevent, and / or diagnose diseases, disorders, or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathies is provided. The article of manufacture comprises a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, IV solution bags, and the like. The container can be formed from a variety of materials, such as glass or plastic. The container holds a composition that is effective, alone or in combination with another composition, for treating, preventing, and / or diagnosing diseases, disorders, and / or abnormalities associated with TDP-43, particularly TDP-43 aggregates, or TDP-43 proteinopathies, and can have a sterile access port (e.g., the container can be an intravenous solution bag or a vial with a stopper pierceable by a hypodermic needle). At least one active agent in the composition is an antibody or immunoconjugate of the present invention. The label or package insert indicates that the composition is used to treat the selected condition. In addition, the article of manufacture may comprise: (a) a first container comprising a composition, wherein the composition comprises an antibody (a preferred type of TDP-43-specific binding molecule) or immunoconjugate of the present invention; and (b) a second container comprising a composition, wherein the composition comprises an additional therapeutic agent. The article of manufacture in this embodiment of the invention may further comprise a package insert indicating that the composition can be used to treat a specific condition. Alternatively or in addition, the article of manufacture may further comprise a second (or third) container comprising a pharmaceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer's solution, or dextrose solution. It may also contain other materials desirable from a commercial and user perspective, including other buffers, diluents, filters, needles, and syringes.

[0370] In another embodiment, the present invention relates to a method for maintaining or improving cognitive memory ability, motor and language function, or preventing and / or slowing down the decline of cognitive memory ability, motor and language function in a subject, comprising administering a binding molecule of the present invention, an immunoconjugate of the present invention, a composition of the present invention, or a pharmaceutical composition of the present invention.

[0371] In another embodiment, the invention relates to a method of reducing TDP-43 levels comprising administering a binding molecule of the invention, an immunoconjugate of the invention, a composition of the invention, or a pharmaceutical composition of the invention.

[0372] The methods of the present invention may comprise administering at least one additional treatment, preferably wherein the additional treatment is selected from, but not limited to, neurological drugs, anti-Aβ antibodies, anti-Tau antibodies, Tau aggregation inhibitors, beta amyloid aggregation inhibitors, anti-BACE1 antibodies, and BACE1 inhibitors.

[0373] The present invention also relates to a method for detecting TDP-43, comprising contacting a sample with a binding molecule of the present invention, preferably, wherein the sample is a brain sample, a cerebrospinal fluid sample, a urine sample or a blood sample. BRIEF DESCRIPTION OF THE DRAWINGS

[0374] Figure 1 : Detection of TDP-43 in tissue sections from subjects with frontotemporal dementia (FTD) with type A pathology. Immunohistochemistry was performed on 10 μm thick frozen sections of the frontal cortex from FTD subjects with type A pathology using fluorescently labeled secondary antibodies for detection. The following antibodies were used as controls: rabbit polyclonal pan-TDP-43 antibody (Proteintech, 10782-2-AP) for detection of pathological inclusions and physiological nuclear TDP-43; rabbit monoclonal phospho-TDP-43p409 / 410 antibody (Cosmobio, TIP-PTD-P02) for detection of pathological aggregated and phosphorylated TDP-43. Arrows indicate TDP-43 aggregates; thick arrows indicate physiological TDP-43 in the nucleus (nuclei were visualized by DAPI staining). The hybridoma name or commercial antibody source is shown in the upper left corner of each image.

[0375] Figure 2 : Detection of TDP-43 in detergent-soluble and insoluble fractions obtained from postmortem brain tissue (frontal cortex) of FTD type A. Immunoblotting with commercial antibodies that bind to the N-terminal region (A, B) or the C-terminal region (C) shows the presence of TDP-43 in the sarkosyl-soluble (lane 1) and insoluble (lane 2) fractions. Immunoblotting of mAbs against TDP-43 generated in this study whose epitope is in the N-terminal region of TDP-43 (D to I). Immunoblotting of mAbs against TDP-43 that bind to the C-terminal region of TDP-43 (J to N). All mAbs against TDP-43 specifically recognize full-length TDP-43. In addition, some mAbs (K, M, N) recognize pathological features of diseased states, such as the C-terminal fragment in the insoluble fraction.

[0376] Figure 3: The density of pTDP-43 immunoreactive objects measured in two brain regions: striatum (A) and cerebral cortex (B) of mice treated with vehicle (n=30, gray bars) and ACI-7069-633B12-Ab1 (IgG2a variant) (n=25, dotted gray bars) is shown. (C) Insoluble fractions obtained from the cortex of the left cerebral hemisphere were quantified for total TDP-43 in the vehicle (n=30) and ACI-7069-633B12-Ab1 (IgG2a variant) (n=25) treated groups (*p<0.05, **p<0.01, ****p<0.0001).

[0377] Figure 4 TDP-43 aggregation induced by TEV cleavage in the presence of ACI-7069-633B12-Abl (IgG2a variant) or isotype control was measured by turbidity at 600 nm after 30 h. The endpoint after 30 h was normalized to the isotype control (grey bars), and aggregated TDP-43 (%) was calculated for ACI-7069-633B12-Abl (dotted grey bars). Mean ± SD of three independent experiments is shown, and statistical differences between the isotype control and ACI-7069-633B12-Abl (IgG2a variant) were analyzed by Welch's t-test (***p < 0.001).

[0378] Figure 5 : (A) shows the area of ​​Iba1 positive immunoreactivity measured in the cerebral cortex of mice treated with vehicle (n=16, gray bars) and ACI-7069-633B12-Ab1 (IgG2a variant) (n=16, dotted gray bars). Error bars represent the standard error of the mean (SEM). (B to C) show the average microglial size measured in the cerebral cortex of mice treated with vehicle (n=16, gray bars) and ACI-7069-633B12-Ab1 (IgG2a variant) (n=16, dotted gray bars). Microglia were classified into three categories based on their morphology: (B) large hypertrophic, (C) small ramifying, and (D) ramified resting. Statistical differences between vehicle control and ACI-7069-633B12-Ab1 (IgG2a variant) were analyzed by t-test (*p<0.05).

[0379] Figure 6: TDP-43 levels in the CSF of multiple FTLD-TDP patients and healthy controls were quantified using AlphaLISA assays using ACI-7069-633B12-Ab1 (IgG2a variant) and ACI-7071-809F12-Ab1 (IgG2a variant). Raw AlphaLISA counts (y-axis) of total TDP-43 were obtained for multiple CSF samples (x-axis). Statistical analysis of raw counts was performed using a linear mixed model with data from three independent experiments using group, experiment, sex, and age as fixed factors and individual as a random factor (**p < 0.01).

[0380] Figure 7 : Immunodepletion of TDP-43 and pTDP-43 from detergent-insoluble fractions obtained from postmortem brain tissue of FTD type A using antibodies ACI-7069-633B12-Ab1 (IgG2a variant) (1), ACI-7069-642D12-Ab1 (IgG2a variant) (2), and mouse IgG2a control (3). Immunodepleted fractions 1 to 3 were analyzed by Western blotting using TDP-43 or pTDP-43-specific detection antibodies. IN refers to input material (before immunodepletion).

[0381] Example

[0382] Example 1: Preparation of TDP-43 vaccine composition

[0383] Liposome-based vaccines were prepared according to the protocol published in WO 2012 / 055933. A vaccine comprising full-length TDP-43 (FL TDP-43) protein as antigen (Table 2, SEQ ID NO: 1) was used for antibody generation.

[0384] Table 2: Description of TDP-43 protein and peptide antigens

[0385]

[0386] Example 2: Generation of anti-TDP-43 antibodies

[0387] A. Immunization of mice

[0388] Female C57BL / 6O1aHsd (C57BL / 6) and BALB / c O1aHsd (BALB / c) wild-type mice (Harlan, USA) were administered at 9 weeks of age. Vaccination began at 10 weeks of age in the presence of monophosphoryl hexaacyl lipid A, 3-deacylated (synthetic) (3D-(6-acyl) ), mice were vaccinated with full-length TDP-43 protein present on the surface of liposomes.

[0389] Mice were vaccinated by subcutaneous injection (sc) on days 0, 4, 8, 21, 35, and 60. Heparinized plasma was prepared from the mice 7 days before immunization (preimmune plasma) and on days 14, 28, 42, 81, and 121 after the first immunization. In addition, mice used for myeloma fusions were vaccinated by three daily booster injections of TDP-43 protein by ip injection without adjuvant.

