HUMANIZED ANTI-BETA AMYLOID N-TRUNCATED MONOCLONAL ANTIBODY

Abstract

The present invention relates to an antibody comprising a heavy chain variable domain (VH domain) and a light chain variable domain (VK domain), characterized in that a) the amino acid sequence of the VH domain is defined in SEQ ID NO: 3, the amino acid sequence of the VK domain is defined in SEQ ID NO: 7; ob) the amino acid sequence of the VH domain is defined in SEQ ID NO: 4, and the amino acid sequence of the VK domain is defined in SEQ ID NO: 7; oc) the amino acid sequence of the VH domain is defined in SEQ ID NO: 5, and the amino acid sequence of the VK domain is defined in SEQ ID NO: 8; od) the amino acid sequence of the VH domain is defined in SEQ ID NO: 5, and the amino acid sequence of the VK domain is defined in SEQ ID NO: 7; and where the antibody binds to the amyloid peptides AßpE3-42 and Aß-42 and does not bind to the amyloid peptide Aß1-42.

Description

HUMANIZED ANTI-N-TRUNCATED BETA AMYLOID MONOCLONAL ANTIBODY FIELD OF INVENTION The present invention relates to humanized antibodies that bind to amyloid peptides. BACKGROUND OF THE INVENTION The murine anti-amyloid beta (Aβ) antibody NT4X-167 was initially designed against the amyloid peptide Aβ4-40 and is reported to bind specifically to the N-truncated amyloid peptides AβρE3-42 and Aβ4-42 but not to the amyloid peptide Aβ1-42 (Antonios et al., Acta Neuropathol. Commun. (2013) 6:156). Passive immunization using NT4X-167 has been shown to be therapeutically beneficial in mouse models of Alzheimer's disease (Antonios et al., Scientific Reports 5:17338; 2015). Humanized versions of NT4X-167 could be useful for clinical applications, for example, in the treatment of Alzheimer's disease (AD). BRIEF DESCRIPTION OF THE INVENTION The inventors of this product have unexpectedly discovered that the binding activity of humanized versions of the NT4X-167 antibody is enhanced by mutating certain residues within the heavy and / or light chain variable domains. This may be useful, for example, in the development of candidate molecules for clinical use. A first aspect of the invention provides an anti-Aβ antibody comprising a heavy-chain variable domain and a light-chain variable domain, wherein a) the variable heavy chain domain (VH domain) comprises SEQ ID NO:2 with four or fewer additional alterations, such as substitutions, in the frame regions, and b) the variable light chain domain (VK domain) comprises SEQ ID NO:6 optionally with up to four or fewer additional alterations, such as substitutions, in the frame regions. The anti-Aβ antibody can specifically bind to N-terminal truncated amyloid peptides (AβρE3-χ or Aβ4-χ). For example, the anti-Aβ antibody can specifically bind to one or more, preferably all, of AβρE3-38, AβρE3-40, AβρE3-14, AβρE3-42, Aβ4-38, Aβ4-40, Aβ4-14, and Aβ4-42. / αορηη / ίζηζ / Ε / νίΛΐ The anti-Aβ antibody may not show specific binding to full-length amyloid peptides or amyloid peptides without N-terminal truncations (Aβί-χ), such as Aβ1-42, Aβ1-38, Aβ1-40, or Aβ1-14. Preferably, the heavy chain variable domain (VH domain) of the anti-Aβ antibody comprises SEQ ID NO: 3, SEQ ID NO: 4 or SEQ ID NO: 5. Preferably, the variable light chain domain (VL domain) of the anti-Aβ antibody comprises SEQ ID NO: 7 or SEQ ID NO: 8. A second aspect described herein provides a pharmaceutical composition comprising an antibody of the first aspect and a pharmaceutically acceptable carrier. A third aspect described herein provides a nucleic acid encoding an antibody of the first aspect or a heavy chain variable domain and / or light chain variable domain thereof. A fourth aspect described herein provides a vector comprising a nucleic acid of the third aspect. A fifth aspect described herein provides a host cell comprising a nucleic acid of the third aspect or a vector of the fourth aspect. A sixth aspect described herein provides a method for making an antibody according to the first aspect; the method comprises expressing, in a host cell culture, a vector according to the fourth aspect to produce said antibody; and recovering the antibody from the cell culture. A seventh aspect described herein provides a method for the treatment of Alzheimer's disease by administering, to an individual in need of such treatment, an effective amount of an antibody according to the first aspect or the pharmaceutical composition according to the second aspect. An eighth aspect described herein provides an antibody according to the first aspect or the pharmaceutical composition according to the second aspect, for use in a method for treating the human or animal body. A ninth aspect described herein provides an antibody according to the first aspect or the pharmaceutical composition according to the second aspect, for use in a method for treating Alzheimer's disease in an individual. These and other aspects and modalities described herein are described in more detail below. / QOrnn / L7Π7 / E / YILI BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 shows the binding of the murine antibody NT4X-167 to amyloid peptides. Figure 2 shows the binding of murine and chimeric NT4X-167 antibodies to PSL amyloid peptides. Figure 3 shows the binding of murine, humanized, and chimeric NT4X-167 to AβρE3-42 amyloid peptides: Initial versions Figure 4 shows the joining of the humanized variants to Aβ1-42. Figure 5 shows the joining of the humanized variants to AβρE3-42. Figure 6 shows the binding of the second round of humanized NT4X-167 antibodies to the AβρE3-42 peptide: HC to HR versions in combination with RKA Figure 7 shows the binding of the second round of humanized NT4X-167 antibodies to the AβρE3-42 peptide by means of ELISA. Figure 8 shows the binding of the third round of humanized NT4X-167 antibodies to the AβρE3-42 peptide: HS to HY versions in combination with RKA Figure 9 shows the binding of the fifth round of humanized NT4X-167 antibodies to the AβρE3-42 peptide: Versions rcNT4XS6A, rcNT4XS7A, rcNT4XS8A in combination with RKA Figure 10 shows the thermal stability of the humanized rcNT4X_SA, BA, TA, UA, VA, WA, YA antibody that binds to the amyloid peptide AβρE3-42 Figure 11 shows the Thermal Change Analysis of the humanized rcNT4X_SA and rcNT4X_S7A antibodies. Figure 12 shows non-specific protein-protein interactions (cross-interaction chromatography) Figure 13 shows the non-specific protein-protein interactions (cross-interaction chromatography) of the lead humanized candidate rcNT4XS7A. Figure 14 shows the purified antibody candidates evaluated for solubility. Figure 15 shows the bonding of serum stability assessment of the humanized antibody rcNT4X_SA and rcNT4X_S7A to the amyloid peptide PSL AβρE3-42. Figure 16 shows the bonding of serum stability assessment of the humanized antibody rcNT4X_SA and rcNT4X_S7A to the amyloid peptide Anaspec Aβ4-42. Figure 17 shows the amount of protection provided to primary embryonic rat neurons in vitro by humanized rcNT4X_SA and rcNT4X_S7A, antibodies against amyloid peptide 4-42-induced cell death. / Qoenn / Lznz / E / YiAi Figure 18 shows the amount of protection provided to primary embryonic rat neurons in vitro by humanized rcNT4X_SA and rcNT4X_S7A, (ΑβΰΙρ3) antibodies from amyloid peptide induced cell death 3-42. Figure 19 shows the amount of protection provided to primary embryonic rat neurons in vitro by humanized rcNT4X_SA and rcNT4X_S7A antibodies against cell death induced by the amyloid peptide Aβ1-42. Figure 20 shows the amount of protection provided to human CNS.4U neurons in vitro by humanized rcNT4X_SA and rcNT4X_S7A, antibodies against cell death induced by the amyloid peptide Aβ4-42. Figure 21 shows the amount of protection provided to human CNS.4U neurons in vitro by humanized rcNT4X_SA and rcNT4X_S7A antibodies against cell death induced by the amyloid peptide AβρE3-42. Figure 22 shows the amount of protection provided to human CNS.4U neurons in vitro by humanized rcNT4X_SA and rcNT4X_S7A antibodies against cell death induced by the amyloid peptide Aβ1-42. Figure 23 shows that neuronal loss in the hippocampus of 6-month-old Tg4-42 mice is rescued by both rcNT4X antibodies. Neuron quantification in the CAI was performed using unbiased stereology. The number of neurons in the hippocampus of six-month-old Tg4-42 mice was shown after passive immunization with rcNT4X_SA and rcNT4X_S7A. Tg4-42 mice immunized with rcNT4X antibodies displayed significantly more neurons than mice injected with IgG1 of the same age. One-way analysis of variance (ANOVA) followed by Bonferroni multiple comparisons; n = 5–6. *p < 0.05; ***p < 0.001; data presented as mean ± SEM Figure 24 shows that rcNT4X_S7A has the highest power to rescue neuronal loss in Tg4-42. A comparison of original NT4X data with rcNT4X_SA and rcNT4X_S7A. T-test between NT4X and rcNT4X_S7A. n = 5-7. *p < 0.05; data presented as mean ± SEM Figure 25 shows no significant difference between the MRCT control IgG1 antibody group compared to the IgG2b and PBS control groups. Data for the IgG2ba and PBS groups were taken from Antonios et al. [6]. Neither t-tests nor ANOVA showed significant differences. Data are presented as mean ± SEM, n = 5–7. Figure 26 shows that passive immunization with rcNT4X_S7A rescues learning deficits in Tg4-42 mice. Tg4-42 mice received weekly injections with the antibody and a control antibody (MRCT control) for 12 weeks. The mice were tested at 6 months of age in the Morris water maze. Spatial reference memory was impaired in Tg4-42 mice treated with MRCT control antibodies, as they showed no preference for the target quadrant in the probe test. In contrast, Tg4-42 mice immunized with the rcNT4X_S7A antibody did not exhibit learning deficits. ***p < 0.001; **p < 0.01; *p < 0.05. n = 8 per group. One-way analysis of variance (ANOVA) followed by Bonferroni multiple comparisons. T target quadrant, L left quadrant, R right quadrant, O opposite quadrant. Data presented as mean ± SEM; m = months. Figures 27A to 27E show the reduction in cortical plaque loads in immunized 5XFAD mice. Plaque load analysis of 5XFAD mice immunized with rcNT4X_SA and rcNT4X_S7A compared to 5XFAD mice injected with IgG1. (FIG. 27A) Immunostaining with an antibody against pan-Aβ showed a reduction in plaque load after immunization with rcNT4X_S7A, but not in the group immunized with rcNT4X_SA. (FIG. 27B) Both groups treated with rcNT4X showed significantly reduced fibrillar Aβ deposits as demonstrated by thioflavin S staining. (FIG. 27C) Immunostaining with an antibody against Aβ-χ showed a reduction in plaque burden after immunization with rcNT4X_S7A, but not in the group immunized with rcNT4X_SA. (FIG.27D) Immunostaining with an antibody against αβ3-χ pyroglutamate revealed a reduced plaque burden with both rcNT4X antibodies, which was only significant for the rcNT4X_S7A antibody and showed a trend for the rcNT4X_SA antibody (FIG. 27E). Immunostaining with an antibody against αβ4-χ revealed a reduced plaque burden with both rcNT4X antibodies. One-way analysis of variance (ANOVA) followed by Dunnett's multiple comparison test against the control group; n = 7-11; ***p < 0.001, **p < 0.01, *p < 0.05. Data presented as mean ± SEM DETAILED DESCRIPTION OF THE INVENTION This invention relates to the discovery that the binding activity of humanized versions of the murine anti-amyloid beta (Aβ) antibody NT4X-167 is significantly enhanced by mutation of certain residues within the variable domains. An anti-Aβ antibody as described herein may comprise a heavy chain variable domain (HV) and a light chain variable domain (LV). The heavy chain variable domain may comprise SEQ ID NO: 2 with four or fewer additional amino acid mutations, e.g., substitutions, deletions, or insertions, in the framework regions. The antibody can specifically bind to N-terminal truncated amyloid peptides, for example pyroglutamate-modified amyloid peptides (pE) (also called AβρE3-χ, AβρΟΙυ3-χ, Aβ(ΰΙρ3)3-χ, and p3-x), such as AβρE3-38, AβρE3-40, AβρE3-14 and AβρE3-42, and non-pyroglutamate-modified amyloid peptides, such as Aβ4-38, Aβ4-40, Aβ4-14 and Aβ4-40. The binding can be determined, for example, using an anti-Aβ antibody described herein in an IgG1 format using standard techniques such as ELISA or Surface Plasmon Resonance, as described below. The VH may comprise the amino acid sequence of SEQ ID NO: 2; or SEQ ID NO: 2 with, independently, one or more, for example, two, three, or four additional amino acid alterations or mutations in the framework regions relative to SEQ ID NO: 2 (for example, single amino acid substitutions, deletions, or insertions), preferably substitutions. The additional amino acid alterations or mutations in the framework regions may be at residues other than 27F, 29L, 63R, and 70V, preferably at residues other than 27F, 29L, 63R, 70V, 52BX1, 52CX6, 53X2, 54X3, 55X4, and 56X5 of SEQ ID NO: 2. The VL domain may have the amino acid sequence of SEQ ID NO: 6; or SEQ ID NO: 6 with, independently, one or more, for example, two, three, or four additional amino acid alterations or mutations in the framework regions relative to SEQ ID NO: 6 (for example, single amino acid substitutions, deletions, or insertions), preferably substitutions. The additional amino acid alterations or mutations in the framework regions may be at residues other than 92X7 of SEQ ID NO: 6. The substitutions may be conservative. For example, an anti-Aβ antibody described herein may comprise a VH domain of SEQ ID NO: 3, 4, or 5, optionally with 1, 2, 3, or 4 amino acid substitutions in the frame regions. An anti-Aβ antibody described herein may comprise a VL domain of SEQ ID NO: 7 or 8, optionally with 1, 2, 3, or 4 amino acid substitutions in the frame regions. A suitable anti-Aβ antibody may comprise (i) the VH domain of SEQ ID NO: 3 and the VL domain of SEQ ID NO: 7, (ii) the VH domain of SEQ ID NO: 4 and the VL domain of SEQ ID NO: 7, (iii) the VH domain of SEQ ID NO: 5 and the VL domain of SEQ ID NO: 7, (iv) the VH domain of SEQ ID NO: 3 and the VL domain of SEQ ID NO: 8, (v) the VH domain of SEQ ID NO: 4 and the VL domain of SEQ ID NO: 8, and / or (vi) the VH domain of SEQ ID NO: 5 and the VL domain of SEQ ID NO: 8. Some preferred anti-Aβ antibodies may comprise the VH domain of SEQ ID NO: 5 and the VL domain of SEQ ID NO: 8. The terms immunoglobulin and antibody can be used interchangeably to refer to any protein comprising an antibody antigen-binding site that has the ability to bind specifically to one or more antigens. An antigen is an entity (e.g., a protein or peptide) to which an immunoglobulin or antibody (or antigen-binding fragment of the antibody itself) specifically binds. Anti-Aβ antibody antigens described herein may include the N-truncated amyloid peptides AβρE3-42 and Aβ4-42. Native antibodies are typically heterotetrameric glycoproteins of approximately 150,000 Daltons, composed of two identical light (L) chains and two identical heavy (H) chains. Each light chain is linked to a heavy chain by a covalent disulfide bond, with varying numbers of disulfide bonds between the heavy chains of different antibody isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Antibodies comprise globular regions of heavy-chain or light-chain polypeptides, called domains. A domain may comprise peptide loops, typically 3 to 4 loops, stabilized, for example, by a β-pleated sheet and / or an intrachain disulfide bond. Domains are generally termed constant or variable, depending on the relative lack of sequence variation within domains of several class members in the case of a constant domain, or the significant variation within domains of several class members in the case of a variable domain. Antibody or polypeptide domains are frequently referred to interchangeably in the technical terminology as antibody regions or polypeptide regions. The constant domains of an antibody light chain can be referred to as light chain constant regions, light chain constant domains, CL regions, or CL domains. The constant domains of an antibody heavy chain can be referred to as heavy chain constant regions, heavy chain constant domains, CH regions, or CH domains. The constant domain of the light chain aligns with the first constant domain of the heavy chain. The heavy chain constant domain comprising the antibody tail region is referred to herein as the Fe domain (crystallizable fragment) or Fe region. The Fe region can interact with Fe receptors on the cell surface and certain complement system proteins, thereby enabling the antibody to activate the immune system. Each polypeptide chain contains three heavy chain constant domains. The variable domains of an antibody light chain may be referred to as light chain variable regions, light chain variable domains, VL regions, or VL domains (the L here stands for light rather than the lambda light chain isotype). The variable domains of an antibody heavy chain may be called heavy chain variable regions, heavy chain variable domains, VH regions, or VH domains. Intact light chains have, for example, two domains (VL and CL), and intact heavy chains have, for example, four or five domains (VH, CH1, CH2, and CH3). The light and heavy chain variable domains include hypervariable regions (HVR or HV), also known as complementarity-determining regions (CDRs), which are hypervariable in sequence and can form structurally defined loops. Antibodies typically comprise six hypervariable regions: three in the heavy chain (H1, H2, H3) and three in the light chain (L1, L2, L3), interspersed among relatively conserved frame regions (FRs). In the antibodies described herein, the amino acid sequences of the variable domains are shown below. CDRs can be readily identified within these sequences using standard techniques (see, for example, Kabat, EA, Wu, TT, Perry, HM, Gottesmann, KS & Foeller, C. (1991). Sequences of Proteins of Immunological Interest, 5th ed., NIH Publication no. 91-3242. US Department of Health and Human Services). In Kabat's nomenclature, VHCDR1 is located at positions 31-35, VHCDR2 is located at positions 50-65, VHCDR3 is located at positions 95-102, VLCDR1 is located at positions 24-34, VLCDR2 is located at positions 50-56, and VLCDR3 is located at positions 89-97. The variable regions of each light / heavy chain pair form the antigen-binding site. The term antigen-binding site refers to a site that specifically binds to (immunoreacts with) an antigen. The antibodies described herein comprise at least one antigen-binding site and preferably two. An antigen-binding site is formed from the heavy and light chain CDRs, aligned by the frame regions, which allow binding to a specific epitope. An antigen-binding region or antigen-binding domain is a region or domain of the antibody that includes an antibody-binding site. The antibodies described herein have at least one antigen-binding site that recognizes the amyloid peptides AβρE3-42 and Aβ4-42. Naturally occurring or recombinantly produced antibody chains can be expressed with a leader sequence that is removed during cell processing to produce a mature chain. Mature chains can also be produced recombinantly, containing a non-natural leader sequence, for example, to enhance secretion or alter the processing of a particular chain of interest. The constant regions of an antibody's heavy and light chains can exhibit phenotypic variation. Antibody light chains are classified as kappa (k) or lambda (λ) based on the amino acid sequence of the light chain constant region and are approximately 230 residues long. An antibody described herein / Qoenn / Lznz / E / YiAi comprises a kappa light chain (the variable domain of the kappa light chain is referred to herein as VK). The heavy chains of antibodies in humans and higher mammals are classified as gamma (γ), mu (μ), alpha (α), delta (δ), or epsilon (ε), are approximately 450–600 residues long, and define the antibody isotype as IgG, IgM, IgA, IgD, and IgE, respectively. There are two IgM subclasses (H and L), three IgA subclasses (IgA1, IgA2, and secretory IgA), and four IgG subclasses (IgG1, IgG2, IgG3, and IgG4). An antibody described herein is preferably an immunoglobulin G (IgG) antibody. An antibody described herein is more preferably an IgG4 antibody or an IgG1 antibody with minimal effector function. The antibodies described herein may comprise heavy chains belonging to any of the immunoglobulin isotypes described herein. The antibodies described herein may comprise sequences from more than one class or isotype. An anti-Aβ antibody described herein may exhibit cytotoxic activity. In this antibody, the constant domain is typically complement-fixing, and the class is typically IgG1. The human isotypes IgG1 and IgG4 are exemplary. An antibody described herein may comprise a fragment of a complete antibody. The term fragment refers to a part or portion of an antibody or antibody chain comprising fewer amino acid residues than an intact or complete antibody or antibody chain, wherein the portion preferably retains at least one, and preferably most or all, of the functions normally associated with that portion when present in an intact antibody. Fragments may be obtained by chemical or enzymatic treatment of an intact or complete antibody or antibody chain. Fragments may also be obtained by recombinant means. The antibody fragments described herein can bind to the antigen or compete with the intact antibody (i.e., the intact antibody from which they were derived) for antigen binding (i.e., specific binding). The antibodies described herein bind to the amyloid peptides AβρE3-42 and Aβ4-42. The binding fragments are produced by recombinant DNA techniques or by enzymatic or chemical cleavage of intact immunoglobulins. The antibodies described herein may exist as binding fragments that include, but are not limited to, Fab, Fab', F(ab')2, chemically bound F(ab')2, monospecific Fab2, bispecific Fab2, trispecific Fab2, monovalent IgG, scFv (single-chain variable fragment), di-scFv (divalent scFv), bispecific diabody, trispecific tribody, scFv-Fc, minibody or sdAb (single-domain antibody), and retain the ability to bind to the amyloid peptides AβρE3-42 and Aβ4-42. / αορηη / ίζηζ / Ε / γίΛΐ An antibody described herein may be part of a bispecific or trispecific antibody. A bispecific antibody is an artificial hybrid antibody having two different heavy / light chain pairs and two different antigen-binding sites; a trispecific antibody is an artificial hybrid antibody having three different light / heavy chain pairs and three different antigen-binding sites. Bispecific and trispecific antibodies may be produced by a variety of methods, including hybridoma fusion or Fab fragment linkage. See, for example, Songsivilai & Lachmann, Clin. Exp. Immunol. 79:315-321 (1990); Kostelny et al., Immunol. 148, 1547-1553 (1992). An exemplary antibody described herein may be a bispecific antibody comprising at least two different antigen-binding sites. Specific binding refers to the situation in which an antibody will not exhibit any significant binding to molecules other than its specific epitope on an antigen. The term is also applicable where, for example, an antigen-binding domain is specific for a particular epitope carried by a number of antigens, in which case the antibody carrying the antigen-binding domain will be able to bind to the various antigens carrying the epitope. The anti-Aβ antibodies described herein, or the nucleic acids encoding such antibodies, are in an isolated state. The antibodies and nucleic acids shall be free or substantially free of material with which they naturally associate, such as other polypeptides or nucleic acids with which they are found in their natural environment or the environment in which they are prepared (e.g., cell culture) when such preparation is performed using recombinant DNA technology in vitro or in vivo. The antibodies and nucleic acids may be formulated with diluents or adjuvants and still be isolated for practical purposes—for example, antibodies will normally be mixed with gelatin or other carriers if used to coat microtiter plates for use in immunoassays, or mixed with pharmaceutically acceptable carriers or diluents when used in diagnosis or therapy. Another aspect of the invention provides a nucleic acid encoding an antibody or a light chain, heavy chain, VH domain, or VL domain thereof, as described herein. A nucleic acid, for example, may encode a variable heavy chain domain (VH domain) comprising SEQ ID NO: 2, such as SEQ ID NO: 3, SEQ ID NO: 4, or SEQ ID NO: 5, and / or a variable light chain domain (VK domain) comprising SEQ ID NO: 6, such as SEQ ID NO: 7 or SEQ ID NO: 8, as described above. Optionally, the encoded VH domain and / or VL domain may have up to four additional amino acid mutations in the frame region. Nucleic acids can include DNA and RNA sequences, where the thymine nucleobases are replaced with uracil. / Qoenn / Lznz / E / YiAi An antibody described herein can be produced by recombinant expression. The nucleic acids described above, which encode variable light and heavy chain regions optionally linked to constant regions, can be inserted into expression vectors. The vectors comprising the nucleic acids encoding the antibodies described herein are themselves an aspect of the invention. The light and heavy chains can be cloned into the same or different expression vectors. The nucleic acids encoding the antibody chains described herein can be operably linked to one or more control sequences in the expression vectors that ensure the expression of the antibody chains.Expression control sequences include, but are not limited to, promoters (e.g., naturally associated or heterologous promoters), signal sequences, enhancer elements, and transcription termination sequences. Preferably, expression control sequences are eukaryotic promoter systems in vectors capable of transforming or transfecting eukaryotic host cells (e.g., COS, CHO, or Exp1293 cells). Such vectors can be incorporated into a suitable host, whereby the host is maintained under conditions suitable for high-level expression of the nucleotide sequences and the collection and purification of antibodies. The aspects of the invention provide a nucleic acid encoding an antibody described herein; a vector, preferably an expression vector, comprising one or more nucleic acids encoding an antibody described herein; and a vector comprising one or more nucleic acids encoding an antibody described herein, operably linked to a promoter. Exemplary expression vectors are pHuK and pHuGl, which, in combination with the nucleic acids described herein, comprise nucleotide sequences encoding the antibodies described herein. Other vectors providing nucleotide sequences encoding the constant regions of the antibody light and heavy chains may also be used. The expression vectors used as described herein are typically replicable in host organisms either as episomes or as an integral part of the host chromosomal DNA. Expression vectors commonly contain selection markers (e.g., ampicillin resistance, hygromycin resistance, tetracycline resistance, kanamycin resistance, or neomycin resistance) to enable the detection of cells transformed with the desired DNA sequences (see, for example, Itakura et al. US4704362). Host cells can be transformed with expression vectors and cultured in conventional nutrient media as appropriate to induce promoters, select transformants, and / or amplify genes encoding the required sequences. / Qoenn / Lznz / E / YiAi A host cell comprising a nucleic acid or vector described above is provided as an aspect of the invention. Another aspect of the invention provides a method for making an antibody described herein. The method comprises expressing, in a host cell culture, a vector described herein to produce said antibody, and recovering the antibody from the cell culture. This method may comprise transferring a vector comprising one or more nucleic acids encoding an antibody or antibody chain, as described above, into a host cell, as described herein, growing the host cell culture under conditions that permit expression of the nucleic acids, and recovering the expressed antibody. Any suitable method known in the art may be employed. Suitable microbial host organisms for cloning and expressing the nucleic acids and vectors described herein include prokaryotic hosts such as Escherichia coli, bacilli such as Bacillus subtilis, and other Enterobacteriaceae such as Salmonella, Serratia, and various Pseudomonas species. Expression vectors can also be made in these prokaryotic hosts, typically containing expression control sequences compatible with the host cell (e.g., an origin of replication). In addition, any number of a variety of well-known promoters will be present, such as the lactose promoter system, a tryptophan (trp) promoter system, a beta-lactamase promoter system, or a lambda phage promoter system.Promoters typically control expression, optionally with an operator sequence, and contain ribosome binding site sequences and the like, to initiate and complete transcription and translation. Vectors for use in prokaryotic cells may also require an origin of replication component. Other microbes, such as yeast, can also be used to express the nucleic acids or vectors described herein. Saccharomyces is a preferred yeast host, with suitable vectors containing expression control sequences (e.g., promoters), an origin of replication, termination sequences, and the like, as desired. Typical promoters include 3-phosphoglycerate kinase and other glycolytic enzymes. Inducible yeast promoters include, but are not limited to, promoters for alcohol dehydrogenase, isocytochrome C, and enzymes responsible for maltose and galactose utilization. In addition to microorganisms, mammalian tissue cell culture can also be used to express the nucleic acids or vectors described herein and produce antibody polypeptides (e.g., polynucleotides encoding antibodies or fragments thereof) (see, e.g., Winnacker, From Genes to Clones, VCH Publishers, NY 1987). A eukaryotic or mammalian cell host comprising a nucleic acid or vector described herein is itself an aspect of the invention. Eukaryotic cells are actually preferred because a number of suitable host cell lines capable of secreting heterologous proteins (e.g., intact antibodies) have been developed in the art, including CHO cell lines, various COS cell lines, HeLa cells, ExpiCHO cells, myeloma cell lines, or transformed B cells or hybridomas.The cells may be human or non-human, for example, non-human mammalian cells. In some preferred modalities, the cells are human cells (Exp1293). The antibodies described herein may be produced in cell lines engineered to produce afucosylated proteins, such as the Potelligent® CHOK1SV cell line (BioWa / Lonza), the engineered GlymaxX® cells (ProBioGen), or the EB66 duck embryonic stem cell line (Valneva). Expression vectors for mammalian cells typically include, but are not limited to, one or more of the following: a signal sequence, one or more marker genes, an enhancer element, a promoter, and necessary processing information sites, such as ribosome binding sites, RNA splicing sites, polyadenylation sites, and transcription terminator sequences.The preferred expression control sequences are promoters derived from immunoglobulin genes, SV40, adenovirus, bovine papillomavirus, cytomegalovirus and the like (see for example Co et al., 1 ImmunoL 148:1149 1992). A vector described herein for use in a eukaryotic host cell may also encode a signal sequence or another polypeptide having a specific cleavage site at the N-terminus of the mature antibody chain or polypeptide. Suitable signal sequences may be heterologous and can be recognized and processed (i.e., cleaved by a signal peptidase) by the host cell. For expression in mammalian cells, mammalian signal sequences and viral secretory guides, such as the herpes simplex gD signal, are available. Alternatively, the antibody-coding sequences described herein may be incorporated into transgenes for introduction into the genome of a transgenic animal and subsequent expression in the milk of the transgenic animal (see, for example, Deboer et al., US5741957, Rosen, US5304489, and Meade et al., US5849992). Suitable transgenes include sequences encoding light and / or heavy chains operabably linked to a promoter and enhancer of a mammary gland-specific gene, such as casein or β-lactoglobulin. The vectors described herein, which contain the polynucleotide sequences of interest (e.g., sequences encoding the heavy and light chains and expression control sequences), can be transferred to the host cell using well-established methods that vary depending on the cell type. For example, calcium chloride transfection is commonly used for prokaryotic cells, while calcium phosphate treatment, electroporation, lipofection, biolistics, or virus-based transfection can be used for other cell hosts. (See, generally, Green and Sambrook, Molecular Cloning: A Laboratory Manual (Coid Spring Harbor Press, 4th ed., 2012). Other methods used to transform mammalian cells include the use of polybrene, protoplast fusion, liposomes, electroporation, and microinjection (see, generally, Sambrook et al., sup).For the production of transgenic animals, transgenes can be microinjected into fertilized oocytes, or they can be incorporated into the genome of embryonic stem cells, and the nuclei of these cells are transferred into enucleated oocytes. When heavy and light chains are cloned into separate expression