Anti-amyloid beta peptide antibodies and methods of use thereof

Abstract

The present disclosure relates to anti-amyloid beta peptide antibodies that specifically bind N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta (Ab)), multi-specific binding proteins comprising a pyroGlu3 Abeta (Ab) antigen-binding domain and a blood brain barrier (BBB) antigen-binding domain that specifically binds to a BBB target selected from human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc), and uses (e.g., therapeutic uses) thereof.

Description

ANTI-AMYLOID BETA PEPTIDE ANTIBODIES AND METHODS OF USE THEREOFCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the priority benefit of U. S. Provisional Appl. No. 63 / 759,812, filed February 18, 2025; U. S. Provisional Appl. No. 63 / 801,714, filed May 7, 2025; U. S.Provisional Appl. No. 63 / 819,343, filed June 6, 2025; U. S. Provisional Appl. No. 63 / 884,900, filed September 19, 2025; U. S. Provisional Appl. No. 63 / 911,900, filed November 5, 2025; and U. S. Provisional Appl. No. 63 / 979,679, filed February 10, 2025; each of which is herein incorporated by reference in its entirety.REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

[0002] The content of the electronically submitted sequence listing (Name:4503_038PC07_SequenceListing_ST26.xml; Size: 347,615 bytes; and Date of Creation:February 12, 2026) is herein incorporated by reference in its entirety.FIELD OF THE PRESENT DISCLOSURE

[0003] The present disclosure relates to anti-amyloid beta peptide antibodies that specifically bind N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 A0), multi-specific binding proteins comprising a pyroGlu3 A0 antigen-binding domain and a blood brain barrier (BBB) antigen-binding domain that specifically binds to a BBB target selected from human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc), and uses (e.g., therapeutic uses) thereof.BACKGROUND

[0004] Amyloid beta (A0, Abeta, or A-beta) is formed by the proteolytic cleavage of a larger glycoprotein called amyloid precursor protein (APP). APP is an integral membrane protein expressed in many tissues, but especially in neuron synapses. APP is cleaved by g-secretase to release the A0 peptide, which encompasses peptides ranges from 37-49 amino acid residues. A0 monomers aggregate into several types of higher order structures, such as oligomers, protofibrils, and amyloid fibrils. Amyloid oligomers are soluble and may spread throughout the brain, while amyloid fibrils are larger, insoluble, and can further aggregate to form amyloid deposits or plaques. Amyloid deposits found in brains of human patients include a heterogeneous mixture ofA0 peptides, some of which include N-terminal truncations and further may include N-terminal modifications, such as an N-terminal pyroglutamate residue (pGlu).

[0005] Amyloid plaques found in human patients include a heterogeneous mixture of Ap peptides. PyroGlu3 Abeta (also referred to as pyroGlu3 AP, N3pE Ap, N3pGlu Ap, Ap pE3-42) is a truncated form of Ap peptide and is found only in amyloid plaques. N3pE Ap lacks the first two amino acid residues at the N-terminus of human Ap and has a pyroglutamate which is derived from glutamic acid at the third amino acid position of Ap. Although pyroGlu3 AP is a minor component of the deposited Ap in the brain, studies suggest that pyroGlu3 AP has aggressive aggregation properties and accumulates early in the deposition cascade. Passive immunization by long term chronic administration of antibodies against Ap found in plaques, including pyroGlu3 AP, has been shown to disrupt the Ap aggregates and promote the clearance of plaques in the brain.

[0006] Anti-AP antibodies, including anti-pyroGlu3 AP antibodies, have been previously described. See, e.g., USPN 7,122,374, W02002 / 046237, W02004 / 080419, WO2012 / 021469, WO2012 / 021475, WO2017 / 123517, WO2017 / 160555, WO2018 / 005282, W02018 / 031361, WO2018 / 194951, WO2022 / 150735, WO2022 / 192636, WO2022 / 251048, W02024 / 020470, WO2024 / 107683, W02003 / 070760, WO2014 / 056816, W02015 / 120280, W02014 / 089500, WO2016 / 087944, WO2017 / 211827, WO2019 / 040612, W02021 / 081101, WO2022 / 261026, W02023 / 081194, W02020 / 023530, WO2021 / 186245, WO2023 / 111618, WO2024 / 118665, WO2024 / 206161, WO2023 / 170295, WO2023 / 170291, WO2023 / 170290, W02018 / 011353, and WO2024 / 133925.