[0390] Vaccine responses were measured in mouse plasma. Binding of plasma-derived antibodies from immunized mice to immobilized recombinant full-length (FL) TDP-43 indicated high titers of antibodies against TDP-43.

[0391] B. Hybridoma Generation and Subclone Selection

[0392] Mice were euthanized and spleen cells from four individual mice were used for fusion with myeloma cells. Antibody screening from successfully fused hybridoma cell lines was performed as follows. Diluted (1:32) cell culture supernatants were analyzed using a Luminex bead-based multiplex assay (Luminex, The Netherlands). Luminex beads were conjugated to FLTDP-43 and IgG was captured using anti-mouse IgG-Fc antibodies specific for IgG1, IgG2a, IgG2b, IgG2c, and IgG3 subclasses (Jackson Immunoresearch, USA). Binding to beads conjugated to FL TDP-43 identified 386 hits from mice immunized with the FL TDP-43 liposomal vaccine.

[0393] Viable hybridomas were cultured in serum-containing selective medium. Clones that preferentially bound to TDP-43 inclusion bodies in human FTD brain and to the C-terminus of TDP-43 were selected for further subcloning. After limiting dilution, clonal hybridomas were cultured in immunoglobulin-reduced medium, and stable colonies were selected for antibody screening and selection. The antibodies shown in Table 3 were identified from this screening.

[0394] Example 3: Determination of binding potency (EC50)

[0395] As described above, Luminex assays were performed with serial dilutions of the antibodies to determine the half-maximal effective concentration (EC50) for antibody binding to FL TDP-43. All EC50 values ​​are summarized in Table 3. In summary, all tested antibodies bound to full-length TDP-43 with high affinity.

[0396] Table 3: EC50 values ​​determined by Luminex assay

[0397]

[0398]

[0399] Example 4: Antibodies that bind to human FL TDP-43

[0400] Antibody binding to human FL TDP-43 was determined using an indirect ELISA. ELISA plates were coated with 1 μg / ml human FL TDP-43 in carbonate buffer overnight at 4°C. The plates were washed with 0.05% Tween 20 / PBS and then blocked with 1% bovine serum albumin (BSA) in 0.05% Tween 20 / PBS for 1 hour at 37°C. Antibodies purified from hybridoma supernatants were then added in 3-fold serial dilutions (starting at 1 μg / ml) and incubated at 37°C for 2 hours, after which the plates were washed. AP-conjugated anti-mouse IgG secondary antibody (Jackson Immunoresearch Laboratories, United Kingdom) was added at a 1 / 1000 dilution in 0.05% Tween 20 / PBS for 1 hour at 37°C. After the final wash, the plates were incubated with pNPP (Sigma-Aldrich, Switzerland) phosphatase substrate solution and read at 405 nm using an ELISA plate reader (Tecan, Switzerland). All clones tested bound to full-length TDP-43 with varying EC50 values ​​ranging from 10 to 1567 ng / ml (Table 4).

[0401] Table 4: EC50 values ​​by ELISA

[0402]

[0403] Example 5: Epitope mapping by ELISA and peptide array

[0404] Antibodies purified from serum-free hybridoma supernatants were screened by indirect ELISA assay using a library of 40-66aa linear peptides or 15-mer peptides biotinylated at the N-terminus and covering the entire TDP-43 sequence with a 9aa offset and a 6aa overlap to determine the binding region. The peptide sequences are provided in Table 5.

[0405] 96-well plates were coated with 5 μg / ml non-biotinylated peptide in carbonate buffer at 4°C overnight. The plates were washed with 0.05% Tween 20 / PBS and then blocked with 1% bovine serum albumin (BSA) in 0.05% Tween 20 / PBS for 1 hour at 37°C. Antibodies purified from hybridoma supernatants were then added at 1 μg / ml and incubated at 37°C for 2 hours, after which the plates were washed. AP-conjugated anti-mouse IgG secondary antibody (Jackson Immunoresearch Laboratories, United Kingdom) was added at a 1 / 1000 dilution in 0.05% Tween 20 / PBS for 1 hour at 37°C. After the final wash, the plates were incubated with pNPP (Sigma-Aldrich, Switzerland) phosphatase substrate solution and read at 405 nm using an ELISA plate reader (Tecan, Switzerland).

[0406] For biotinylated peptides, a 96-well streptavidin-coated ELISA plate was incubated with 5 μg / mL of biotinylated 15-mer peptide. The plate was washed three times with 0.05% Tween 20 / PBS and then blocked with 1% bovine serum albumin (BSA) in 0.05% Tween 20 / PBS at 37°C for 1 hour. Antibodies purified from hybridoma supernatants were then added at 1 μg / ml and incubated at 37°C for 2 hours, after which the plate was washed. AP-conjugated anti-mouse IgG secondary antibody (Jackson ImmunoResearch Laboratories, United Kingdom) was added at a 1 / 1000 dilution in 0.05% Tween 20 / PBS at 37°C for 1 hour. After the final wash, the plate was incubated with pNPP (Sigma-Aldrich, Switzerland) AP substrate solution and read at 405 nm using an ELISA plate reader (Tecan). The identified binding regions are provided in Table 6. The test antibodies were found to bind to the following peptides: TP-21, TP-23, TP-35, TP-40, TP-48, TDP-6 corresponding to regions 181 to 195, 199 to 213, 307 to 321, 352 to 366, 389 to 411, and 140 to 200 of SEQ ID NO: 1, respectively.

[0407] More precise linear epitopes were mapped using a library of 15-mer peptides (Pepscan, Netherlands) synthesized directly on the solid support and covering the entire TDP-43 sequence according to SEQ ID NO: 1 with a 1 aa offset and a 14 aa overlap. The peptide arrays were blocked with horse serum and ovalbumin and incubated overnight at 4°C with purified antibody solutions at concentrations ranging from 0.75 to 5 μg / ml. After washing, the peptide arrays were incubated with a 1 / 1000 dilution of rabbit anti-mouse IgG (H+L) HRP conjugate (Southern Biotech, USA) for 1 hour at 25°C. After washing, the peroxidase substrate 2,2'-azido-di-3-ethylbenzothiazoline sulfonate (ABTS) and 20 μl / ml of 3% H2O2 were added. After one hour, the color development was quantified using a charge coupled device (CCD) camera and an image processing system. These binding regions were determined by epitope mapping, and the following epitopes were identified (provided in Table 6): amino acids 183 to 188, 203 to 213, 204 to 208, 204 to 211, 205 to 210, 316 to 323, 358 to 361, 400 to 405, 400 to 406, and 400 to 412 of SEQ ID NO: 1.

[0408] Table 5: Peptides used to determine binding regions by ELISA

[0409]

[0410] Table 6: Binding regions and epitopes of the tested antibodies

[0411] Hybridoma clone name Binding region, aa Epitope, aa 631B2A2 397-411 400-406 633B12C8 397-411 400-405 634H10H7 140-200 183-188 636E5B8 352-366 358-361 641H1E7 199-213 204-211 642A10B11 389-411 400-412 642D12B4 181-195 183-188 646B7F7 199-213 205-210 712A6B10 307-321 316-323 809D9C2 199-213 203-213 809F12D8 199-213 204-208

[0412] Example 6: Detection of TDP-43 in brain tissue from FTD / ALS subjects by immunohistochemistry

[0413] Target engagement was assessed in immunohistochemistry experiments on tissue from the brains of FTD subjects. Human FTD brain tissue was purchased from The Netherlands Brain Bank, Netherlands Institute for Neuroscience (Amsterdam) (Open Access: www.brainbank.nl) and Queen Square Brain Bank for Neurological Disorders (UCL). All materials were collected from donors from whom written informed consent for brain autopsy and use of materials and clinical information for research purposes was obtained. Immunohistochemistry was performed on 10 μm thick frozen sections using fluorescently labeled secondary antibodies for detection. The following antibodies were used as controls: rabbit polyclonal pan-TDP-43 antibody (Proteintech, 10782-2-AP) for detection of pathological inclusions and physiological nuclear TDP-43; rabbit monoclonal phosphorylated TDP-43p409 / 410 antibody (Cosmobio, TIP-PTD-P02) for detection of pathological aggregated and phosphorylated TDP-43; and secondary antibody without primary antibody (no 1° Ab) for detection of nonspecific background.

[0414] All antibodies of the present invention bind to nuclear TDP-43, non-aggregated TDP-43, and aggregated TDP-43. Some antibodies of the present invention preferentially bind to aggregated TDP-43 in the cytoplasm in type A pathology ( Figure 1 ). The detailed evaluation of the binding characteristics is summarized in Table 7.