vectors, the vectors are co-transfected to obtain the expression and assembly of intact antibodies described herein. Once expressed, the complete antibodies, their dimers, individual light and heavy chains, or other immunoglobulin forms described herein may be purified according to standard procedures of the art, including ammonium sulfate precipitation, affinity columns, column chromatography, HPLC purification, gel electrophoresis, and the like (see generally Scopes, Protein Purification (Springer-Verlag, NY, 1982)). Substantially pure antibodies of at least approximately 90 to 95% homogeneity are preferred, and 98 to 99% or greater homogeneity is preferred, for pharmaceutical uses as described herein. Standard protein purification methods known in the art may be employed.The following procedures are exemplary of suitable protein purification procedures: fractionation on immunoaffinity or ion exchange columns, precipitation with ethanol, reversed-phase HPLC, chromatography on silica or on a cation exchange resin such as DEAE, chromatography focusing, SDS-PAGE, precipitation with ammonium sulfate, and gel filtration. The production of the antibodies described herein can be carried out using any suitable technique, including those described herein as well as other techniques known in the art. The antibodies described herein can be produced on a commercial scale using methods well established in the art for large-scale antibody manufacturing. For example, recombinant expression systems such as those described herein can be employed. An antibody described herein may specifically bind to the amyloid peptides AβρE3-42 and Aβ4-42. The antibody may show no specific binding or substantially no specific binding to the amyloid peptide Aβ1-42. An antibody described herein may also exhibit the desirable structural, physical, biophysical, and chemical properties described below, with reference to the examples. / Qoenn / Lznz / E / YiAi The affinity of an antibody described herein is the degree or strength of binding of the antibody to the epitope or antigen. The dissociation constant, Kd, ​​and the affinity constant, Ka, are quantitative measures of affinity. Kd is the ratio of the antibody's dissociation rate (koff), the rate at which it dissociates from its antigen, to the antibody's association rate (kOn), the rate at which it binds to its antigen. The binding of an antibody to its antigen is a reversible process, and the rate of the binding reaction is proportional to the concentrations of the reactants. At equilibrium, the rate of formation of the [antibody][antigen] complex is equal to the rate of dissociation into its components [antibody] + [antigen]. The measurement of reaction rate constants can be used to define an equilibrium or affinity constant, Ka (Ka = 1 / Kd). The lower the value of Kd, the greater the antibody's affinity for its target.Most antibodies have Kd values ​​in the low micromolar (10⁶) to nanomolar (10⁷ to 10⁹) range. High-affinity antibodies are generally considered to be in the low nanomolar (10⁻⁹) range, with very high-affinity antibodies in the picomolar (10⁻¹²) range. An antibody described herein may have a dissociation rate constant (kOn) of at least 2xl02M^s'1, at least 5xl02M^s-1, at least 103M^s'1, or at least 5xl03M^s-1. An antibody described herein may have an antibody dissociation rate (koff) less than 5x10-1s-1, less than 10'1s-1, less than 5x10'2s-1, less than 10'2s-1, or less than 5x10'3s-1 In some embodiments, an antibody described herein binds (e.g., specifically binds) to amyloid peptides AβρE3-42 and Aβ4-42 with an affinity constant or Kade of at least 102M'1, at least 5xl02M'1, at least 103M'1, at least 5xl03M'1, at least 104M'1, at least 5xl04M'1, at least 105M'1, at least 5xl05M'1, at least 106M'1, at least 5x10δM'1, or at least 107M'11. An antibody described herein may have a dissociation constant or Kd of the amyloid peptides AβρE3-42 and Aβ4-42 less than 5x10'2M, less than 10'2M, less than 5x10'3M, less than 10'3M, less than 5x10'4M, less than 10'4M, less than 5x10'5M, less than 10'5M, less than 5x10'6M, less than 10'6M, or less than 5x10'7M, Specific antibody binding means that the antibody exhibits appreciable affinity for a particular antigen or epitope and, generally, does not exhibit significant cross-reactivity. An antibody that does not exhibit significant cross-reactivity is one that will not bind appreciably to an undesirable entity (e.g., an undesirable proteinaceous entity). An antibody specific for a particular epitope, for example, does not have significant cross-reactions with remote epitopes on the same protein or peptide. The specific binding of an antibody described herein can be determined according to any recognized means for determining such binding. An antibody can be bound to Aβ4-42 or AβρE3-42 amyloid peptides with a binding affinity of at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% of the binding affinity of murine NT4X-167 antibody to Aβ4-42 or AβρE3-42 amyloid peptide, as measured by ELISA. Suitable ELISA techniques are well known. For example, the immobilized amyloid peptide can be contacted with the antibody in an IgG1 format and washed one or more times in a 0.1% nonionic detergent such as polysorbate 20 (Tween 20) to remove unbound antibody. The antibody bound to the immobilized peptide can then be detected using any convenient technique, for example, using a secondary antibody bound to a detectable tag, such as HRP. An antibody described herein may be thermally stable, i.e., an antibody described herein may bind to the amyloid peptides AβρE3-42 and Aβ4-42 at temperatures between 30°C and 85°C, specifically up to 75°C. An antibody described herein may have a melting temperature between 50°C and 100°C, specifically between 60 and 80°C, more specifically near 66-67°C. An antibody described herein may have a low propensity for aggregation. Aggregation propensity can be analyzed using standard techniques, such as multi-angle light scattering or dynamic light scattering. An antibody described herein may have a low propensity for non-specific protein-protein interactions and good solubility. An antibody described herein may have a low propensity to aggregate when concentrated. A formulation described herein may comprise an antibody concentrated to 50-200 mg / ml, for example 75-150 mg / ml, preferably 80-120 mg / ml and more preferably 90-110 mg / ml, with a preferred concentration of approximately 100 mg / ml, without forming soluble aggregates in an aqueous solution maintained at a physiological pH, for example, by means of Dulbecco's PBS. An antibody described herein may have a low propensity to aggregate when subjected to repeated freezing and thawing, or prolonged temperatures above normal body temperature. For example, a prolonged temperature is 50°C for 30 days in Dulbecco's PBS. An antibody described herein may have an isoelectric point (pl) between pH 8.6 and pH 9, preferably pH 8.1 to pH 8.7 / QOrnn / L7Π7 / E / YILI An antibody described herein may retain binding capacity to the amyloid peptides AβρE3-42 and Aβ4-42 after incubation at 37°C in mouse, human, and / or cynomolgus primate serum. For example, an antibody described herein may retain binding capacity to AβρE3-42 or Aβ4-42 after incubation in mouse, human, and / or cynomolgus serum for 10 to 50 days, preferably 20–40 days, and more preferably 30 days. An antibody that retains binding capacity may exhibit the same or substantially the same binding capacity at 37°C as that observed in an antibody that was not incubated in serum or that was incubated in a control solution. An anti-Aβ antibody described herein may be aglycosylated. The Fe regions of IgG antibodies carry a highly conserved N-glycosylation site, and glycosylation of the Fe fragment is essential for Fe receptor-mediated activity. The N-glycan carbohydrate portions attached to this site are predominantly fucosylated core diantenary structures of the complex type. In addition, small amounts of these N-glycans also carry sialic acid residues linked to GIcNAc and α-2,6, which bisect each other. An aglycosylated antibody may lack one or more carbohydrate portions due to, for example, a chemical or enzymatic process, the absence or mutation of one or more glycosylation sites, or bacterial expression. An anti-Aβ antibody as described herein can be modified to enhance its antibody-dependent cell-mediated cytotoxicity (ADCC). ADCC is a cell-mediated reaction in which nonspecific cytotoxic cells (e.g., natural killer (NK) cells, neutrophils, and macrophages) recognize the antibody bound to a target cell and subsequently cause lysis of the target cell. Such a cell may be a human cell. It is generally thought that the ADCC activity of antibodies requires the binding of the Fe region of an antibody to an antibody receptor on the surface of an effector cell, such as a killer cell, a natural killer cell, or an activated macrophage.By altering the fucosylation (e.g., by reducing or removing it) of the carbohydrate structure of a humanized antibody (i.e., in the Fe region), the antibody's ADCC activity can be potentiated in vitro, for example, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 100-fold, 500-fold, 600-fold, 700-fold, or 1000-fold, relative to an unmodified humanized antibody. Because of the increased ADCC activity, such modified antibodies can be used at lower doses than their unmodified counterparts and generally have fewer or reduced side effects in patients. An anti-Aβ antibody as described herein can be used in complement-dependent cytotoxicity (CDC). CDC involves the central innate complement system, which acts as an effector of adaptive immunity. The classical CDC pathway is triggered by the binding of antibody molecules to an antigen on a target cell and is initiated by the binding of a Clq protein to the Fe domain of the bound antibody. The resulting complement cascade activates a membrane attack pathway, leading to the formation of a membrane attack complex that induces lysis of the target cell.An antibody as described herein can be modified to enhance its ability to trigger CDC by any method known in the art, such as, but not limited to, designing the protein structure to contain amino acid residue substitutions in the antibody heavy chain constant domains. For an example of a combination of IgG1 amino acid substitutions used to enhance CDC activity, see Moore et al., mAbs, 2(2), 181-189 (2010). The CDC activity of an antibody modified as described herein can be enhanced, for example, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 100-fold, 500-fold, 600-fold, 700-fold, or 1000-fold relative to an unmodified humanized antibody. Anti-Aβ antibodies can also be modified by chemical modification, for example by PEGylation, or by incorporation into a liposome, to improve their pharmaceutical properties, for example by increasing their in vivo half-life. An anti-Aβ antibody described herein may be formulated and / or administered as a pharmaceutical composition comprising the active therapeutic antibody agent and a variety of other pharmaceutically acceptable components (see Remington: The Science and Practice of Pharmacy (22nd ed., Pharmaceutical Press, London, PA (2013))). The preferred form depends on the desired route of administration and therapeutic application. Compositions may also include, depending on the desired formulation, pharmaceutically acceptable, nontoxic carriers or diluents, which are defined as vehicles commonly used to formulate pharmaceutical compositions for human or animal administration. The diluent is selected so as not to affect the biological activity of the combination. Examples of such diluents are distilled water, phosphate-regulated physiological saline, Ringer's solution, dextrose solution, and Hank's solution.In addition, the pharmaceutical composition or formulation may also include other non-toxic, non-therapeutic, non-immunogenic carriers, adjuvants or stabilizers and the like. The pharmaceutical compositions containing an anti-Aβ antibody described herein may also include large, slowly metabolized macromolecules such as proteins, polysaccharides such as chitosan, polylactic acids, polyglycolic acids, and copolymers (such as Sepharose™ functionalized latex, agarose, cellulose, and the like), polymeric amino acids, amino acid copolymers, and lipid aggregates (such as oil droplets or liposomes). Furthermore, these carriers may function as immunostimulatory agents (i.e., adjuvants). / Qoenn / Lznz / E / YiAi For parenteral administration, an antibody or composition described herein may be administered as injectable doses of a solution or suspension of the substance in a physiologically acceptable diluent with a pharmaceutical carrier, which may be a sterile liquid such as water, oil, saline solution, glycerol, or ethanol. In addition, excipients such as wetting agents or emulsifiers, surfactants, pH regulators, and the like may be present in the compositions. Other components of pharmaceutical compositions include those of petroleum, animal, vegetable, or synthetic origin, for example, peanut oil, soybean oil, and mineral oil. In general, glycols such as propylene glycol or polyethylene glycol are preferred liquid carriers, particularly for injectable solutions.Antibodies can be administered in the form of a depot injection or implant preparation, which can be formulated in such a way as to allow sustained release of the active ingredient. The term parenteral as used herein includes subcutaneous, intravenous, intradermal, intramuscular, intraperitoneal, and intrathecal administration of an antibody or composition described herein. An anti-Aβ antibody or composition described herein may also be administered by nasal or gastric methods. Typically, the compositions are prepared as injectables, either as liquid solutions or suspensions; solid forms suitable for solution or suspension in liquid vehicles may also be prepared prior to injection. The preparation may also be emulsified or encapsulated in liposomes or microparticles such as polylactide, polyglycolide, or copolymer for an enhanced adjuvant effect, as discussed above (see Langer, Science 249: 1527 (1990) and Hanes, Advanced Drug Delivery Reviews 28:97 (1997)). The agents of this invention may be administered in the form of a depot injection or implant preparation, which may be formulated to permit sustained or pulsatile release of the active ingredient. Additional formulations suitable for other routes of administration include oral, intranasal, and pulmonary formulations, suppositories, and transdermal applications. Oral formulations include excipients such as pharmaceutical-grade mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, and magnesium carbonate. These compositions take the form of solutions, suspensions, tablets, pills, capsules, sustained-release formulations, or powders and contain 10%–95% of the active ingredient, preferably 25%–70%. Topical application can result in transdermal or intradermal delivery. Topical administration can be facilitated by co-administration of the agent with cholera toxin or its detoxified derivatives or subunits, or other similar bacterial toxins (see Glenn et al., Nature 391, 851 (1998)). Co-administration can be achieved by using the components as a mixture or as linked molecules obtained by chemical crosslinking or expression as a fusion protein. Alternatively, transdermal administration can be achieved using a skin patch or transferosomes (Paul et al., Eur. Immunol. 