[0415] Table 7: Detection of TDP-43 in brain tissue from FTD subjects

[0416] Antibody name IHC detection of aggregated TDP-43 IHC detection of non-aggregated TDP-43 in the nucleus ACI-7069-631B2-Ab1 +++ +++ ACI-7069-633B12-Ab1 +++ +++ ACI-7069-634H10-Ab2 ++ ++ ACI-7069-636E5-Ab1 + / - +++ ACI-7069-641H1-Ab2 +++ + ACI-7069-642A10-ab1 +++ + ACI-7069-642D12-Ab1 ++ + ACI-7069-646B7-Ab1 +++ +++ ACI-7071-712A6-Ab1 ++ + ACI-7071-809D9-Ab2 +++ +++ ACI-7071-809F12-Ab1 +++ +++

[0417] NA: Little data available; -: Little presence; + / -: Little clarity; +: Weak; ++: Moderate; +++: Abundant

[0418] Example 7: Detection of TDP-43 in Brain Tissue from FTD / ALS Subjects by Western Blot

[0419] A brain tissue region (frontal cortex) was homogenized at 4°C in a 1:4 (w / v) ratio in homogenization-solubilization buffer (HS buffer) using a precellys CK mixing homogenizer (Labgene, BER0092). The following sequence was used for homogenization: 3 cycles at 5000 rpm for 30 seconds (with a 15 second pause between each cycle). The homogenized sample was aliquoted and stored at -80°C in 1.5 ml low protein binding tubes (Axygen MCT-175-LC).

[0420] • HS buffer - 10 mM Tris.HCl pH 7.5, 150 mM NaCl, 0.1 mM EDTA, 1 mM DTT, Complete EDTA-free protease inhibitors (Roche, 32524300) and PhosSTOP phosphatase inhibitors (Roche, 4906837001).

[0421] The brain homogenate was thawed on ice and resuspended in HS buffer to a final concentration of 2% Sarkosyl, 1 unit / μL Benzonase, and 1 mM MgCl2. The sample was then incubated on a thermomixer at 37°C with constant shaking at 600 rpm for 45 minutes. The supernatant was collected in a new tube. The pellet was resuspended in 1000 μl of myelin flotation buffer and centrifuged at 20,000 g for 60 minutes at 4°C on a benchtop centrifuge. The supernatant was carefully removed to remove all floating lipids. If all lipids could not be removed in a single step, the step was repeated. The pellet was then washed with PBS and centrifuged at 4°C on a benchtop centrifuge for 30 minutes. The final pellet was resuspended in 200 μl PBS and stored at -80°C. The samples were analyzed by immunoblotting under denaturing conditions.

[0422] HS buffer containing Sarkosyl, Benzonase and MgCl2 - 10 mM Tris.HCl pH 7.5, 150 mM NaCl, 0.1 mM EDTA, 1 mM DTT, 4% Sarkosyl, 1 unit / μL Benzonase (Novagen 70746-4), 4 mM MgCl2, complete EDTA-free protease inhibitors (Roche) and PhosSTOP phosphatase inhibitors (Roche).

[0423] Myelin flotation buffer - HS buffer containing 1% Triton X-100 and 30% sucrose

[0424] Western blotting was performed on Bolt 12% Bis-Tris Plus gel 1.0 mm (Thermofisher) using MES SDS running buffer (Thermofisher). Once diluted in PBS, samples (30 μl / sample) were loaded onto the gel, and loading buffer (1×, Licor, 928-40004) contained 100 mM DTT. Protein was decomposed at a constant voltage of 100 V for 1 hour. After electrophoresis, proteins were transferred to a nitrocellulose membrane (Thermofisher, IB23001) at 20 volts using iBLOT (Thermofisher, IB21001) for 7 minutes. After protein transfer, the membrane was blocked for 1 hour in Licor blocking buffer (Odyssey blocking buffer 927-40000) diluted in PBS at 1:3. The membranes were incubated overnight with the following primary antibodies: total TDP-43 (Proteintech, 60019-2-Ig or 10782-2-AP), pTDP-43 (Cosmobio, TIP-PTD-M01). For the primary antibodies, blocking buffer was diluted 1:1 in PBS-T (PBS containing 0.4% Tween 20). After washing four times with PBS-T (PBS containing 0.1% Tween 20), the membranes were incubated with secondary antibodies conjugated to LICOR dyes. Secondary antibodies - donkey anti-mouse (Cat. No. 926-68072) or goat anti-rabbit (Cat. No. 926-32211) - were used at a dilution of 1:10,000 in Licor blocking buffer diluted 1:1 with PBS-T (PBS containing 0.4% Tween 20) for 1 hour at room temperature. The membrane was washed again four times with PBS-T (PBS containing 0.1% Tween 20) and scanned using the LICOR system. Figure 2 All mAbs were shown to specifically recognize full-length TDP-43. In addition, some mAbs (K, M, N) recognized pathological features of the disease state, such as C-terminal fragments and high molecular weight aggregates in the insoluble fraction. Example 8A: Affinity measurements using SPR

[0425] Binding affinity to soluble or aggregated FL TDP-43 was assessed by determining the dissociation constant (KD) using surface plasmon resonance (SPR; Biacore T200, GE Healthcare Life Sciences). Recombinant human soluble or aggregated FL TDP-43 was immobilized on a CM5 Series S sensor chip (GE Healthcare Life Sciences) by amine coupling. Soluble TDP-43 was immobilized at a concentration of 5 μg / ml in 10 mM sodium acetate (pH 4.5) at a flow rate of 5 ul / min for 420 seconds, resulting in an immobilization level of 150 RU. Aggregated TDP-43 was immobilized at a concentration of 50 μg / ml in 10 mM sodium acetate (pH 4.5) at a flow rate of 5 μl / min for 840 seconds, resulting in an immobilization level of 110 RU. Biotinylated TP-73 peptide (amino acids 181 to 190 of SEQ ID NO: 1) was immobilized in PBS-P + The antibody was immobilized on a Series S Sensor Chip SA (GE Healthcare Life Sciences) at a concentration of 5 μg / ml at a flow rate of 5 μl / min for 30 seconds, resulting in an immobilization level of 400 RU. To evaluate the KD value, the purified antibody and control antibody (2E2-D3) were mixed in PBS-P +The antibodies were injected at a flow rate of 50 μl / min starting at 333 nM and diluted down to 0.15 nM. The antibodies were injected at a flow rate of 50 μl / min, with a contact time of 90 seconds and a dissociation phase of 700 seconds, followed by three regenerations in 10 mM glycine-HCl pH 1.7. For the optimized SPR protocol, the antibodies were diluted 3-fold starting at 300 nM and diluted down to 1.2 nM, and injected at 30 ul / min for 300 seconds, followed by a dissociation phase of 600 seconds. The surface was regenerated by a single injection of 10 mM glycine-HCl pH 1.7. The results obtained from the binding kinetics were double-referenced using a blank flow cell and buffer cycles and evaluated using an overall 1:1 fit model with RI. The affinities of the 11 antibodies and two Fab fragments are shown in Table 8. The antibodies of the present invention bound to aggregated TDP-43 with a KD ranging from 0.62 nM to 4.64 nM. In addition, some antibodies showed preferential binding to aggregated TDP-43 compared to soluble TDP-43. Two Fab fragments bound to soluble TDP-43 with KDs ranging from 2.8 nM to 21.8 nM and showed similar KDs for aggregated TDP-43. Two antibodies (marked with *) were reanalyzed using an optimized SPR protocol (with longer association and dissociation phases) that allows for more accurate KD determination, especially for antibodies with slower off-rates. Both antibodies bound to soluble TDP-43 with KDs ranging from 0.22 nM to 3.9 nM and to aggregated TDP-43 with KDs ranging from 0.18 nM to 0.69 nM. Antibody ACI-7069-642D12-Ab1 bound to the TP-73 peptide with a KD of 3.6 nM.

[0426] Table 8: Binding characterization by SPR

[0427]

[0428]

[0429] NA, not applicable, because there are fewer than three curves available for fitting.

[0430] *Binding of recombinantly produced IgG2a isotype antibodies was characterized using an optimized SPR protocol.

[0431] Example 8B: Affinity Measurement Using SPR

[0432] The binding affinity to soluble FL TDP-43 was evaluated by determining the dissociation constant (KD) using surface plasmon resonance (SPR; Biacore T200, GE Healthcare Life Sciences). Goat anti-mouse capture antibodies were immobilized on CM5 Series S sensor chips (GE Healthcare Life Sciences) by amine coupling. The antibodies were allowed to bind to soluble FL TDP-43 in PBS-PBS. + (GE Healthcare Life Sciences) at a concentration of 2 to 5 μg / ml was captured at a flow rate of 10 μl / min for 120 seconds, resulting in a capture level of 350 to 1000 RU. To evaluate the KD value, FL TDP-43 or TP-51 peptide (amino acids 352 to 414 of SEQ ID NO: 1) was added to PBS-P + The binding kinetics were performed in a 3-fold dilution starting from 1.2 nM to 100 nM at a flow rate of 30 μl / min with single cycle kinetic injection for a 300 second contact time. The dissociation was recorded for 1 hour and then regenerated once with 10 mM glycine-HCl pH 1.7. The results obtained from the binding kinetics were double referenced using a blank flow cell and buffer cycle and evaluated using an overall 1:1 fitting model with RI. The association rates (on-rate) (ka), dissociation rates (off-rate) (kd) and affinities (KD) of the three antibodies are shown in Table 9 as the mean ± SD of 12 (ACI-7069-633B12-Ab1), 2 (ACI-7069-642D12-Ab1) or 3 (ACI-7071-809F12-Ab1) replicates. Antibodies ACI-7069-633B12-Abl, ACI-7069-642D12-Abl, and ACI-7071-809F12-Abl bound to soluble TDP-43 with affinities ranging from 15 to 135 pM, 226 to 272 pM, and 389 to 457 pM, respectively. Antibody ACI-7069-633B12-Abl bound to TP-51 peptide with an affinity ranging from 1184 to 1316 pM.

[0433] Table 9: Affinity for soluble FL TDP-43 and TP-51 peptides by SPR

[0434]

[0435] Example 9: Antibody Sequencing

[0436] Cloning hybridoma cell lysates are used for gene sequencing of variable regions. Mouse hybridomas are harvested and lysed using a lysis buffer containing guanidine salts to inactivate RNase. Genomic DNA is then eliminated by RNase-free DNase, and RNA is purified using a silica-based affinity column using multiple washes and eluted from the column using RNase-free water. Once RNA is extracted, its purity and concentration are measured spectrophotometrically. RNA integrity is assessed on a denaturing agarose gel, and reverse transcriptase (RT) is used to reverse transcribe RNA into cDNA. Before adding the RT reaction mixture, RNA is heated to 70°C for 10 minutes to destroy RNA secondary structure. The RT product is directly used for PCR amplification. For high-fidelity PCR amplification of cDNA, each of the variable region primers corresponding to different gene families encoding antibodies is separately mixed with the constant primers of VH and VL in a total reaction volume of 50 μl. Initially, a degenerate primer library (12 for VH and 12 for VL) is used, and based on the results, a second library is used to obtain PCR products. After the PCR reaction, the product was analyzed by gel electrophoresis on a 2% agarose gel stained with ethidium bromide. The PCR products of VL and VH were purified separately on agarose gel using tris acetate EDTA (TAE). The purified fragments cut from the gel were sequenced using the same primers as those used for PCR using a dye terminator sequencing method. Sequencing was performed in two directions to provide overlap at both ends. The sequences were analyzed using a multiple sequence alignment (Clustal tool) and annotated using the Kabat algorithm as described in Kabat et al., Sequences of Proteins of Immunological Interest, 91-3242 (1991). The nucleotide sequences of the heavy and light chain variable domains (VH and VL) are shown in Table 10. The translated protein sequences of the selected heavy chain (VH) and light chain (VL) variable domains and their complementary determining regions (CDRs) are shown in Table 11.

[0437] Table 10: Nucleotide sequences of heavy and light chain variable domains (VH and VL)

[0438]

[0439]

[0440]

[0441]

[0442]

[0443]

[0444] Table 11: Amino acid sequences of heavy and light chain variable domains (VH and VL) and their CDRs

[0445]

[0446]

[0447]

[0448]

[0449]

[0450]

[0451]

[0452] Example 10: In vivo efficacy of ACI-7069-633B12-Ab1 (IgG2a variant) in a transgenic mouse model of TDP-43 proteinopathy

[0453] To evaluate the in vivo efficacy of ACI-7069-633B12-Ab1 (IgG2a variant), the ability of ACI-7069-633B12-Ab1 (IgG2a variant) to reduce TDP-43 pathology in NEFH-tTA x hTDP-43ΔNLS double transgenic mice (rNLS8 mice, Walker et al. 2015) was tested. rNLS8 mice were injected weekly with ACI-7069-633B12-Ab1 (IgG2a variant) (n=30) or vehicle (n=30), and molecular pathology markers, such as phosphorylated TDP-43 and / or total insoluble TDP-43, were analyzed at the end of dosing.

[0454] 10.1 Animals

[0455] Before starting the study, all animals were acclimatized, checked, handled and weighed to ensure sufficient health and to minimize the nonspecific stress associated with the experimental manipulation. During the feeding period and before 8 weeks of age, mice were kept on a food diet containing doxycycline (200 mg / kg). At 8 weeks of age, the diet was changed to a food diet that did not contain doxycycline (DOX) to allow transgenic expression. Throughout this study, light / dark cycles (12 / 12), room temperature (20°C to 23°C) and relative humidity (about 50%) were kept constant. Food, diet and water were provided ad libitum during the study. When mice began to show difficulty in moving, the diet was changed to wet food and hydrogel on the cage bottom. All behavioral tests were carried out during the light cycle phase of the animal.

[0456] 10.2. Compound Administration

[0457] ACI-7069-633B12-Abl (IgG2a variant) (60 mg / kg) and vehicle were prepared fresh on the day of injection and administered ip following a weekly dosing schedule throughout the study.

[0458] 10.3. Brain Collection

[0459] The brain was divided into two hemispheres. The left hemisphere was dissected to collect the cortical brain area. The mouse cortex and the remaining brain tissue were quickly frozen for further biochemical analysis. The remaining right hemisphere was directly immersion fixed after perfusion for 3 hours at room temperature and collected in freshly prepared 1× PBS containing 4% paraformaldehyde (PFA).

[0460] Immunohistochemistry

[0461] The right cerebral hemisphere fixed by immersion was cut sagittally with a slice thickness of 10 microns on a Leica CM1950 cryostat. Systematic random sagittal section collections of each mouse (7 slices from the 2nd, 3rd, 4th, 6th, 8th, 10th and 11th levels of the brain) were performed for immunostaining of TDP-43 and phosphorylated TDP-43. Iba1 staining was performed to quantify the number and morphology of microglial cells in the brain. Antibody binding was visualized using a fluorescently labeled second antibody. Standard negative controls included wild-type brain slices and slices from transgenic animals to which no primary antibody was applied.

[0462] 10.5. Imaging and Determination of Immunoreactivity

[0463] Mounted sections were imaged as a whole on an Axio.Scan z1 slide scanner driven by ZEN software at 10× magnification using LED (Colibri2) illumination and a sensitive Orca Flash 4.0 monochrome camera. Brain dimensions were determined using separate delineations of regions of interest in the cerebral cortex and dorsal striatum. Object density (OD) (in terms of OD per mm) was determined using a 3D imager. 2 The number of objects (count) was determined for: all markers, percent labeled area, and OD relative to the second delineated region of interest size (excluding any tissue artifacts (tissue folds, etc.)).

[0464] 10.6. Preparation of protein samples from cerebral cortex:

[0465] Tissues were thawed on ice and then sonicated in 5X v / w radioimmunoprecipitation assay buffer (RIPA, 50 mM Tris, 150 mM NaCl, 1% IGEPAL CA630, 5 mM EDTA, 0.5% sodium deoxycholate, and 0.1% SDS, pH 8.0) containing 1 mM PMSF and a protease / phosphatase inhibitor cocktail (Roche Applied Science). Samples were centrifuged at 4°C, 100,000 g for 30 minutes, and the supernatant was considered the soluble fraction. The pellet was washed by sonication with RIPA, and the supernatant was discarded. The RIPA-insoluble pellet was sonicated in 2X v / w urea buffer (7 M urea, 2 M thiourea, 4% CHAPS, and 30 mM Tris, pH 8.5) and centrifuged at 22°C, 100,000 g for 30 minutes. The supernatant was considered the RIPA-insoluble / urea-soluble fraction. The protein concentration of the RIPA soluble fraction was determined using the BCA protein assay (Pierce).