25:3521 (1995); Cevc et al., Biochem. Biophys. Acta 1368:201-15 (1998)). Preferably, an anti-Aβ antibody described herein or a composition comprising an anti-Aβ antibody described herein can be administered intravenously (IV) or intramuscularly (IM). The compositions may comprise an anti-Aβ antibody described herein, pharmaceutically acceptable carriers as described herein, and other therapeutic agents, in particular prophylactic or therapeutic agents useful for the prevention, management, or treatment of Alzheimer's disease (AD). Such therapeutic agents may include analgesic drugs, anti-inflammatory drugs, antiviral drugs, drugs that relieve fever or elevated body temperature, therapeutic compounds designed to numb pain, for example, mouth rinses or sprays that can numb mouth pain, and therapeutics that enhance cognition, such as memantine, donepezil, galantamine, and rivastigmine. A composition described herein may further comprise compositions for rehydrating a subject, for example, by means of intravenous therapy. The compositions described herein may comprise nucleic acids, i.e., DNA or RNA, encoding an anti-Aβ antibody described herein, and any method of delivering such nucleic acids, with or without any of the other compositional components discussed above. The compositions may also comprise vectors, for example, but not limited to, the expression vectors described herein, which themselves comprise the nucleic acids described herein. The compositions described herein may comprise viral vectors for use as nucleic acid delivery systems in cells. Suitable viral vector nucleic acid delivery systems include retroviral systems, adenoviral vectors, viral vectors of the poxvirus family, including vaccinia virus and avian poxviruses, and viral vectors of the alpha virus genus. A nucleic acid encoding an antibody described herein, or a vector containing the same, may be packaged in liposomes for delivery to an individual or cell, which may be incorporated into the compositions as described. Vectors and nucleic acids encoding an antibody may also be adsorbed onto or associated with particle carriers. The compositions described herein may comprise gene therapy vectors containing nucleotide sequences encoding the antibodies described in the / Qornn / Lznz / E / YiAi patent herein, or naked antibody polypeptide chains according to the invention. The compositions may comprise such vectors or polypeptides in combination with the antibodies described herein, and any other composition components described herein. An antibody described herein may be used in a kit. The term "kit" refers to a combination of reagents and other materials that facilitate sample analysis. In some embodiments, an immunoassay kit described herein includes a suitable antigen, a binding agent comprising a detectable portion, and detection reagents. A system for amplifying the signal produced by the detectable portions may or may not also be included in the kit. In addition, in other embodiments, the kit includes, but is not limited to, components such as a sample collection apparatus, sample tubes, holders, trays, racks, plates, kit user instructions, solutions or other chemical reagents, and samples for use in standardization, normalization, and / or control sampling. The kit may contain at least one antibody described herein. A kit may comprise a composition described herein, in one or more containers, optionally with one or more different prophylactic or therapeutic agents useful for the prevention, management, or treatment of Alzheimer's disease (AD). If the composition containing the components for administration is not formulated for delivery via the alimentary route, such as oral administration, a device capable of delivering the kit components via another route may be included, for example, a syringe. The kit may further include instructions for preventing, treating, managing, or improving AD, as well as side effects and dosage information for the method of administration. The present invention also provides diagnostic kits. The antibodies described herein may be useful for monitoring, diagnosing, or providing a prognosis for the development or progression of Alzheimer's disease (AD), and may be used in a kit suitable for such purposes. An antibody described herein may be used in a diagnostic kit to detect the presence of an N-truncated amyloid peptide, such as AβρE3-42 or Aβ4-42, in a body fluid sample taken from an individual, where the individual may be a human or a mammal, such as a non-human primate or a laboratory animal, including mice, rats, and rabbits. A body fluid sample, such as, but not limited to, blood, serum, or cerebrospinal fluid (CSF), is taken from an individual and tested for the presence of N-truncated amyloid peptides using the antibodies described herein.Measuring amyloid peptide levels in an individual's blood using an antibody described herein can provide information on the individual's susceptibility, risk of occurrence, diagnosis, or prognosis of Alzheimer's disease (AD), or on appropriate administration schedules or dosages of an antibody or composition described herein for treating the individual. Diagnostic methods are generally performed in vitro. One method for detecting the presence of a truncated N-type amyloid peptide in an individual's sample may involve contacting the sample with an anti-Aβ antibody described herein and determining the binding of the antibody to one or more peptides in the sample. A kit that is useful for the diagnosis described above may comprise antibodies described herein coupled to a detectable substance that includes, but is not limited to: various enzymes for use in assays including EIA and ELISA, such as, but not limited to, horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase; prosthetic groups, such as, but not limited to, streptavidin / biotin and avidin / biotin; particles such as latex beads or bacteria, for use in agglutination tests; fluorescent materials, such as, among others, umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, fluorescein dichlorotriazinylamine, dansyl chloride, or phycoerythrin; luminescent materials, such as, but not limited to, luminol; bioluminescent materials, such as, among others, luciferase, luciferin and aequorin;radioactive materials, such as, among others, iodine (131I, 125I, 123I, 121I), carbon (14C), sulfur (35S), tritium (3H), indium (115In, 113In, 112In, 111I), and technetium (99Tc), thallium (201T), gallium (68Ga, 67Ga), palladium (103Pd), molybdenum (99Mo), xenon (133Xe), fluorine (18F), 153Sm, 177Lu, 159Gd, 149Pm, 140La, 175Yb, 166Ho, 90Y, 47Se, 186Re, 188Re, i42Pr,105Rh,97Ru,68Ge,57Co,65Zn,85Sr,32P,153Gd,169Yb,51Cr,54Mn,75Se,113Sn, and117Sn;Positron-emitting metals using various positron emission tomography techniques, non-radioactive magnetic metal ions, and molecules that are radiolabeled or conjugated with specific radioisotopes. A readily measurable, detectable label can be conjugated to an antibody described herein and used in the diagnosis of a disease as described herein. The detectable substance can be coupled or conjugated either directly to an antibody or indirectly, through an intermediary (such as, for example, a linker known in the art) using techniques known in the art. Metal ions that can be conjugated to antibodies for use as a diagnostic are known in the art (see, for example, US474900). Antigen detection using any of the methods or detectable substances described above may yield a positive result for the presence of the amyloid peptide AβρE3-42 or Aβ4-42 using the antibodies described herein in a kit as described herein and may diagnose an individual with AD or provide prognostic information about a person with AD or at risk of developing AD. Such an individual may subsequently require and / or undergo treatment for AD, as described herein. / Qoenn / Lznz / E / YiAi Aspects of the invention relate, among other things, to the treatment of Alzheimer's disease (AD) and other AD-related diseases and disorders, as well as other neurological diseases characterized by soluble amyloids. Aspects of the invention also relate to a method of treating, including prophylactic treatment, AD by administering to an individual in need of treatment an effective quantity of an antibody or composition described herein. An antibody or composition, preferably a pharmaceutical composition (for example, a composition comprising an antibody described herein, a pharmaceutically acceptable excipient, and optionally an additional therapeutic agent) described herein may be used in a method for treating the human or animal body.An antibody or composition, preferably a pharmaceutical composition, described herein may be used in a method for treating the human or animal body, wherein the treatment is therapeutic or prophylactic treatment of AD in an individual. The treatment methods mentioned above may comprise administering the antibody or composition (e.g., a composition comprising an antibody described herein, a pharmaceutically acceptable excipient and optionally an additional therapeutic agent) described herein to an individual under conditions that generate a beneficial therapeutic response in the individual, e.g., for the prevention or treatment of AD. This individual may suffer from AD. The treatment methods described herein can be used in both asymptomatic patients and those currently exhibiting symptoms of AD. An antibody described herein may be administered prophylactically to an individual who does not have AD. An antibody described herein may be administered to an individual who does not have, or does not exhibit symptoms of, AD. An antibody described herein may be administered to an individual who does not have, or appears to have, AD. Individuals eligible for treatment include individuals at risk of or susceptible to AD but who are asymptomatic, individuals suspected of having AD, and individuals currently exhibiting symptoms. The antibodies described herein may be administered prophylactically to the general population without the need for any risk assessment of the individual in question.In some modalities, suitable individuals for treatment as described herein may include individuals with early-onset AD or one or more symptoms thereof, and individuals in whom amyloid peptide is detected in a body fluid sample, such as CSF. The terms treat, treating, or treatment (or grammatically equivalent terms) mean that the severity of the individual's condition is reduced or at least partially improved or lessened and / or that some relief, mitigation, or decrease is achieved in at least one clinical symptom and / or there is an inhibition or delay in the progression of the condition and / or prevention or delay of the onset of a disease or ailment. An antibody described herein that can be used in a treatment method for AD may be an antibody of any sequence and format described herein that specifically binds to the N-truncated amyloid peptides AβρE3-42 and / or Aβ4-42. The antibodies used for the treatment methods as described herein may be fragments of antibodies described herein, for example, antigen-binding fragments. An antibody described herein may be administered to an individual with AD. An antibody described herein may be administered to an individual in need of treatment with a pharmaceutical carrier or pharmaceutical composition, or any composition described herein. Alternatively, the antibody can be administered to an individual by administering a polynucleotide that encodes at least one antibody chain. The polynucleotide is expressed to produce the antibody chain in the patient. Optionally, the polynucleotide encodes both heavy and light antibody chains. The polynucleotide is expressed to produce the heavy and light chains in the individual. An antibody described herein may be used in a method for preventing or treating AD that involves administering to the patient an effective dose of the antibody as described herein. As used herein, an effective amount, effective dose, or sufficient amount (or grammatically equivalent terms) of a therapeutic antibody described herein refers to an amount of antibody or composition described herein that is effective in producing a desired effect, which is optionally a therapeutic effect (i.e., by administering a therapeutically effective amount).For example, an effective amount or an effective dose or a sufficient amount may be an amount such that the severity of the individual's condition, e.g., AD, is reduced or at least partially improved or relieved and / or that some relief, mitigation or decrease is achieved in at least one clinical symptom and / or there is an inhibition or delay in the progression of AD and / or prevention or delay in the onset of AD. The terms patient, individual, or subject include human subjects and other mammals receiving prophylactic or therapeutic treatment with one or more agents (e.g., immunotherapeutic agents or antibodies) described herein. Mammalian subjects include primates, e.g., non-human primates. Mammals also include laboratory animals commonly used in research, such as, but not limited to, rabbits and rodents like rats and mice. / Qoenn / Lznz / E / YiAi A quantity of an antibody or composition described herein suitable for achieving therapeutic or prophylactic treatment is defined as an effective dose, e.g., a therapeutically or prophylactically effective dose. In both prophylactic and therapeutic treatment regimens, reagents may be administered in multiple doses until a sufficient immune response has been achieved. The term immune response or immunological response includes the development of a humoral (antibody-mediated) and / or cellular (antigen-specific T cell-mediated response or response to their secreted products) directed against an antigen in a recipient. Typically, the immune response is monitored, and repeat doses are given if the immune response begins to wane. The effective doses of the compositions described herein for the treatment of the conditions described above vary depending on many different factors, including the means of administration, the target site, the patient's physiological state, whether the patient is human or animal, other medications administered, and whether the treatment is prophylactic or therapeutic. Normally, the patient is a human, but non-human mammals, such as primates, rabbits, rats, and mice, including transgenic mammals, can also be treated. Treatment dosages should be adjusted to optimize safety and efficacy. For passive immunization with an antibody described herein, the dosage ranges from approximately 0.01 to 100 mg / kg, and more generally from 0.1 to 50 mg / kg, of the host's body weight. For example, doses may be at least 1 mg / kg of body weight or at least 10 mg / kg of body weight, or within the range of 1–100 mg / kg. In another example, doses may be at least 0.5 mg / kg of body weight or at least 50 mg / kg of body weight, or within the range of 0.5 to 50 mg / kg, preferably at least 5 mg / kg. In a preferred example, dosages may be approximately 50 mg / kg. The methods described herein may involve administering an antibody to a subject as a single dose, in two doses, or in multiple doses. The antibody dose may be approximately 100 pg / kg to 100 mg / kg of the patient's body weight, approximately 300 pg / kg to 60 mg / kg of the patient's body weight, or approximately 10 mg / kg to 50 mg / kg of the patient's body weight. Subjects may be administered such doses daily, every other day, weekly, or according to any other schedule determined by empirical analysis. Treatment may involve administration in multiple doses over a prolonged period, for example, at least six months. Additional treatment regimens may involve administration once every two weeks, once a month, or once every three to six months.Exemplary dosing regimens include 1-20 mg / kg or 15 mg / kg on consecutive days, 30 mg / kg on alternate days, or 60 mg / kg weekly. / Qoenn / Lznz / E / YiAi The antibody described herein can be administered on multiple occasions. The intervals between individual doses can be weekly, monthly, or annually. Intervals can also be irregular, as indicated by measuring the patient's blood levels of the anti-Aβ antibody. In some methods, the dose is adjusted to achieve a plasma antibody concentration of 1–1000 pg / ml, and in others, 25–300 pg / ml. Alternatively, the antibody described herein can be administered as a sustained-release formulation, in which case less frequent administration is required. The dose and frequency vary depending on the antibody's half-life in the patient. In general, humanized antibodies exhibit a longer half-life than chimeric and non-human antibodies. The dosage and frequency of administration may vary depending on whether the treatment is prophylactic or therapeutic. In prophylactic applications, compositions containing the antibodies described herein, or a cocktail thereof, are administered to a patient who is not yet in the diseased state to enhance the patient's resistance. This amount is defined as an effective prophylactic dose. In this use, the precise amounts again depend on the patient's health status and overall immunity, but generally range from 0.1 to 25 mg per dose, particularly 0.5 to 2.5 mg per dose. A relatively low dose is administered at relatively infrequent intervals over a long period of time. The doses of nucleic acids encoding antibodies described herein range from approximately 10 ng to 1 g, 100 ng to 100 mg, 1 pg to 10 mg, or 30–300 pg of DNA per patient. Doses for infectious viral vectors range from 0 to 100, or more, virions per dose. The antibodies and compositions described herein may be administered for therapeutic and / or prophylactic treatment by parenteral, topical, intravenous, oral, gastric, subcutaneous, intra-arterial, intracranial, intraperitoneal, intranasal, or intramuscular methods, as described herein. Intramuscular injection or intravenous infusion is preferred for antibody administration. Other aspects and modalities described herein provide the aspects and modalities described above with the term comprising replaced by the term consisting of and the aspects and modalities described above with the term comprising replaced by the term consisting essentially of. It should be understood that the request describes all combinations of any of the aspects and modalities described above with each other, unless the context requires otherwise. Similarly, the request describes all combinations of the preferred and / or optional features / Qoenn / Lznz / E / YiAi, either individually or in conjunction with any of the other aspects, unless the context requires otherwise. Modifications to the above modalities, additional modalities, and modifications thereof will be evident to the expert in reading this description and, as such, are within the scope described herein. All documents and sequence database entries mentioned in this specification are incorporated herein by reference in their entirety for all purposes. The antibody residue positions described herein are numbered according to the scheme established in Kabat, EA, Wu, TT, Perry, HM, Gottesmann, KS & Foeller, C. (1991). Sequences of Proteins of Immunological Interest, 5th ed., NIH Publication No. 91-3242. USD Department of Health and Human Services. Where appropriate, the position of a substitution may be described relative to a Kabat-numbered residue that is invariant in immunoglobulin sequences. Alternative antibody numbering schemes are described in Honegger, A and Plückthun, A. (2001). J. Mol. Biol 309, 657-67. and / or where used herein shall be taken as a specific description of each of the two specified features or components with or without the other. For example, A and / or B shall be taken as a specific description of each of (i) A, (ii) B and (iii) A and B if each were set out individually herein. Experimental Materials and methods 1. Preparation of RNA from hybridoma cells. Thomas Bayer supplied frozen pellets of NT4X167 mouse hybridoma cells, which were stored at -80°C, and processed using the Qiagen RNeasy kit to isolate RNA following the manufacturer's protocol. 