[0466] 10.7. Quantification of insoluble TDP-43

[0467] Total TDP-43 levels in the RIPA-insoluble fraction were analyzed by a commercial human TDP-43 AlphaLISA kit (PerkinElmer, AL387HV).

[0468] Statistical Analysis

[0469] IHC and AlphaLISA data are expressed as mean ± SEM. The statistical difference between the animals treated with vehicle and the animals treated with ACI-7069-633B12-Ab1 (IgG2a variant) was analyzed by Welch t test and indicated by the asterisk above the corresponding bar (*p < 0.05, **p < 0.01, ****p < 0.0001). The outliers in histological measurements were excluded and were Grubbs outliers (Grubbs outlier) (single measurement) in the group or level, or due to technical reasons (image artifacts, tissue folds, etc.).

[0470] 10.9. Results

[0471] Treatment with ACI-7069-633B12-Ab1 (IgG2a variant) reduces phosphorylated TDP-43 and insoluble TDP-43 in rNLS8 mice

[0472] In the rNLS8 mouse model, overexpression of a DOX-repressible form of the K82A / R83A / K84A mutant human TDP-43 (hTDP-43ΔNLS) leads to a marked accumulation and aggregation of TDP-43 in the neuronal cytoplasm. The pathological hallmark of this model is the deposition of insoluble and phosphorylated TDP-43 inclusions (pTDP-43). These small, spherical cytoplasmic inclusions are present only in transgenic animals and are completely absent in WT or monogenic transgenic tTA control mice. Furthermore, pTDP-43 is generally absent during the first week in the absence of DOX and accumulates with a dramatic progression during the 3-4 week period when DOX is removed (Walker et al., 2015). Treatment with ACI-7069-633B12-Ab1 (IgG2a variant) results in a statistically significant decrease in the density of phosphorylated TDP-43 in both the striatum and cerebral cortex compared to vehicle-treated mice ( Figure 3 A to B), demonstrating its functional efficacy in reducing TDP-43 pathology. The striatum and cerebral cortex were chosen for quantification due to high transgene expression in these regions.

[0473] 10.10. Treatment with ACI-7069-633B12-Ab1 (IgG2a variant) reduces insoluble TDP-43 in rNLS8 mice

[0474] To confirm the reduction in TDP-43 pathology observed in the immunohistochemical readout, the amount of total insoluble / aggregated TDP-43 in the brain was quantified following biochemical fractionation. RIPA insoluble fractions were prepared from the cortex of the left cerebral hemisphere, which contained insoluble / aggregated TDP-43. A significant decrease in the amount of insoluble TDP-43 was observed in mice treated with ACI-7069-633B12-Ab1 (IgG2a variant) compared to vehicle-treated animals ( Figure 3 C). This reduction in molecular TDP-43 pathology is consistent with the results observed by immunohistochemistry, confirming the efficacy of treatment with ACI-7069-633B12-Ab1 (IgG2a variant). To our knowledge, this is the first time that administration of a peripheral antibody has ameliorated the development of TDP-43 pathology in an in vivo model for a TDP-43 proteinopathy.

[0475] ACI-7069-633B12-Ab1 (IgG2a variant) treatment in rNLS8 mice increases the area of ​​microglial immune response

[0476] Functional recovery in rNLS8 mice after suppression of transgenic expression involved an increase in microglial activity. Microglial cell body area increased at this stage and led to clearance of TDP-43 pathology and functional restoration of motor deficits, demonstrating a therapeutic paradigm in the rNLS8 mouse model (Spiller KJ et al., Nature Neuroscience, 2018).

[0477] To evaluate the mode of action of ACI-7069-633B12-Ab1 in reducing TDP-43 pathology in rNLS8 mice, its effect on microglial activation was assessed. Iba1 staining was performed by immunohistochemistry to quantify the number and status of microglia in the mouse cerebral cortex. Microglial proliferation was found in rNLS8 mice at the end of the Dox period (5 weeks). ACI-7069-633B12-Ab1 treatment significantly increased the area of ​​Iba1 positive immunoreactivity in the cortex ( ) compared to vehicle-treated controls. Figure 5 A). This increase may be due to an increase in microglial number or changes in microglial morphology. To this end, the density of Iba1-positive cells in the cortex was first assessed. Compared to vehicle-treated controls, ACI-7069-633B12-Ab1 treatment did not affect microglial density, which represents cell number.

[0478] Next, the effect of ACI-7069-633B12-Ab1 on microglial morphology was evaluated. In order to correlate the increase in Iba1 immunoreactive area with changes in the activation state of microglial cells, which represents morphology, microglial cells were classified into three states (large hypertrophic, small branched, and branched quiescent) based on their size and morphology. Compared with the control treated with vehicle, the average cell size of large hypertrophic microglia was significantly increased in ACI-7069-633B12-Ab1 (IgG2a variant) treatment ( Figure 5 B). No significant differences were found in the other two types of microglia representing lower activation states ( Figure 5 C to D). This analysis demonstrates that the increase in total Iba1-positive immunoreactive area observed in the ACI-7069-633B12-Ab1-treated group is due to morphological changes that reflect increased microglial size and activation. This suggests that ACI-7069-633B12-Ab1 (IgG2a variant) reduces TDP-43 pathology in this animal model, at least in part, by recruiting and activating microglia.

[0479] Example 11: In vitro function of ACI-7069-633B12-Ab1 (IgG2a variant) in a recombinant TDP-43 aggregation assay

[0480] To evaluate the in vitro function of ACI-7069-633B12-Ab1 (IgG2a variant), the ability of ACI-7069-633B12-Ab1 (IgG2a variant) to inhibit TDP-43 aggregation was tested. FL TDP-43 was fused at the C-terminus to a recombinantly produced maltose binding protein (MBP) separated by a Tobacco Etch Virus (TEV) protease cleavage site. Aggregation of 2.5 μM TDP-43-TEV-MBP fusion protein in 30 mM Tris, 150 mM NaCl, pH 7.4 in the presence of 2.5 μM ACI-7069-633B12-Ab1 (IgG2a variant) or an isotype control that does not bind to TDP-43 was induced by adding TEV protease (AcTEV, Invitrogen) and the absorbance was monitored at 600 nm in μclear 96-well plates (Greiner) over 30 h. For evaluation, the endpoint was normalized to the isotype control and the percentage of aggregated TDP-43 was calculated for ACI-7069-633B12-Ab1. Antibody ACI-7069-633B12-Ab1 significantly inhibited TDP-43 aggregation by 98% (98%) compared to the isotype control. Figure 4 ).

[0481] Example 12: Detection and Quantification of TDP-43 in Biological Fluids Using ACI-7069-633B12-Abl (IgG2a Variant) and ACI-7071-809F12-Abl (IgG2a Variant)

[0482] Methods: A PerkinElmer bead-based AlphaLISA immunoassay was developed using ACI-7069-633B12-Ab1 (IgG2a variant) and ACI-7071-809F12-Ab1 (IgG2a variant). Dilution linearity was established in spike-recovery experiments for CSF samples. TDP-43 concentrations were then measured in diluted CSF samples. The assay was performed on a white optiplate. TM Samples were prepared in a 384 microplate and the emission at 615 nm was measured as raw AlphaLISA counts.

[0483] Results: In this immunoassay, total TDP-43 was quantified in cerebrospinal fluid (CSF) samples from healthy controls and patients with FTLD-TDP (semantic dementia, C9orf72, or GRN). Figure 6 In three independent experiments, relative TDP-43 quantification of multiple patient CSF samples from FTLD-TDP patients with GRN mutations showed significantly higher TDP-43 levels compared to healthy controls ( Figure 6 In three independent experiments, relative TDP-43 quantification of multiple patient CSF samples from FTLD-TDP patients with C9orf72 mutations and semantic dementia also showed higher TDP-43 levels compared to healthy controls ( Figure 6 ).

[0484] Example 13: Binding to pathological TDP-43 assessed by immunodepletion in FTD brain extracts

[0485] To evaluate the efficacy of the antibodies in specifically binding to TDP-43 aggregates in their native state, immunodepletion experiments in brain extracts enriched for pathological TDP-43 were performed.

[0486] Methods: The insoluble fraction from FTD type A (FTD-A) postmortem brain was prepared as described in Example 7. Immunodepletion was performed using DynabeadsTM magnetic beads, protein G (Thermoscientific 10003D). After resuspension in the tube, 130 μl of beads were transferred to a 1.5 ml low binding tube. The beads were rinsed twice with PBS supplemented with 0.05% Tween 20 using a magnet to remove the supernatant. The beads were divided equally among three different low binding tubes. Antibodies (ACI-7069-633B12-Ab1 (IgG2a isotype), ACI-7069-642D12-Ab1 (IgG2a isotype), mouse IgG2a control) were diluted to 100 μg / ml and 100 μl was added to each tube after removing the supernatant (using a magnet). The antibody-bead mixture was incubated at room temperature for 1 hour. The beads-antibody complex was washed once with 500 μl PBS-0.05% Tween 20 and once with PBS, and then resuspended in 250 μl PBS. The antibody-beads were divided into two new tubes (120 μl per tube). The insoluble fraction was thawed on ice and sonicated on ice at an amplitude of 30 for 30 seconds. After removing the supernatant, 30 micrograms of brain material was added to each antibody-bead tube and incubated overnight at 4°C under continuous rotation. The tube was placed on a magnet and the supernatant was collected as the fraction after immunodepletion. The input material and the immunodepleted material were further analyzed by Western blotting. Western blotting was performed as described in Example 7. Each lane was loaded with 20 μl of sample. Immunoblotting was performed using the following antibodies: total TDP-43 (ACI-7069-633B12-Ab1 coupled to DyLight680), pTDP-43 (Biolegend, 829901) at a dilution of 1:2000 and 1:1000, respectively. Goat anti-rat secondary antibody (Cat. No. 925-32219) was used at a dilution of 1:10000.

[0487] Results: Compared to the isotype control antibody, ACI-7069-633B12-Ab1 and ACI-7069-642D12-Ab1 were able to specifically bind to and deplete TDP-43 and pTDP-43 from the sarkosyl-insoluble fraction obtained from FTD type A brain tissue ( Figure 7 ). This data establishes the properties of these antibodies for engagement with their targets in human patients.

[0488] References

[0489] Arai et al.,TDP-43 is a component of ubiquitin-positive tau-negativeinclusions in frontotemporal lobar degeneration and amyotrophic lateralsclerosis,Biochemical and Biophysical Research Communications 351(2006)602-611.

[0490] Buratti and Barallc,Nuclear factor TDP-43 can affcct sclcctcdmicroRNA levels,FEBS Journal 277 (2010)2268-2281.

[0491] Brettschneider J et al.,Spreading of pathology in neurodegenerativediseases:a focus on human studies,Nature Rev.Neuroscience,2015,109.

[0492] Brettschneider et al.,Stages of pTDP-43 pathology in amyotrophiclateral sclerosis,Ann Neurol.2013 July;74(1):20-38.

[0493] Charlton,Methods in Molecular Biology,Val.248(B.K.C.Lo,ed.,HumanaPress,Totowa,NJ,2003),pp.245-254.

[0494] Chowdhury,Methods Mol.Biol.207:179-196(2008)

[0495] Clyncs et al.,Fc receptors are required in passive and activeimmunity to melanoma Proc.Nat’! Acad.sci.USA 95:652-656(1998)

[0496] Cohen et al.,An acetylation switch controls TDP-43 function andaggregation propensity,Nat Commun.6:5845,2015.

[0497] Cragg.M.S.et al.,Complement-mediated lysis by anti-CD20 mAbcorrelates with segregation into lipid rafts,Blood 101:1045-1052(2003)

[0498] Cragg.M.S.and M.J.Glennie,Antibody specificity controls in vivocffector mechanisms of anti-CD20 reagents,Blood 103:2738-2743(2004)).

[0499] Cunningham and Wells,High-resolution epitope mapping of hGH-receptorinteractions by alanine-scanning mutagenesis.(1989)Science.244:1081-1085.

[0500] Duncan&Winter,The binding site for C1q on IgG,Nature 322:738-40(1988)

[0501] Edclman.G.M.et al.,The Covalent Structure of an Entire gammaGImmunoglobulin Molecule,Proc.Natl.Acad.USA,63,78-85(1969)

[0502] Feiler et al.,TDP-43 is intercellularly transmitted across axonTerminals,J.Cell Biol.Vol.211 No.4897-911.

[0503] Feneberg et al.,Towards a TDP-43-Based Biomarker for ALS andFTLD.Molecular Neurobiology,2018;55(10):7789-7801.

[0504] Gazzano-Santoro et al.,Engineered Antibodies with Increased Activityto Recruit Complement,J.Immunol.Methods 202:163(1996)

[0505] Gemgross,Advances in the production of human therapculic proteins inyeasts and filamentous fungi,Nat.Biotech.22:1409-1414(2004).

[0506] Gerhardt et al.,Methods for General and Molecular Bacteriology,ASMPress(1994).

[0507] Golemis,Protein-Protein Interactions;A Molecular Cloning Manual,ColdSpring Harbor Laboratory Press(2002).

[0508] Graham et al.,Characteristics of a human cell line transformed by DNAfrom human adenovirus type 5.J.Gen Viral.36:59(1977)

[0509] Guyer et al.,Immunoglobulin binding by mouse intestinal epithelialcell receptors,J.Immunol.117:587(1976).

[0510] Harlow and Lane,Antibodies:A Laboratory Manual,Cold Spring HarborLaboratory Press,(1988).

[0511] Harlow and Lane.Using Antibodies:A Laboratory Manual,Cold SpringHarbor Laboratory Press,(1999).

[0512] Hascgawa et al.,Phosphorylated TDP-43 in frontotemporal lobardegeneration and amyotrophie lateral sclerosis,(2008)Annals of Neurology Vol64 No 1,60-70.

[0513] Hascgawa et al.,Prion-like mechanisms and potential therapeutictargets in neurodegenerative disorders.Pharmacol Ther.2017 Apr,172:22-33.

[0514] Hellstrom,I.et al.,Proc.Nat’l Acad.Sci.USA 83:7059-7063(1986).

[0515] Hellstrom,I et al,Proc.Nat’l Acad.Sci.USA 82:1499-1502(1985).

[0516] Hoogenboom et al.,Methods in Molecular Biology 178:1-37(O’Brien etal.,ed.,Human Press,Totowa,NJ,(2001).

[0517] Howard and Bcthell.(2000)Basic Methods in Antibody Production andCharacterization, Cre.Pr.Inc.Idusogie et al.,J.Immunol.164:4178-4184(2000).

[0518] Ishii et al.,Formation and spreading of TDP-43 aggregates in culturedneuronal and glial cells demonstrated by time-lapse imaging,PLoS ONE 12(6):e0179375,2017

[0519] Jones et al.,Replacing the complementarity-dctermining regions in ahuman antibody with those from a mouse,Nature 321(1986).522-525.

[0520] KA et al.,TDP-43 is a key player in the clinical features associatedwith Alzheimer’s disease,Acta Neuropathol.2014:127(6):811-824.

[0521] KA et al.,Staging TDP-43 pathology in Alzheimer’s disease ActaNeuropatbol.2014:127(3):441-450.

[0522] Kam et al., Proc.Natl.Acad.Sci.USA 102:11600-11605(2005).

[0523] Kanda Y.et al.,Biotcchnol.Biocng.,94(4):680-688(2006).

[0524] Kabat et al.,Sequences of Proteins of Immunological Interest.5thedit.NIH Publication no.91-3242U.S.Department of Health and Human Services(1991).

[0525] Kim et al.,J.Immunol.24:249(1994).

[0526] Kohler,Nature 256(1975),495.

[0527] Khor,Proceedings of the American Society of Clinical OncologyAbstract(1997),847.

[0528] Lagier-Tourenne et al.,TDP-43 and FUS / TLS:emerging roles in RNAprocessing and neurodegeneration,Human Molecular Genetics,2010,Vol.19,ReviewIssue 1 R46-R64.

[0529] Lagier-Tourenne and Cleveland.Rethinking ALS:the FUS about TDP-43,Cell 136,2009.1001-1004.

[0530] Le Ber,Genetics of frontotemporal lobar degeneration:an up-date anddiagnosis algorithm,Revue Neurologique 169(2013)811-819.

[0531] Lefkovits,Immunology Methods Manual:The Comprehensive Sourcebook ofTechniques:Academic Press(1997).

[0532] LoBuglic,Proceedings of the American Society of Clinical OncologyAbstract(1997).1562.

[0533] Li et al.,Optimization of humanized IgGs in glycoengineered Pichiapastoris,Nat.Biotech.24:210-215(2006).