2. Synthesis of the cDNA of the first strand NT4X-167 RNA (~26 pg) was reverse transcribed to produce cDNA using the GE Life Sciences First Strand cDNA Synthesis Kit following the manufacturer's protocol. This was repeated twice to generate 3 independent NT4X-167 cDNA products (rounds 1, 2, and 3) in order to detect and avoid reverse transcriptase-induced cDNA mutations. / αορηη / ίζηζ / Ε / γίΛΐ 3. Determination of the cDNA sequence NT4X-167 cDNA was amplified by PCR in three separate sections. Immunoglobulin cDNA was amplified by PCR with either kappa light chain primers plus MKC or heavy chain primers plus an MHC mix using the Phusion Flash high-fidelity PCR master mix. The result of each PCR reaction was a single amplification product that was purified using the QIAquick PCR purification kit and sequenced (by GATC Biotech) in both directions using the M13-Forward and M13-Reverse primers to obtain three independent sets of sequence information for each immunoglobulin chain. 4· NT4X-167 VK and VH DNA sequence The consensus DNA sequence of the NT4X-167 VK PCR product was designated NT4X167 VK and the consensus DNA sequence of the NT4X-167 VH PCR product was designated NT4X-167 VH and are shown in SEQ ID Nos 9-12, respectively. Germline analysis of the NT4X-167 sequences shows that the Kappa light chain is a murine MKV4 and the heavy chain is a murine MHV7. 5. Construction of the NT4X-167 chimeric expression vectors The construction of chimeric expression vectors involved cloning the amplified variable regions into IgG / kappa vectors (pHuK and pHuGl) using ligase-dependent cloning (LIC). The vectors (modified pCMV) were digested with BfuAl (BspMl), and then coatings compatible with the 3'-5' exonuclease activity of T4 DNA polymerase (+ dATP) were generated. Antibody sequences were generated by first amplifying the variable region by PCR from NT4X-167 cDNA with primers containing either the 3' end of the leader sequence (the majority of the sequence present in the vector) – forward primer – or the beginning of the constant region (IgGl or kappa) – reverse primer – followed by the beginning of the variable region (in each direction). Complementary coatings were generated on the PCR products by treatment with T4 DNA polymerase + dTTP.The vector and inserts were incubated at room temperature, transformed into chemically competent TOP10 bacteria, and plated on kanamycin plates. Several clones were isolated, and the colonies were screened by PCR using the HCMVi and HuGl LIC Rev primers for VH or HuK LIC Rev for VK. Clones that generated PCR products of the correct size were selected, miniprepared using the QIAGEN kit, and sequenced with the same primers. 6. Generation of chimeric antibodies ExpiFectamine 293 suspension cells grown in ExpiFectamine 293 / Qoenn / Lznz / E / YiAi transfection medium and antibiotics were co-transfected with CNT4X-167 VH.pHuGl and CNT4X-167 VK.pHuK (1 pg of DNA each) using ExpiFectamine 293 Reagent. Cells were grown in 1 mL of growth medium for 5 days. Up to 81 pg / mL of the chimeric antibody NT4X-167 was measured in the conditioned medium by ELISA. 7. Amyloid peptides Amyloid peptides, Aβ1-42, AβρE3-42 and 4-42 were acquired from Peptide Specialty Laboratories (PSL) or California Peptides. 8. Transgenic mice The Tg4-42hom (hereafter referred to as Tg4-42) and 5XFAD transgenic homozygous mouse lines used in this study have been previously described [1,2]. 10. Passive immunization The potential therapeutic effects of reverse-cloned (re) humanized NT4X antibodies (rcNT4X_SA and rcNT4X_S7A) were studied using passive immunization in Tg4-42 and 5XFAD mice. Passive immunization was performed by intraperitoneal injections and compared to a control group using an antibody of the same immunoglobulin class as both rcNT4X antibodies (IgG, MRCT control antibody). Male and female Tg4-42 mice were immunized by injection of the antibodies, 10 mg / kg body weight, diluted in sterile PBS (pH 7.4). Mice received weekly injections starting at three months of age. Each mouse received a total of 12 injections. The behavioral test was performed between the 10th and 11th injections. Animals were euthanized after the last injection. The control group received intraperitoneal injections with the MRCT IgG1 control antibody (10 mg / kg body weight). Animals were euthanized after the last injection at six months of age. Six-week-old female 5XFAD mice received weekly injections of either rcNT4X_SA and rcNT4X_S7A (10 mg / kg body weight, diluted in sterile PBS) or MRCT control (IgG1; 10 mg / kg body weight, diluted in sterile PBS). Each mouse received a total of 12 intraperitoneal injections. The animals were sacrificed after the last injection at 18 weeks of age. The control groups were treated the same as the therapeutic groups. 11. Spatial reference memory of Morris' water maze Spatial reference memory in Tg4-42 mice was assessed using the Morris water maze [3] as previously described [2]. 12. Quantification of the number of neurons using unbiased stereology Stereological analysis was performed as previously described [2,4]. The CAI hippocampal cell layer (Bregma -1.22 to -3.52 mm) was delineated in cresyl violet stained sections and analyzed using a stereology workstation (Olympus BX51 with a motorized sample stage for automatic sampling), Stereolnvestigator 7 (MicroBrightField, Williston, USA) and a 100x oil lens (NA = 1.35). 13. Immunohistochemistry and histology Mouse tissue samples were processed as previously described [5]. The following antibodies were used for plate loading staining: antibody 1-57 (αβ3-χ pyroglutamate, 1:5000, mouse monoclonal [5]), antibody 80C2 (anti-αβ-χ, Synaptic Systems Göttingen, 1:500, mouse monoclonal), polyclonal antibody 24311 (anti-pan-β, 1:500, mouse [2]), and polyclonal antibody 029 (anti-β4-χ, 1:500, guinea pig). Biotinylated secondary anti-rabbit and anti-mouse antibodies (1:200) were purchased from DAKO. Staining was visualized using the ABC method with a Vectastain kit (Vector Laboratories) and diaminobenzidine as the chromogen. Counterstaining was performed with hematoxylin. For DAPI staining, sections were deparaffinized and washed in PBS followed by incubation in 4',6-diamidin-2'-phenylindole (DAPI, 1 pg / ml) for 1 min.For fluorescent staining with thioflavin S, tissue sections were deparaffinized and rehydrated, washed twice with deionized water, treated with 1% (w / v) aqueous thioflavin S, and counterstained in 1% (w / v) aqueous 4',6-diamidin-2-phenylindole. Embedding was performed in aqueous fluorescent mounting medium (DAKO). 14· Quantification of the Aβ care Plaque burden was quantified in 5XFAD-immunized mice. For each animal, three paraffin-embedded sections, at least 40 µm apart, were prepared. Relative plaque burden was assessed in the cortex using an Olympus BX-51 microscope equipped with an Olympus DP-50 camera and ImageJ software (NIH, USA). Representative images were systematically captured at 20x magnification. Using ImageJ, the images were binized to 8-bit black and white images, and a fixed intensity threshold was applied to define DAB staining. Measurements were taken for the percentage area covered by DAB staining, as well as for the number of grains per mm² and the mean grain size. / Qoenn / Lznz / E / YiAi 15. Statistical analysis Differences between groups were tested using one-way analysis of variance (ANOVA) followed by Bonferroni multiple comparisons, ANOVA followed by Dunnett's multiple comparisons, or Student's t-test as indicated. All data are given as mean ± standard error of the mean (SEM) as indicated. All statistics were calculated using GraphPad Prism version 5.04 for Windows (GraphPad Software, San Diego, CA, USA). 16. Approval of the study The animal experiments were approved by the local animal protection authorities (Niedersáchisches Landesamt für Verbraucherschutz und Lebensmittelsicherheit) in accordance with approval number 17 / 2447. The experiments were carried out in accordance with the approved protocols. 17. ELISA Each well of a 96-well MaxiSorp plate (Nunc) was coated with 50 pL of 200 ng / mL aliquots of amyloid peptides 1-42, pE3-42, or 4-42 in PBS and incubated overnight at 4°C. The wells were washed 3x with PBS-T (0.1% Tween20) and blocked with 150 pL of 5% milk in PBS / 0.05% Tween20 per well. The wells were then incubated at room temperature with shaking for 1 hour and washed 3x with PBS-T (0.1% Tween20). Fifty milliliters (pL) of serially diluted primary antibody in 1% milk PBS / 0.05% Tween20 were added to the wells of the assay plate using a series of 3-fold dilutions starting from approximately 100 pg / mL. The incubation and washing steps were then repeated. The anti-human kappa HRP chain (Sigma A7164-lmL) was diluted 4,000-fold in 1% milk PBS / 0.05% Tween20, and 50 pL were added to each well. The incubation and washing steps were repeated, and then 75 pL of K-Blue substrate (Neogen) was added to each well and incubated for 5–10 minutes at room temperature.The reaction was stopped by adding 50 pl of RED STOP solution (Neogen) to each well and the optical density was read at 650 nm. Results Generation of a chimeric version of the NT4X-167 antibody The binding of the amyloid peptides Aβ1-42, AβρE3-42, and 4-42 to the chimeric antibody NT4X-167 was measured by ELISA and compared to the original mouse NT4X-167 antibody. The chimeric antibody NT4X-167 bound to the AβρE3-42 peptide in the ELISA assay but did not bind to the Aβ1-42 or 4-42 peptides (Figure 2), with comparable CEso values ​​to the murine NT4X-167 antibody (Figure 1). To further characterize the binding of mouse and chimeric NT4X-167 antibodies to the amyloid peptides / Qoenn / Lznz / E / YiAi, SPR analysis was performed using the Biacore T200 (GE Healthcare). The chimeric NT4X-167 antibody bound to the peptides AβρE3-42 and 4-42 but did not bind to Aβ1-42, with apparent Kd values ​​comparable to those of the original mouse NT4X-167 antibody. The NT4X-167 sequence was used to design the humanized version of the anti-NT4X-167 antibody. Design of the NT4X-167 humanized antibody variants Human VH and VK cDNA databases Human and mouse immunoglobulin protein sequences from the International Database of Immunogenetics 20099 and version 5 of the Kabat database of protein sequences of immunological interest (last updated 17-Nov-1999)8 were used to compile a database of human immunoglobulin sequences in Kabat alignment. The database contains 10,406 VH sequences and 2,894 VK sequences. Molecular model of NT4X-167 A variable region homology model of the mouse NT4X-167 antibody was calculated. The atomic coordinates of 2DQU_L.pdb and lWEJ_H.pdb were the highest-scoring sequence templates for VL and VH, respectively, as determined by BLAST analysis of the Accelrys antibody pdb structure database, and the atomic coordinates of lYNL_LH.pdb were the highest-scoring overall sequence template (interface). These templates were used to generate 20 initial models; the top-scoring model was refined by modeling each CDR loop with its 5 best loop templates. The final 20 models were used to determine a consensus of residues that were within the 4A of the CDR loops. Selection of the human framework The human VH and VK databases containing the NT4X-167 VH and VK protein sequences were queried using various selection criteria. FW residues within 4A of the CDR residues (Kabat definition) in the mouse NT4X167 antibody structures were identified and designated as 4A Proximity Residues. Humanized and incomplete sequences were excluded from the analysis. The sequence AF062228 was chosen as the human heavy chain donor candidate. This sequence exhibits high sequence identity and similarity and has no somatic germline mutations. AF062228 has eight 4α Proximity Residue changes (Tables 1 and 2). Similarly, the sequence AY942002 was chosen as the human kappa light chain donor candidate (Table 4). AY942004, AF054661, and AF113887 were rejected due to the number of somatic mutations in the frames. AJ698329 was rejected due to a G->Q change in Frame 4. All other observed sequences were very similar, but AY942002 showed the best frame identity and similarity for NT4X_VK. AY942002 has no germline somatic mutations and has three possible proximity residue changes of 4A (Table 4). NT4X-167 RHA and RHB Design Since a suitable human framework has been identified, the synthetic protein and DNA sequence can be designed. The initial design of the humanized version of NT4X-167 is the grafting of CDRs 1, 2, and 3 of NT4X-167 VH into the FW acceptor of AF062228, thereby creating the NT4X-167 RHA variant. The eight proximity residues of 4A are remutated to the equivalent mouse residues, thus creating the NT4X-167 RHB variant, and are mutated one at a time into the following variants: the sequences were assembled in silico and designated NT4X-167 RHA for NT4X-167 RH1. Tables 1 to 3 compare the humanized and murine versions of the NT4X-167 VH protein sequences. All humanized variants were cloned into pMoGl and pMoK vectors so that antibodies could be purified from these constructs for in vivo studies in a number of mouse models (5XFAD and Tg4-42). The final major humanized candidates will be cloned into pHuG4 and pHuK vectors. NT4X-167 RKA and NT4X-167 RKB Design The AY942002 framework was used to design the DNA and protein for the humanized constructs. CDRs 1, 2, and 3 of NT4X-167 VK are shown grafted onto the FW acceptor of AY942002 to generate the initial version of humanized NT4X-167 RKA. There are three non-matching 4A proximity residues in NT4X-167 RKA that were backmutated to the equivalent mouse residue in the NT4X-167 RKB variant. These residues are backmutated one at a time into the following variants: the sequences are assembled in silico and designated NT4X-167 RKA to NT4X-167 RKE (Table 4). Generation of humanized antibodies NT4X-167 The genes for NT4X-167 HA, HB, KA, and KB were synthesized using GenScript. The natural human frame sequences AF062228 and AY942002, for the heavy and light chains, respectively, and the natural mouse CDR sequences were assembled in silico and designated NT4X-167 RHA for NT4X-167 RHJ and NT4X-167 RKA for NT4X-167 RKE. Using proprietary GenScript software algorithms, the sequences for RHA / RHB and RKA / RKB were optimized by silent mutagenesis to use codons preferentially used by human cells and synthesized codons. The RKA / RKB and RHA / RHB constructs were amplified by PCR with expression vector-specific / Qoenn / Lznz / E / YiAi primers + insert (as previously described for chimeric versions) and inserted into pMoK and pMoGl, respectively, in