[0534] Mackenzie and Neumann,Molecular neuropathology of frontotemporaldementia:msights into disease mechanisms from postmortem studies,J.Ncurochem.(2016)138(Suppl.1),54-70.

[0535] Mather,Establishment and characterization of two distinet mousetesticular epifheliat cell lines,Biol.Reprod.23:243-251(1980)

[0536] Mather et al.,Culture of testicular cells in hormone-supplemcntedserum-free medium,Annals N.Y Acad.Sci.383:44-68(1982)

[0537] McAleese et al.,TDP-43 pathology in Alzheimer’s disease,dementia withLewy bodies and ageing,Brain Pathol.2017 Jul;27(4):472-479.

[0538] Morris,Epitope Mapping Protocols,Methods in Molecular Biology vol.66(1996)(Humana Press,Totowa. NJ).

[0539] Nonaka et al.,Prion-like Properties of Pathological TDP-43 Aggregatesfrom Diseased Brains,Cell Reports 4(2013),124-134.

[0540] Neumann et al.,Ubiquitinated TDP-43 in frontotemporal lobardegencration and amyotrophic lateral sclerosis,Science 314,(2006),130-133.

[0541] Neumann et al.,Phosphorylation of S409 / 410 of TDP-43 is a consistentfeature in all sporadic and familial forms of TDP-43 proteinopathies,ActaNeuropathol.(2009)117:137-149.

[0542] Petkova,S.B.et al.,Int′l.Immunol.18(12):1759-1769(2006).

[0543] Plückthun,The Pharmacology of Monoclonal Antibodies.Rosenburg andMoore eds.Springer-Verlag,N.Y.(1994),269-315.

[0544] Porta S.et al.,Patient-derived frontotemporal lobar degenerationbrain extracts induce formation and spreading of TDP-43 pathology in vivoNat.Comm.,2018

[0545] Okazaki et al.,Fucose depiction from human IgG1 oligosaccharidecnhances binding enthalpy and association rate between IgG1 andFcgammaRIlla.J.Mol.Biol.336:1239-1249(2004).

[0546] Presta LG.,Antibody engineering,Curr Op Struct Biol 2(1992),593-596.

[0547] Ravctch and Kinet,Annu.Rev.Immunol.9:457-492(1991)

[0548] Reichmann.Reshaping human antibodies for therapy,Nature 332(1998).323-327.

[0549] Remington′s Pharmaceutical Sciences 16th edition,Osol,A.Ed.(1980)

[0550] Ripka et al.,Two Chinese hamster ovary glyeosylation mutants affectedin the conversion of GDP-mannose to GDP-fucose Arch.Biochem. Biophys.249:533-545(1986).

[0551] Sambrook et al.,Molecular Cloning:A Laboratory Manual;Cold SpringHarbor Laboratory Press,2nd edition(1989)and 3rd edition(2001).

[0552] Shepherd and Dean(2000),Monoclonal Antibodies:A Practical Approach,Oxford University Press Shields et al.,High Resolution Mapping of the BindingSite on Human IgG1 for FcγRI,FcγRII,FcγRIII,and FcRn and Design of IgG1Variants with Improved Binding to the FcγR,J.Biol.Chem.9(2):6591-6604(2001).

[0553] Spiller K.J et al.,Microglia-mediated rccovcry from ALS-relevantmotor neuron degencration in a mouse model of TDP-43 protcinopathy,NatureNeuroseience,2018

[0554] Ticozzi et al.,Protein Aggregation and Defective RNA Metabolism asMechanisms for Motor Neuron Damage,9(3):285-296 CNS Neurol.Disord.DrugTargets.2010,9(3),285-296.

[0555] Urlaub et al.,Isolation of Chinese hamster cell mutants deficient indihydrofolate reductasc activity,Proc.Natl.Acad. eli.USA 77:4216(1980)

[0556] Vcrhocyen et al.,Reshaping human antibodies:Grafting an antilysozymeactivity.,Science 239(1988),1534-1536.

[0557] Walker et al.,Neuropathologically mixed Alzheimer’s and Lcwy bodydisease:burden of pathological protein aggregates differs between clinicalphenotypes,Acta Neuropathol(2015)129:729-748

[0558] Wang et al.,TDP-43:an emerging new player in neurodegenerativediseases Trends in Molecular Medicine Vol.14 No.11,2008,479-485.

[0559] Warraich et al.,TDP-43:a DNA and RNA binding protein with roles inneurodegenerative diseases,The Intemational Journal of Biochemistry&CellBiology 42(2010)1606-1609.Wright et al.,Effect of glycosylation on antibodyfunction.TIBTECH 15:26-32(1997).

[0560] Yamane-Ohnuki et al.,Establishment of FUT8 knockout Chinese hamsterovary cells:An ideal host cell line for producing completely defucosylatedantibodies with enhanced antibody-dependent cellulareytotoxicity.Biotech.Bioeng.87:614(2004).

[0561] Yazaki and Wu,Methods in Molecular Biology,Val.248(B.K.C.Lo,ed.,Humana Press,Totowa,NJ),pp.255-268(2003).

Claims

1. A TDP-43 binding molecule which is a murine antibody or antigen-binding fragment thereof that binds to misfolded, aggregated TDP-43 and non-aggregated physiological TDP-43, that binds to an epitope within amino acid residues 400 to 405 or 400 to 406, and which comprises: a) a VH-CDR1 consisting of the amino acid sequence of SEQ ID NO: 11; a VH-CDR2 consisting of the amino acid sequence of SEQ ID NO: 12; and a VH-CDR3 consisting of the amino acid sequence ES (Glu-Ser); a VL-CDR1 consisting of the amino acid sequence of SEQ ID NO: 15; a VL-CDR2 consisting of the amino acid sequence of SEQ ID NO: 16; and a VL-CDR3 consisting of the amino acid sequence of SEQ ID NO: 17; or b) a VH-CDR1 consisting of the amino acid sequence of SEQ ID NO: 21; a VH-CDR2 consisting of the amino acid sequence of SEQ ID NO: 22; and a VH-CDR3 consisting of the amino acid sequence ES (Glu-Ser); a VL-CDR1 consisting of the amino acid sequence of SEQ ID NO: 25; a VL-CDR2 consisting of the amino acid sequence of SEQ ID NO: 16; and a VL-CDR3 consisting of the amino acid sequence of SEQ ID NO: 27; or chimeric or humanized versions thereof.

2. The TDP-43 binding molecule of claim 1, which binds to misfolded aggregated human TDP-43 and non-aggregated physiological human TDP-43.

3. The TDP-43 binding molecule of claim 1, comprising: a. a heavy chain variable region (VH) consisting of the sequence of SEQ ID NO: 10 and a light chain variable region (VL) consisting of the sequence of SEQ ID NO: 14; or b. A heavy chain variable region (VH) consisting of the sequence of SEQ ID NO: 20 and a light chain variable region (VL) consisting of the sequence of SEQ ID NO:

24.

4. The TDP-43 binding molecule of claim 1, comprising: a. VH-CDR1 consisting of the amino acid sequence of SEQ ID NO: 21; VH-CDR2 consisting of the amino acid sequence of SEQ ID NO: 22; and VH-CDR3 consisting of the amino acid sequence ES (Glu-Ser); VL-CDR1 consisting of the amino acid sequence of SEQ ID NO: 25; VL-CDR2 consisting of the amino acid sequence of SEQ ID NO: 16; and VL-CDR3 consisting of the amino acid sequence of SEQ ID NO: 27; and / or b. A heavy chain variable region (VH) consisting of the sequence of SEQ ID NO: 20 and a light chain variable region (VL) consisting of the sequence of SEQ ID NO:

24.

5. The TDP-43 binding molecule of claim 4, wherein: a. Alleviate TDP-43 pathology in vivo; and / or b. reducing the level of aggregated TDP-43 and / or phosphorylated TDP-43 in vivo. The TDP-43 binding molecule of claim 1 , which is a monoclonal antibody or an antigen-binding fragment thereof.

7. The TDP-43 binding molecule of claim 1, which is an IgA, IgD, IgE, IgM, IgG1, IgG2, IgG2a, IgG2b, IgG3 or IgG4 antibody, or an antigen-binding fragment thereof.

8. The TDP-43 binding molecule of claim 4, for use in preventing, alleviating, treating and / or diagnosing a disease, disorder and / or abnormality associated with TDP-43, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD) or amyotrophic lateral sclerosis (ALS).

9. A pharmaceutical composition comprising the TDP-43 binding molecule according to any one of claims 1 to 7, and a pharmaceutically acceptable carrier and / or excipient.

10. A nucleic acid molecule encoding the TDP-43 binding molecule of claim 1, wherein the nucleic acid molecule comprises a nucleotide sequence as shown below: a. the heavy chain variable region (VH) coding sequence of SEQ ID NO: 18 and the light chain variable region (VL) coding sequence of SEQ ID NO: 19; or b. The heavy chain variable region (VH) coding sequence of SEQ ID NO: 28 and the light chain variable region (VL) coding sequence of SEQ ID NO:

29. An expression vector comprising the nucleic acid molecule according to claim 10 .

12. A host cell comprising the nucleic acid molecule of claim 10 and / or the expression vector of claim 11.

13. A host cell expressing the TDP-43 binding molecule according to any one of claims 1 to 7. A cell-free expression system comprising the expression vector according to claim 11 .

15. A method for producing a TDP-43 binding molecule, wherein the TDP-43 binding molecule is an antibody or an antigen-binding fragment thereof, the method comprising the steps of: a) culturing the host cell of claim 12 or 13 or the cell-free expression system of claim 14 under conditions suitable for production of the binding molecule; and b) isolating said binding molecule.

16. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for quantifying TDP-43 in a sample obtained from a subject, wherein the kit is configured to contact the sample with the TDP-43 binding molecule according to any one of claims 1 to 7, and compare the level of TDP-43 in the sample with the level of TDP-43 in a control sample.

17. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for diagnosing a disease, disorder and / or abnormality associated with TDP-43, wherein the kit is formulated for use according to claim 16, wherein a higher level of TDP-43 in the sample compared to a control level based on a healthy subject is indicative of a disease, disorder and / or abnormality associated with TDP-43, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD), Down syndrome, familial English dementia, polyglutamine disease, hippocampal sclerosis dementia, myopathy, traumatic brain injury (TBI), dementia with Lewy bodies (DLB), or Parkinson's disease (PD).

18. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for diagnosing a disease, disorder and / or abnormality associated with TDP-43, wherein the kit is formulated for performing the use of claim 16 or the use of claim 17, wherein similar or higher levels of TDP-43 in the sample compared to a diseased control are indicative of a disease, disorder and / or abnormality associated with TDP-43, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD), Down syndrome, familial British dementia, polyglutamine disease, hippocampal sclerosis dementia, myopathy, traumatic brain injury (TBI), dementia with Lewy bodies (DLB) or Parkinson's disease (PD).

19. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for classifying diseases, disorders and / or abnormalities associated with TDP-43, wherein the classification comprises: a. Carry out the use according to claim 17 or 18; b. optionally identifying a mutation in the sample, including but not limited to a progranulin (GRN) mutation, a C9orf72 mutation, a TARDBP mutation, an angiogenic protein (ANG) mutation, a mutation in a valosin-containing protein (VCP), a myotilin (MYOT) gene mutation, or a mutation in a gene encoding desmin (DES); and c. Classifying the disease, disorder and / or abnormality associated with TDP-43, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD), Down syndrome, familial British dementia, polyglutamine disease, hippocampal sclerosis dementia, myopathy, traumatic brain injury (TBI), Lewy body dementia (DLB) or Parkinson's disease (PD).

20. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for monitoring diseases, disorders and / or abnormalities associated with TDP-43 at two or more time points using a sample from a subject, wherein the kit is formulated for contacting the sample with a TDP-43 binding molecule according to any one of claims 1 to 7, wherein: a. a higher level of TDP-43 in a later sample compared to one or more earlier samples indicates progression of a disease, disorder, and / or abnormality associated with TDP-43; or b. lower TDP-43 levels in the later sample compared to one or more earlier samples indicates resolution of the disease, disorder, and / or abnormality associated with TDP-43; wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD), Down syndrome, familial British dementia, polyglutamine disease, hippocampal sclerosis dementia, myopathy, traumatic brain injury (TBI), Lewy body dementia (DLB) or Parkinson's disease (PD).

21. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for monitoring the treatment of a disease, disorder and / or abnormality associated with TDP-43 at two or more time points using samples from a subject being treated with a specific therapy, wherein the kit is formulated for contacting the sample with a TDP-43 binding molecule according to any one of claims 1 to 7, wherein a lower level of TDP-43 in a later sample compared to one or more earlier samples is indicative of a disease, disorder and / or abnormality associated with TDP-43. or abnormality is successfully treated, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD), Down syndrome, familial British dementia, polyglutamine disease, hippocampal sclerosis dementia, myopathy, traumatic brain injury (TBI), dementia with Lewy bodies (DLB) or Parkinson's disease (PD).

22. The use of claim 21, wherein the first time point is prior to treatment with the therapy and the second time point is after treatment with the therapy.

23. Use of a reagent for determining the level of TDP-43 in a sample in the preparation of a kit for selecting a therapy for treating a disease, disorder and / or abnormality associated with TDP-43, wherein the kit is formulated for contacting samples collected before and after treatment with the therapy with a TDP-43 binding molecule according to any one of claims 1 to 7, wherein a lower level of TDP-43 in the sample collected after treatment compared to the sample collected before treatment indicates that the disease, disorder and / or abnormality associated with TDP-43 is successfully treated, and And thus the treatment is selected for treatment wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alexander disease (AxD), limbic-dominant age-related TDP-43 encephalopathy (LATE), chronic traumatic encephalopathy (CTE), Perry syndrome, Alzheimer's disease (AD), Down syndrome, familial British dementia, polyglutamine disease, hippocampal sclerosis dementia, myopathy, traumatic brain injury (TBI), dementia with Lewy bodies (DLB) or Parkinson's disease (PD).

24. The use of claim 23, wherein the treatment comprises a TDP-43 binding molecule according to any one of claims 1 to 7 or a pharmaceutical composition according to claim 9.

25. The use of any one of claims 16, 17, 20, 21 or 23, wherein the sample comprises a blood, CSF, ISF or urine sample.

26. The use according to any one of claims 17, 20, 21 or 23, wherein the disease, disorder and / or abnormality associated with TDP-43 is sporadic frontotemporal dementia, familial frontotemporal dementia with motor neuron disease (MND), familial frontotemporal dementia without motor neuron disease (MND), frontotemporal dementia with progranulin (GRN) mutation, frontotemporal dementia with C9orf72 mutation, frontotemporal dementia with TARDBP mutation, frontotemporal dementia with valosin-containing protein (VCP) mutation, frontotemporal dementia linked to chromosome 9p, corticobasal degeneration, frontotemporal lobar degeneration (FTLD) with ubiquitin-positive TDP-43 inclusions (FTLD-TDP), argyrophilic grain disease, Pick's disease, semantic variant primary progressive aphasia (svPPA), behavioral variant FTD (bvFTD), nonfluent variant primary progressive aphasia (nfvPPA), sporadic amyotrophic lateral sclerosis, amyotrophic lateral sclerosis with a TARDBP mutation, amyotrophic lateral sclerosis with angiogenic protein (ANG) mutations, sporadic forms of Alzheimer's disease, familial forms of Alzheimer's disease, Huntington's disease, spinocerebellar ataxia type 3 (SCA3), myofibrillar myopathy with a mutation in the myotilin (MYOT) gene or a mutation in the gene encoding desmin (DES), oculopharyngeal muscular dystrophy with rimmed vacuoles, sporadic inclusion body myositis, or inclusion body myopathy with a mutation in a valosin-containing protein.

27. The use according to any one of claims 17, 20, 21 or 23, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), chronic traumatic encephalopathy (CTE) or limbic-dominant age-related TDP-43 encephalopathy (LATE).

28. The use according to any one of claims 17, 20, 21 or 23, wherein the disease, disorder and / or abnormality associated with TDP-43 is amyotrophic lateral sclerosis (ALS).

29. The use according to any one of claims 17, 20, 21 or 23, wherein the disease, disorder and / or abnormality associated with TDP-43 is Alzheimer's disease (AD).

30. The use according to any one of claims 17, 20, 21 or 23, wherein the disease, disorder and / or abnormality associated with TDP-43 is frontotemporal dementia (FTD).

31. A kit for diagnosing a disease, disorder and / or abnormality associated with TDP-43, or for the use according to any one of claims 16 to 30, comprising a TDP-43 binding molecule according to any one of claims 1 to 7.

Images