ligase-independent cloning reactions and used to transform TOP10 bacteria.The HA version was subsequently modified by PCR mutagenesis to obtain other humanized variants noted in Table 4. The clones were sequenced and plasmid DNA was prepared using the QIAGEN Plasmid Kit or the Qiagen Plasmid Maxiprep Kit. The expression construct sequences (HA, HB, KA, and KB) are shown in SEQ ID Nos: 13-20. Expression plasmid preparations encoding (humanized or chimeric) VH and VK were used to transfect Exp1293 cells, cultured for 5 to 7 days in serum-free medium, after which the conditioned medium containing the secreted antibody was collected. Antibody expression IgGiK antibody concentrations in Exp1293 cell-conditioned media were measured by ELISA. Most antibodies were produced at good expression levels. Antigen binding by early versions (round 1 vs round 2) of humanized NT4X-167 antibodiesThe data shown in Figure 3 illustrate the binding of the RHA / RHB heavy chains in combination with the RKA / RKB light chain versions of the humanized NT4X-167 antibody to the amyloid peptides Aβ1-42 and AβρE3-42. No difference in binding was observed between the versions containing the RKA or RKB kappa light chain, implying that the introduced reverse mutations in KB are not essential for binding. The RHA version showed no evidence of binding to the amyloid peptides, and the humanized versions containing the RHB versions bound to AβρE3-42 showed no evidence of binding. Considering these data, only the KA light chain was advanced and additional versions of the humanized heavy chain were synthesized using Stratagene's QuikChange Lightning Site-Directed Mutagenesis Kit (Stratagene), which generates NT4X RHC-RHJ versions (Table 1).The results of a binding ELISA using the humanized versions RHC-RHJ against the peptide AβρE3-42 are shown in Figure 5. The humanized versions RHB / RKA and RHB / RKB bound to the peptide AβρE3-42 while the other humanized versions showed no evidence of binding and a third round of humanized variants were synthesized. / QQPnn / l 7Π7 / E / YILI Antigen binding by means of the third and fourth rounds of humanized NT4X167 antibodies A third round of humanized NT4X-167 heavy chain variants was generated, RHK to RHR (Table 2). The RHK-RHR variants were obtained using the Stratagene mutagenesis kit (Kit QuikChange Lightning Site-Directed Mutagenesis). The RHC-RHR heavy chains were combined with the RKA version of the light chain, and binding ELISA was performed using the AβρE3-42 peptide (Figure 6). The humanized versions RHB, RHM, RHN, RHO, and RHR, in combination with RKA, bound to the AβρE3-42 peptide, with RHB / RKA, RHM / RKA, RHN / RKA, RHO / RKA, and RHR / RKA being the most optimal binders. Four of the eight key heavy chain CDR framework residues in RHP (arginine remutated by SDM to valine), RHQ (valine remutated to phenylalanine), RHK (phenylalanine remutated to glycine), and RHL (leucine remutated to isoleucine) showed reduced binding, suggesting that these four residues must be maintained as mouse residues for complete binding.Additional humanized variants were generated by incorporating the four mouse residues represented by RHP, RHQ, RHK, and RHL, while retaining the four key frame residues represented by RHM, RHN, RHO, and RHR as human frame residues to generate the RHS, RHT, RHU, RHV, RHW, RHX, and RHY versions (Table 2). Peptide-binding ELISA using ΑβρE3-42 showed that the RHS to RHY variants in combination with RKA have ΑβρE3-42 PSL binding profiles similar to RHB / RKA (Figure 8). The RHS / RKA variant is preferable in terms of the number of key human CDR frame residues it contains (lower immunogenicity). Antigen binding by humanized antibodies SA, S6A, S7A and S8A NT4X-167 Additional RHS / KA (SA) variants were generated to achieve >85% identity to the human germline. IMGT domain gap analysis was performed on the SA version to identify residues that could be mutated to increase the percentage identity to the human germline. The RKA light chain had 89.6% identity with the human germline sequences IGKV1-39*O1 and IGKJ4*01. However, the RHS heavy chain had 79.4% identity with the human germline sequence IGHV4-4*08. Germline analysis of the RHS sequence identified six residues that could be mutated back to the human germline to generate RHS6, RHS7, and RHS8 versions in combination with RKA. Peptide-binding ELISAs using AβρE3-42 showed that the RHS7RKA (S7A) variant was the optimal humanized candidate and had 84.5% identity with the human germline. / Qornn / Lznz / E / YiAi Antigen binding by means of additional variants generated based on the crystal structure of NT4X-167 antibodies Additional RHS7 variants to increase the percentage of identity to the human germline were generated based on the crystal structure of the mouse NT4X FAB-bound pE3-14 peptide. The crystal structure highlighted F67, Y68, and I39 as potential amino acids that could be changed without affecting peptide binding. Therefore, F67Y, Y68N, I39W, and an additional variant combining F67Y and Y68N were generated (Table 2), and binding to Aβ1-42, AβρE3-42, and 4-42 was investigated. RHS71, which has the F67Y mutation, retained binding properties equivalent to the parental heavy chain variant S7A and was therefore chosen as a potential humanized heavy chain variant in combination with the light chain RKA.Since the affinity of these antibodies was on the nanometer scale, it was desired to investigate whether it was possible to predict which amino acids could be mutated based on the crystal structure and Schrödinger modeling prediction software to increase the affinity of the RHS71 / RKA antibody. Five additional heavy chain variants were generated with the following mutations: S53M, S53H, R100H, L103R, and L103H (Table 4b). In addition, five light chain variants were also generated: RKF (N92W), RKG (N92Y), RKH (N92H), RKI (L94R), and RKJ (L94H). The sequences of the light chain variants are shown in Table 4. The binding of these additional humanized variants to Aβ1-42, AβρE3-42, and 4-42 was investigated by ELISA (Figure 9) and Biacore. Among all the variants tested, RHS71 (containing the F67Y mutation in the heavy chain) in combination with RKH (containing the N92H mutation) showed a double improvement in Biacore. Thermal stability of the humanized antibody candidate BA, SA, TA, UA, VA, WA, YA at high temperatures The aim of this experiment was to test the thermal stability of humanized antibodies when subjected to higher temperatures, ranging from 30° to 85°C for 10 minutes. The antibodies were then cooled to 4°C and used in an ELISA assay at the CEso concentration of each candidate. All humanized versions appeared stable, retaining their ability to bind to the AβρE3-42 peptide up to 75°C, at which point peptide binding decreased. Determination of the Tm (melting temperature) of the humanized antibody candidate NT4X-167-SA and NT4X-167-S7A To determine the melting temperature of the lead antibody candidates NT4X-167-SA and NT4X-167-S7A, the antibodies were tested in a heat shift assay. Samples were incubated with a fluorescent dye (Sypro Orange) for 71 cycles at a 1°C increment per cycle in a qPCR thermocycler. The Tm for the humanized / Qoenn / Lznz / E / YiAi antibodies was calculated to be 66–67°C. Aggregation of the humanized antibody candidate NT4X-167-SA and NT4X-167-S7A Samples were injected at 0.4 mL / min into a size exclusion column on an HPLC system and analyzed by multi-angle light scattering to determine absolute molar masses and verify aggregation (see Figure 10). The profile shows no signs of aggregation, with an average molecular weight of approximately 133.98 kDa for NT4X-167_SA and 129.92 kDa for NT4X-167_S7A, which is within the expected range for an IgG monomer in this assay setup. The antibody is monodisperse (MW weight / MW number < 1.05). Mass recovery is 100% (calculated mass over injected mass), indicating good protein recovery and that the sample does not appear to stick to the column or contain insoluble aggregates, which would be retained by the column guard. In particular, the data suggest that there are no aggregation concerns for the humanized antibodies NT4X-167_SA and NT4X167_S7A. Non-specific protein-protein interactions (CIOs) Cross-interaction chromatography using bulk purified human polyclonal IgG is a technique for monitoring nonspecific protein-protein interactions and can be used to discriminate between soluble and insoluble antibodies (Section 8.19). A high retention index (k') indicates a propensity for self-interaction and low solubility. The humanized antibodies NT4X-167 RHS / RKA, RHB / RKA, and RHS7 / RKA (cloned as MoGlK) show a retention index below 0.2, indicating a low propensity for nonspecific interactions and good solubility (Figure 11). Solubility of humanized candidate antibodies NT4X-167 RHS / RKA vs RHS7 / RKA The humanized antibodies NT4X-167 RHS / RKA (SA) and RHS7 / RKA (S7A) were concentrated using solvent absorption concentrators (MWCO 7500 kDa) and the concentration was measured at timed intervals. The antibody was concentrated to >50 mg / mL without apparent precipitation. Freeze-thaw stress analysis of humanized antibody candidate NT4X-167 RHS / RKA and RHS7 / RKA Samples of the purified candidate antibodies were subjected to 10 cycles of 15 minutes at -80°C followed by thawing for 15 minutes at room temperature. The samples were then analyzed by SEC-MALS to verify aggregation (Figures 12 and 13). The data suggest that freeze-thaw cycles do not induce aggregation in the humanized NT4X-167 antibodies. Heat-induced strain analysis of humanized candidate antibodies NT4X167 RHS / RKA and RHS7 / RKA Samples of the purified candidate antibodies were exposed to a) 4°C, b) 25°C, c) 37°C, and d) 50°C for 30 days. The samples were then analyzed by SEC-MALS to verify aggregation (Figure 14). Overall, the data suggest that there are no aggregation concerns for the humanized NT4X-167 antibodies. Serum stability assessment of humanized antibody candidate NT4X-167 RHS / RKA and RHS7 / RKA Purified samples of the humanized antibodies NT4X-167 RHS / RKA and RHS7 / RKA were incubated in mouse, human, and cynomolgus serum. Antibody binding capacity after incubation was measured by ELISA for binding to the peptides AβρE3-42 and 4-42. The binding of the humanized NT4X-167 antibodies incubated in the three different sera was compared to the binding of the unincubated antibody and the antibody incubated in PBS. The ELISA assay showed that the binding of the serum-incubated antibody to the peptides AβρE3-42 and 4-42 is very similar to the binding of the unincubated PBS antibody and the unincubated antibody. Therefore, the humanized antibodies NT4X-167 RHS / RKA and RHS7 / RKA have retained their binding capacity after being incubated in mouse, human and cynomolgus serum for 30 days. Humanized NT4X-167 was shown to bind to amyloid peptides 4-42 and AβρE3-42, but not to Aβ1-42. The humanized antibody also showed protection against neuronal cell death in rat and human neurons. The antibody was designed and expressed as a fully humanized antibody without significant loss of binding potency. Experiments with chimeric antibodies, consisting of murine variable regions within human constant regions, showed similar or improved binding potency in ELISA or kinetic studies using Biacore, compared to that of the murine antibody (Figures 1 and 2). Initial experiments showed that fully humanized NT4X-167, i.e., without the frame mutations to introduce murine 4A proximity residues, did not bind to the AβρE3-42 peptide or the chimeric positive control antibody, but versions with the full set of mutations bound on par with the chimeric positive control. This reduction in binding was isolated to the fully humanized heavy chain. However, it was unexpectedly discovered that the introduction of specific reverse mutations allowed the generation of two leading antibody candidates, NT4X RHS / RKA (SA) and NT4X RHS7 / RKA (S7A). These leading candidates have also been cloned into the HuGlK and HuG4K vectors, as well as the initial MoGl vectors. Both candidates exhibited excellent binding activity, expression, thermostability, affinity, and functionality. In vitro cell assays Neuronal protection by humanized NT4X SA and NT4X S7A antibodies in primary rat vs human cortical cultures The humanized antibodies NT4X_SA and NT4X_S7A were found to maintain the properties of the original mouse NT4X antibody in protecting rat neurons from cell death induced by the N-truncated amyloid peptides (4-42 and pyroGul3-42; Figures 17 and 18) but not against the full-length amyloid peptide Aβ1-42 (Figure 19). All three antibodies are fairly equipotent against the 4-42 peptide, but mouse NT4X is slightly more potent against pyro3-42 than SA or S7A, with S7A being slightly more potent than SA. The humanized antibodies NT4X_SA and NT4X_S7A were found to maintain the properties of the original mouse NT4X antibody in protecting against cell death induced by the N-truncated amyloid peptides (4-42 and pyroGul3-42; Figures 20 and 21) but not against the full-length amyloid peptide Aβ1-42 in human neurons (Figure 22). All three humanized antibodies are more potent than the original mouse NT4X antibody in protecting against cell death with peptide 4-42; however, the humanized antibody N92H is more potent against pyro3-42. None of the NT4X antibodies or their humanized versions protect human neurons from death induced by the amyloid peptide Aβ1-42. The results are largely consistent with those obtained in rat neurons, with some increased potency against pyro3-42 in human neurons. In vivo testing in trans-enic mouse models Alzheimer's therapy with rcNT4X SA v rcNT4X S7A in 5XFAD v Tq4-42 mouse models Tg4-42 mice expressing Aβ4-42 were immunized starting at 12 weeks of age for 12 weeks. rcNT4X_SA and rcNT4X_S7A were shown to rescue the loss of CAI neurons in the hippocampus of Tg4-42, with a greater therapeutic effect for rcNT4X_S7A. rcNT4X_S7A was further tested in the Morris water maze test for spatial reference memory performance. The spatial reference memory deficits in Tg4-42 at six months of age were completely rescued. 5XFAD mice were immunized starting at six weeks of age for [Qoenn / Lznz / E / YiAi] weeks. The effect on plaque burden was analyzed in the cortex. rcNT4X SA reduced plaques stained with thioflavin and specific N-terminal antibodies against Αβ3-X and Αβ4-X pyroglutamate. No effect was observed with antibodies against pan-Αβ and Αβί-X. ​​In contrast to mice immunized with rcNT4X_S7A, which showed a significant reduction in plaque with all staining assays, plaques stained with thioflavin or with antibodies recognizing Αβί-X, Αβ3-X pyroglutamate, Αβ4-X, and pan-Αβ were significantly reduced. rcNT4X SA and rcNT4X S7A rescue neuronal loss and memory impairment in Tq4-42 mice The loss of anterior chamber innate (ACI) neurons in the hippocampus of Tg4-42 mice is significant by four months of age [6]. Therefore, passive immunization treatment was initiated at three months of age and continued for 12 weeks. Tg4-42 mice immunized with rcNT4X_SA and rcNT4X_S7A displayed significantly more neurons compared to the IgG1 control group. The level of significance was highest in the rcNT4X_S7A group (Figure 23). Immunization of Tg4-42 mice with rcNT4X_S7A was significantly more potent compared to NT4X (Figure 24). There was no difference in the number of neurons between NT4X and the original rcNT4X_SA. Data from immunization with rcNT4X_SA and rcNT4X_S7A are plotted against immunization with the original murine NT4X antibodies. The control groups injected with IgG1, IgG2b, and PBS did not differ significantly (Figure 25). IgG2ba and PBS data taken from Antonios et al. [6].Passive immunization with rcNT4X_S7A completely rescued spatial reference memory deficits in Tg4-42 mice tested by Morris Water Maze (Figure 26). rcNT4X SA vs rcNT4X S7A bottom plate charging in 5XFAD mice 5XFAD mice were treated between six and 18 weeks of age. Passive immunization with both rcNT4X antibodies reduced plaque load for different Aβ species compared to an isotype control IgG1 antibody. rcNT4X significantly reduced plaques stained against pyroglutamate Aβ3-χ, Aβ4-χ, and thioflavin. No effect was detected on Aβί-χ and pan-Aβ positive plaques. The plaque-reducing effect of rcNT4X_S7A was significantly altered, as plaques positive for pyroglutamate Aβ3-χ, Aβ4-χ, and thioflavin, but also positive for Aβί-χ and pan-Aβ, were reduced (Figs. 27A to 27E). Tg4-42 mice developed severe loss of hippocampal neurons and spatial reference memory deficits [2,6]. The Tg4-42 model represents the first mouse model that exclusively expresses N-truncated β4-42. At six months of age, this model exhibits significant spatial reference memory loss, assessed by the Morris water maze test, and massive degenerated CAI neurons in the hippocampus of Tg442 mice, which can be rescued by passive immunization with the NT4X antibody [6]. In the present study, a novel humanized version of the NT4X antibody cloned into a murine IgG1 core segment was used. Passive immunization with rcNT4X_S7A, initiated at three months of age for 12 weeks, also rescued spatial reference memory deficits in Tg442 mice.Furthermore, the number of CAI neurons in the hippocampus was significantly rescued compared to the Tg4-42 group of animals treated with IgG1. Interestingly, when comparing the treatment effect between NT4X, rcNT4X_SA, and rcNT4X_S7A, the Tg4-42 mice exposed to rcNT4X_S7A showed a significantly higher effect compared to NT4X. Therefore, rcNT4X_S7A is assumed to have the highest potency among the different NT4X variants. Sequences QVQLQESGPG LVKPSETLSL TCTVSGGSIS SYGIHWIRQP PGKGLEWIGV MWSGGITDFY AAFISRVTIS VDTSKNQFSL KLSSVTAADT AVYYCARGSR YALDYWGQGT LVTVSS SEQ ID NO: 1 RHA Sequence QVQLQESGPG LVKPSETLSL TCTVSGFSLS SYGIHWIRQP PGKGLEWIGV MWSGGITXiX6Y X2X3X4X5SRVTIS RDTSKNQVSL KLSSVTAADT AVYYCARGSR YALDYWGQGT LVTVSS SEQ ID NO: 2: Secuencia de RHA con 27F, 29L, 63R y 70V y 52BX1, 53X2, 54X3, 55X4, 56X5y 52CX6, en donde Xi es D o N, X2es A, N, o P, X3es A o S, X4es F o L, X5es I o K, y Xe es F o Y. QVQLQESGPG LVKPSETLSL TCTVSGFSLS SYGIHWIRQP PGKGLEWIGV MWSGGITDFY AAFISRVTIS RDTSKNQVSL KLSSVTAADT AVYYCARGSR YALDYWGQGT LVTVSS SEQ ID NO: 3 Secuencia de RHS QVQLQESGPG LVKPSETLSL TCTVSGFSLS SYGIHWIRQP PGKGLEWIGV MWSGGITNFY PSLKSRVTIS RDTSKNQVSL KLSSVTAADT AVYYCARGSR YALDYWGQGT LVTVSS SEQ ID NO: 4 Secuencia de RHS7 QVQLQESGPG LVKPSETLSL TCTVSGFSLS SYGIHWIRQP PGKGLEWIGV MWSGGITNYY PSLKSRVTIS RDTSKNQVSL KLSSVTAADT AVYYCARGSR YALDYWGQGT LVTVSS SEQ ID NO: 5 Secuencia de RHS71 DIQMTQSPSS LSASVGDRVT ITCRASQDIS NYLNWYQQKP GKAPKLLIYY TSRLHSGVPS RFSGSGSGTD FTLTISSLQP EDFATYYCQQ GX7TLPPTFGG GTKLEIK SEQ ID NO: 6 pieces of RKA with 92X7, including X7 and N, H, Y or W DIQMTQSPSS LSASVGDRVT ITCRASQDIS NYLNWYQQKP GKAPKLLIYY TSRLHSGVPS RFSGSGSGTD FTLTISSLQP EDFATYYCQQ GNTLPPTFGG GTKLEIK SEQ ID NO: 7 Secuencia de RKA DIQMTQSPSS LSASVGDRVT ITCRASQDIS NYLNWYQQKP GKAPKLLIYY TSRLHSGVPS RFSGSGSGTD FTLTISSLQP EDFATYYCQQ GHTLPPTFGG GTKLEIK SEQ ID NO: 8 Secuencia de RKH GATATCCAGATGACACAGACTACATCCTCCCTGTCTGCCTCTCTGGGAGACAGAGTCACCATCAGTTGCAGGGCAAGTCAGGACATTAGCAATTATTTAAACTGGTATCAGCAGAAACCA θ' UJ / Qoenn / Lznz / E / YiAi CAGGGTGCAGCTGCAGGAGAGCGGACCCGGACTGGTGAAGCCCTCCGAGACCCTGAGCCTGACCTGSACCGTGAGCGGAGGCAGCATCAGCAGCTACGGCA / QQmn / l 7Π7 / Ε / ΥΙΛΙ / QQmn / l 7Π7 / Ε / ΥΙΛΙ CAGGTGCAGCTGCAGGAAAGCGGACCCGGCCTGGTGAAGCCTAGCGAGACCCTGAGCCTGACCTGCACCGTGAGCGGCTTCAGCCTGACCAGCTACGGCA / QQmn / l 7Π7 / Ε / ΥΙΛΙ GACATCCAGATGACCCAAAGCCCTAGCAGCCTGAGCGCCAGCGTGGGAGACAGGGTGACCATCACCTGCAGGGCCAGCCAGGACATCAGCAACTACCTGA / QQrnn / l 7Π7 / Ε / ΥΙΛΙ GACATCCAGATGACCCAGAGCCCTAGCAGCCTGAGCGCTAGCGTGGGCGACAGGGTGACCATCACCTGCAGGGCCAGCCAGGACATCAGCAACTACCTGA Μ Γτΐ ΓτΊ ΓτΊ Μ Μ muí en cq en en w en en fr en en en en WWK Μ K Μ W δ δ1δ δ1δ1δ1δ1§§§§§§§ En En en en en en en en Yes SS Yes Yes Yes Yes Μ Η Μ Μ Η Μ H á1tí1á1á1 S Η Η Η Η Η H >. (Νααααααααααααακακκ» NT4X-167 Stopped Chain Humanization Strategy CM o and σι WCQWCOCQCOCQWWWWWWWWCQWWWW to Ο<ωωω[χ1[χ1[χ1[χ1[χ1[1ΰ[χ1[χ1Γι3[χ|[χ1[χ1[χ1[χ1[χ1[χ1[χ1[χ1[χ1[χ1[χ1[χ1]. ωωωωωωωωωωωωωωωωωωωωω cq σ π· π. η. η. η. η. η. η. η. η, η. η. η. η. η. η. η. η. η. η. η. «η oStSSSSSSSKStíSsSSSSSSS Γ** otCoíococococococoKcoWCootcococococoíoco ίο Οίωωωωωωωωωωωωωωωωωωωω m ^qQqqqqqqqQQQDQQqq S &&&&&&&&&&&&&&&&&&&&& (SEQ ID NO: 45), NT4X*RHS6 (SEQ ID NO: 46), NT4X+RHS7 (SEQ ID NO: 47), NT4X*RHS8 (SEQ ID NO: 48), NT4X*RHS71(F67Y) (SEQ ID NO: 49), NT4X*RHS72(Y68N) (SEQ ID NO: 50), NT4X*RHS73(F67Y / Y68N) (SEQ ID NO: 51), NT4X*RHS74(I39W) (SEQ ID NO: 52), NT4X*RHS81(S53M) (SEQ ID NO: 53), NT4X*RHS82(S53H) (SEQ ID NO: 54), NT4X*RHS83(R100H) (SEQ ID NO: 55), NT4X*RHS84(L103R) (SEQ ID NO: 56), y NT4X*RHS85(L103H) (SEQ ID NO: 57). CUADRO 2 Human Development Strategy NT4X-167 - Versions of the academic school that incrementan % of the identity of the human being Overall % of HuID (excluding J region) 79.4 O 00 84.5 85.6 LO LO 00 85.6 86.6 85.6 Changes AA τ-1 LO LO i—1 i—1 ΓΧΙ τ—1 changed to HuID Residues S4069 & P269 S40; N66 & PSLK (69-72) F67Y 00 > F67Y & Y68N I39W mo (Residuos) H40 D66&AAFI (69-72) H40; D66 & AAFI (69-72) F67 00 LO F67 & Y68 139 Sequence of NT4X (VH) RHS RHS6 RHS7 RHS8 RHS71 RHS72 RHS73 RHS74 O < 3 U / Qoenn / Lznz / E / YiAi Humanization of heavy chain NT4X-167 versions RHS % of Human Identity a (Ti (oj ! « I 3 8 á fe OI O tú ΟΊ Qj co µ r- c & EEEEI δ o< K δ δ δ δ µ & 00 O >4 <Λ H 00 fd (o Q DFATYYYYYY DFATYDFI DFATYYDFI 1 41 M Cb § co É co s 0 W 0 0 s 0 | g 0 g 0 WE 0 WE 0 ESSLQPE 00 H sg [ΙΙΑ [ΙΙΑ 1 i SGID SGID aiss SGID 0I3S co SGID OSGID SGID r- co φ 0 8 co 0 8 co 0 8 co 0 8 co 0 8 co 0 8 co 0 8 0 0 0 0 8 0 8 0 8 0 0 8 0 8 0 8 0 8 0 0 8 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 e co § E 0 E 0 E 0 E 0 § 0 E 0 E 0 E 0 E 0 0 5 67890 SRU-IS 9 SRLHS ii SRLHS SRLHS SRLHS iii Οί ps J ' 8 g 1 G í mqi PGGAP PGGAP i PGKAP PGKAP i Λ ES 8 no >¡ (Λ S_ ii ¡ i 8 1 1 3 coff 8β TCRASQDIS SKÍXWDI TCRASQPIS SICOSWI sicCswm TCRASQDIS TCRASQDIS 8 g CO cá C Θ iiiii I iiim 0 sg 0 0 0 0 0 0 0 0 S 3 «-4 o co c 0 co 0 m 0 0 0 0 0 & 8 3 δ ass 0 & I 0 DigyiTQSE DIQMTQSE DiaVirQSE DIQMTQSE 0 |RKJ(L94H) with «r 0 &< ​​2 2 σ> 5 2 2 2 2 «r H 2 2 2 2 2 Q O $ co LO CO 4—» í2 Tu Ό O 3 g 'co <u oí fü ω (D c g Ό ZJ Ό íU 4-J O 4-J GJ a ω o 73 C V) ω σ <u c c GJ S (Λ íU O Ό íü 'c M— ω GJ O a 'Q íü j_ Ό íü E GJ Ό en GJ C g Ό ω 4—» ZJ E en ω c ro y 73 c (Λ £ Έ O) GJ C c GJ UJ en en G LO Ó Q θ' UJ en Oí X έ co LO Ó Q <y UJ en CM o o CM d07 X < o i—1 ó Q θ' UJ en 7Γ GJ 73 en g ej c GJ ej GJ ω ω (ü ro GJ c 76 fe s ó z Q θ' UJ LO 07 en LO Ó Q <y UJ en UJ fe CM O Ó Z Q θ' UJ en Q c¿ X fe «s i—1 eO Ó Z Q θ' UJ en c¿ $ ó Q O UJ en X σ>—1 2 c¿ X faith LO Ó z Q θ' UJ in 2* 07 _1 s at X faith Co eD Ó ZQ θ' UJ in X CM σ> 2 XX c¿ X faith in eO in θ ø' * 2 Q ZJ c ZJ go CM GJ ω ω 7D o LJ O GJ en o íü O LO ó •s»✓ O -J LL¡ c¿ / Qoenn / Lznz / E / YiAi / Qoenn / Lznz / E / YiAi Light Chain Humanization Strategy NT4X-167 Kappa KD (M) ó T- X oj Ó S--- X 0) T” ό ό V“ T” X fc o LO V— ω oo T* XX c CO co o LO 0) --- - ΑβρΕ3-42 CM Ύ​ 0.1 H tí tí tí Ó tí bd U) <9 09 ¿d 19 ·· >4 tí O tí ' 1 1 [tí 1 | •.fe 1 :tí 1 ·.·.' 1 What? fC those.' >1 : · fe CX< CX 09 > · : U' t“i 09 on tí ti ti* 09 E< ti 09 m ti ih ti ¡¿j ti ti ex ti :(X4 ti ti ee ti km ex 09 0 LO ^5 <0 x < 09 ¿ 9 ti ?-' ti x> ti ti 09 cj tíl ti ti ó.< M ti 09 >··< Γλι ti ti ti ti P Ώ £ CX ti ti ti ti ti ti ώ rn ti I—¡ t<5 ti ti 09 M Uj $ «-i ti ti bl o ω ¡Ώ § ti ti UJ ti ti ti í>¡ íü Kl 09 le K< ti ti ti ti 09 R—· 09 0 M GX m ' n UJ ti 0 A Η 1 CJ 1 Ei 1 ρ1 1 in 1 ►-1 1 faith 1 Μ κι faith lZ faith CO r> ω Ί fe LO Mi ni ej fe fe κΐ > O xC fe fe O fe L> QVQLQESGPGLVKPSETLSLTCTVSGFSLTSYGIHWIRQPPGífGLEWLGT^WSGGITDFYAAFISgLTISRDTSKNQVSLKMSSVT AADTAVY-GLDY-CARGSRYA-G thy pn 09 ​​É· thyroid M thyroid 09 Kl t-Mq thyroid KM 09 thyroid 19 thyroid Μ thyroid 09 thyroid O l-1 and en ϊ1 E 9 i 1 uJ 1 09 1 kJ 1 ÍX4 1 un «q in >4 ftj tM r.''; > W ί u<: LO Mi fti CJ ω > fel QX KtL¡ tí CX d-í O' * £ ^QLQEXGPGLVZPSKTLSLTCTVSGzSITSYGIHKIRQPPGZGLEWLGV^SGGIT^^ AADTA7YY1CARGSRYA --Y-W-Q--- ¡7^ 09 E'4 to 69 M ix; 09 í Du to EXt Η-Ί to 09 to XI XM U; ti OM and 1-4 !·^ S ti ti ti 09 XI 09 O i -Ί CX 09 O ΐ·:·ΐ tSv <9 >4 in Q > 1--I AND< 1 y ; 1 r-ι 1 XI l 09 1 XI 1 £-4 1 in 09 ú.i ιΐιΐ i ί ai íR κΐ a< cj tb :>-< with QX Stlq tí.fe O* << &SA1ATISÜEÍ4ZGI--------VaS£GWGXÁAY¿ÜY¥ Ι�D£8®Ί£Δδθ£ΐσ'82Ι1Ί1Ι9υ§νί>ΐΐΙ399έ»,ώΙ®3Ί3Ή9£3δ3Ι^Ι3Χ931Ί£ί£·£Λ131Ί9Ί1Ή8βΜΊ93585δΊδΑδ E?-i 09 09 tí X., 09 E'H Θ σϊ tí Oí fe (G ha O CQ > fe tí tS tí ¡¿J 0 1X4 P-4 ex 1-4 rrj 09 íx: > h-4 IX ex¡ ¡> tí tí 0-9 H E--4 19 x ex 09 0 ί>·ί . <d>4 09 tí tí EM 1 OI EH l tí 1 en 1 tí 1 E^ 1 en 09 k9j tí tí 09 í 9 tí tí OC tí ex 09 tí ín tí ex ávi tí ití Oí 0 ΐΐ.1 02 tí S t«q 09 C4 O tí tí 09 h-4 tí ”3 tXlq tí tí b'4 O 09 tí tí OOX< tí tí l· tí tí > tí tí 09 l:-4 tí yj 09 tí ííSx U) tí tí 0 > Ι-Ί Η 1 O 1 E-ι 1 μΐ i 01 1 h-1 1 ιμ 1 Μ κΐ á, a; tí tí LO tí 19.1<J Sí? > H tí > O* tí tí Xi CX tí fe O- tí 09 Ώ tí· κ—Ί 09 Ití' e ;* MX 09 Ei tí 09 K4 tí b—3 áq E'H tí s &J C2I tH l·-· 'n 1 L? tí tí tí 0 tí 0 tí tí ex tí b-4 09 CC 09 x¡ > tí > > · μι ro ht ·· tb . rv C.'-J tb í>a Sí tí CO Ω > Pi 1 X 1 Pl 1 rb 1 Pl 1 EH 1 κι m:<n ;> -i n.1 μ; X 0Ί t> i Ί . 1 M tí Mi Cu tí W c* κι; Pi HO d k1 Xj tí 0'9 09 Em tí ti XI tí ti ti 09 bl tí g tí ti ΗΊ tí 09 «íCl tí W tí ti ti ti ex tí m rn ώ tí ti ti 09 tí ti O tí ex 09 U) H-ι ESC U) tí 09 tí > tí ti 1 tí 1 Ε1 I tí I 09 I tí I tí I un Ktq 09 ^4 tí C>q ><1 r / 3 > 0 . Ί i.¿ 15 Kt; til O (9 >4 Q .faith K-Jq you^ you CX IW! •a' ιΐ-χ x: •X) <£1 tí CJ tí c 1 faith S ^5 faith oa faith tz »5 faith e<: bi £· R? If r-ι td F5 ffl -AJ1 FX UJ 5^4 Wq Q EH X¡ £3 tí X PL) Ei dd t?; or; S Hear XJ you x¡ Pm. Table 6 shows the sequences of NT4X_VH (SEQ ID NO: 12), AF062228 (SEQ ID NO: 21), NT4X RHA (SEQ ID NO: 22), NT4X RHB (SEQ ID NO: 23), NT4X: RHK (SEQ ID NO: NO: 32 NT4X RHM (SEQ ID NO: 34), NT4X RHN (SEQ ID NO: 35), NT4X RHO (SEQ ID NO: 36), NT4X RHP (SEQ ID NO: 37), NT4X RHQ (SEQ ID NO: 38), and NT4X RHR (SEQ ID NO: References 1. Oakley, H., et al J Neurosa 2006, 26,10129-10140 2. Bouter, Y. et al Acta Neuropathol2013, 126, 189-205 3. Morris, R. J Neurosci Methods 1984, 11, 47-60. 4. Jawhar, S. et al NeurobiolAging 2012, 33, 196.el29 -196.el40. 5. Wirths, O. et al J Neura! Transm 2010, 117, 85-96 6. Antonios, G., et al Sdentific report 20\6, 5, 17338. doi:10.1038 / srepl7338 7. Wittnam, 1L. et al JBiol Chem 2012, 287, 8154-8162 8. Kabat, EA, et al. Sequences of Proteins of Immunological Interest. 5th ed. NIH National Technical Information Service. (1991) 1-3242. 9. Lefranc, M.-P., et al.. Nucí. Acids Res. (2015) 43 (DI): D413-D422. doi:10.1093 / nar / gkul056 Additional statements of the invention: The following listed statements of the invention are part of the description; 1. An antibody comprising a heavy chain variable domain and a light chain variable domain, wherein a) the variable heavy chain domain (VH domain) comprises SEQ ID NO:2 with four or fewer additional alterations, such as substitutions, in the frame regions, Y b) the variable light chain domain (VK domain) comprises SEQ ID NO:6 with four or fewer additional alterations, such as substitutions, in the frame regions. 2. An antibody according to statement 1 wherein the antibody binds to the amyloid peptides AβρE3-42 and Aβ4-42 and does not bind to the amyloid peptide Aβ1-42. 3. An antibody in accordance with any of the above statements wherein the antibody binds to the amyloid peptide AβρE3-42 with a binding affinity of at least 85% of the binding affinity of murine NT4X-167 antibody to the amyloid peptide AβρE3-42, as measured by ELISA. 4. An antibody in accordance with any of the above statements wherein the VH domain comprises SEQ ID NO: 2. / Qoenn / Lznz / E / YiAi 5. An antibody in accordance with any of the above statements wherein the VL domain comprises SEQ ID NO: 6. 6. An antibody according to any of the above statements wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein XI (Kabat position 52B) is D. 7. An antibody according to any of statements 1 to 5 wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein XI (Kabat position 52B) is N. 8. An antibody according to any of the above statements wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X2 (Kabat position 53) is A. 9. An antibody according to any of statements 1 to 7 wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X2 (Kabat position 53) is P. 10. An antibody according to any of the above statements wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X3 (Kabat position 54) is A. 11. An antibody according to any of statements 1 to 9 wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X3 (Kabat position 54) is S. 12. An antibody according to any of the above statements wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X4 (Kabat position 55) is F. 13. An antibody according to any of statements 1 to 11 wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X4 (Kabat position 55) is L. / Qoenn / Lznz / E / YiAi 14. An antibody according to any of the above statements wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X5 (Kabat position 56) is I. 15. An antibody according to any of statements 1 to 13 wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X5 (Kabat position 56) is K. 16. An antibody according to any of the above statements wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X6 (Kabat position 52C) is F. 17. An antibody according to any of statements 1 to 15 wherein the heavy chain variable domain comprises SEQ ID NO: 2 wherein X6 (Kabat position 52C) is Y. 18. An antibody according to any of statements 1 to 5 wherein the heavy chain variable domain comprises SEQ ID NO: 3 with four or fewer additional substitutions in the frame regions. 19. An antibody according to statement 18 wherein the heavy chain variable domain comprises SEQ ID NO: 3 20. An antibody according to any of statements 1 to 5 wherein the heavy chain variable domain comprises SEQ ID NO: 4 with four or fewer additional alterations, such as substitutions, in the frame regions. 21. An antibody according to statement 20 wherein the heavy chain variable domain comprises SEQ ID NO: 4. 22. An antibody according to any of statements 1 to 5 wherein the heavy chain variable domain comprises SEQ ID NO: 5 with four or fewer additional alterations, such as substitutions, in the frame regions. / Qoenn / Lznz / E / YiAi 23. An antibody according to declaration 22 wherein the heavy chain variable domain (VH domain) comprises SEQ ID NO: 5. 24. An antibody according to any of the above statements wherein the light chain variable domain comprises SEQ ID NO: 6 wherein X7 (Kabat position 92) is N. 25. An antibody according to any of statements 1 to 23 wherein the light chain variable domain comprises SEQ ID NO: 6 wherein X7 (Kabat position 92) isH. 26. An antibody according to any of statements 1 to 23 wherein the light chain variable domain comprises SEQ ID NO: 6 wherein X7 (Kabat position 92) is Y. 27. An antibody according to any of statements 1 to 23 wherein the light chain variable domain comprises SEQ ID NO: 6 wherein X7 (Kabat position 92) is W. 28. An antibody according to any of statements 1 to 25 wherein the light chain variable domain comprises SEQ ID NO: 7 with four or fewer additional alterations, such as substitutions, in the frame regions. 29. An antibody according to statement 28 wherein the light chain variable domain comprises SEQ ID NO: 7. 30. An antibody according to any of statements 1 to 23 and 25 wherein the light chain variable domain comprises SEQ ID NO: 8 with four or fewer additional alterations, such as substitutions, in the frame regions. 31. An antibody according to statement 30 wherein the light chain variable domain comprises SEQ ID NO: 8. 32. An antibody according to any of statements 1 to 5 comprising a VH domain of SEQ ID NO: 3 and a VK domain of SEQ ID NO: 7. / Qoenn / Lznz / E / YiAi 33. An antibody according to any of statements 1 to 5 comprising a VH domain of SEQ ID NO: 4 and a VK domain of SEQ ID NO: 7. 34. An antibody according to any of statements 1 to 5 comprising a VH domain of SEQ ID NO: 5 and a VK domain of SEQ ID NO: 8. 35. An antibody according to any of statements 1 to 5 comprising a VH domain of SEQ ID NO: 5 and a VK domain of SEQ ID NO: 7. 36. A pharmaceutical composition comprising an antibody according to any of the above statements with a pharmaceutically acceptable carrier. 37. A nucleic acid that codes for an antibody from any of statements 1 to 35. 38. A vector comprising the nucleic acid of declaration 37 operably linked to a promoter. 39. A host cell comprising the nucleic acid of statement 37 or vector of statement 38. 40. A method for making an antibody according to any of statements 1 to 35, the method comprising expressing, in a host cell culture, a vector according to statement 36 to produce said antibody; and recovering the antibody from the cell culture. 41. A method for the treatment or prophylaxis of Alzheimer's disease by administering, to an individual in need of such treatment, an effective amount of an antibody in accordance with any of statements 1 to 35 or the pharmaceutical composition of statement 36. 42. An antibody according to any of statements 1 to 35 or the pharmaceutical composition of statement 36, for use in a method for treating the human or animal body. / Qoenn / Lznz / E / YiAi 43. An antibody according to any of statements 1 to 35 or the pharmaceutical composition of statement 36, for use in a method for the treatment of Alzheimer's disease in an individual.< / d>

Claims

1. An antibody comprising a heavy chain variable domain and a light chain variable domain, characterized in that a) the heavy chain variable domain (VH domain) comprises SEQ ID NO:2 with four or fewer additional alterations in the frame regions, and b) the light chain variable domain (VK domain) comprises SEQ ID NO:6 with four or fewer additional alterations in the frame regions.

2. The antibody according to claim 1, further characterized in that the antibody binds to the amyloid peptides AβρE3-42 and Aβ4-42 and does not bind to the amyloid peptide Aβ1-42.

3. The antibody according to any of the preceding claims, further characterized in that the VH domain comprises SEQ ID NO: 2 and the VK domain comprises SEQ ID NO:

6. 4.- The antibody according to any of claims 1 to 3, further characterized in that the heavy chain variable domain comprises SEQ ID NO: 3 with four or fewer additional alterations in the frame regions, optionally wherein the heavy chain variable domain comprises SEQ ID NO:

3.

5. The antibody according to any of claims 1 to 3, further characterized in that the heavy chain variable domain comprises SEQ ID NO: 4 with four or fewer additional alterations in the frame regions, optionally wherein the heavy chain variable domain comprises SEQ ID NO:

4.

6. The antibody according to any of claims 1 to 3, further characterized in that the heavy chain variable domain comprises SEQ ID NO: 5 with four or fewer additional alterations, such as substitutions, in the frame regions, optionally, wherein the heavy chain variable domain comprises SEQ ID NO:

5.

7. The antibody according to any of claims 1 to 6, further characterized in that the light chain variable domain comprises SEQ ID NO: 7 with four or fewer additional alterations, such as substitutions, in the frame regions, optionally, wherein the light chain variable domain comprises SEQ ID NO:

7.

8. The antibody according to any of claims 1 to 6, further characterized in that the light chain variable domain comprises SEQ ID NO: 8 with four or fewer additional alterations, such as substitutions, in the frame regions, optionally wherein the light chain variable domain comprises SEQ ID NO:

8. / Qornn / Lznz / E / YiAi 9. The antibody according to any of claims 1 to 3, further characterized in that it comprises the VH domain of SEQ ID NO: 3 and the VK domain of SEQ ID NO:

7.

10. The antibody according to any of claims 1 to 3, further characterized in that it comprises the VH domain of SEQ ID NO: 4 and the VK domain of SEQ ID NO:

7.

11. The antibody according to any of claims 1 to 3, further characterized in that it comprises the VH domain of SEQ ID NO: 5 and the VK domain of SEQ ID NO:

8.

12. The antibody according to any of claims 1 to 3, further characterized in that it comprises the VH domain of SEQ ID NO: 5 and the VK domain of SEQ ID NO:

7.

13. A pharmaceutical composition comprising an antibody according to any of the preceding claims with a pharmaceutically acceptable carrier. 14.- The antibody according to any of claims 1 to 12 or the pharmaceutical composition according to claim 13, for use in a method for treating the human or animal body.

15. The antibody according to any of claims 1 to 12 or the pharmaceutical composition according to claim 13, for use in a method for the treatment of Alzheimer's disease in an individual.