[0007] However, such anti-pyroGlu3 AP antibody therapies intended for central nervous system (CNS) disorders may be limited in their effectiveness due to their limited ability to cross the blood brain barrier (BBB). Therefore, there is a need for novel anti-pyroGlu3 Ap therapies having improved delivery across the BBB.SUMMARY OF THE PRESENT DISCLOSURE

[0008] Provided herein is an isolated antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) and comprises the heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and light chain variable region (VL) CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 4, 5, 6, 21, 22, and 23, respectively; SEQ ID NOs: 7, 8, 9, 24, 25,and 26, respectively; SEQ ID NOs: 10, 11, 12, 27, 28, and 29, respectively; SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; SEQ ID NOs: 16, 17, 18, 32, 33, and 34, respectively; SEQ ID NOs: 4, 5, 6, 21, 22, and 23, respectively; SEQ ID NOs: 7, 8, 9, 24, 25, and 26, respectively; SEQ ID NOs: 10, 19, 12, 27, 35, and 29, respectively; SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; or SEQ ID NOs: 16, 20, 18, 36, 33, and 34, respectively.

[0009] Also provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) selected from the group consisting of: (a) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 37 and a VL comprising the amino acid sequence of SEQ ID NO: 38; (b) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 39 and a VL comprising the amino acid sequence of SEQ ID NO: 40; (c) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 41 and a VL comprising the amino acid sequence of SEQ ID NO: 42; (d) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 44; (e) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 45 and a VL comprising the amino acid sequence of SEQ ID NO: 46; (f) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 47 and a VL comprising the amino acid sequence of SEQ ID NO: 48; (g) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 49 and a VL comprising the amino acid sequence of SEQ ID NO: 50; (h) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 51 and a VL comprising the amino acid sequence of SEQ ID NO: 52; (i) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 53; (j) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 54 and a VL comprising the amino acid sequence of SEQ ID NO: 55; and (k) an antibody that binds to the same human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) epitope as the antibody or antigen-binding fragment thereof of any one of (a)-(j).

[0010] Also provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) selected from the group consisting of: (a) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 37 and a VL comprising the amino acid sequence of SEQ ID NO: 38; (b) an antibody comprising a VH comprising the amino acid sequence of SEQ IDNO: 39 and a VL comprising the amino acid sequence of SEQ ID NO: 40; (c) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 41 and a VL comprising the amino acid sequence of SEQ ID NO: 42; (d) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 44; (e) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 45 and a VL comprising the amino acid sequence of SEQ ID NO: 46; (f) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 47 and a VL comprising the amino acid sequence of SEQ ID NO: 48; (g) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 49 and a VL comprising the amino acid sequence of SEQ ID NO: 50; (h) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 51 and a VL comprising the amino acid sequence of SEQ ID NO: 52; (i) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 53; (j) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 54 and a VL comprising the amino acid sequence of SEQ ID NO: 55; and (k) an antibody that competitively inhibits the binding of any one of (a)-(j) to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta).

[0011] In some aspects, the antibody or antigen-binding fragment disclosed herein binds at least 2-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human nonmodified E3-24 Ab polypeptide. In some aspects, the antibody or antigen-binding fragment thereof binds about 2-fold to about 5-fold more strongly to human pyro-Glu3-24 Ab polypeptide than to human non -modified E3-24 Abeta polypeptide.

[0012] In some aspects, the antibody or antigen-binding fragment disclosed herein binds at least 40-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human nonmodified E3-40 Abeta polypeptide or wherein the antibody or antigen-binding fragment thereof binds about 40-fold to about 65-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human non -modified E3-40 Abeta polypeptide.

[0013] In some aspects, the the antibody or antigen-binding fragment promotes phagocytosis of Abeta plaques, optionally wherein 0.1 pg / mL increases phagocytosis of Abeta plaques by about 50% as compared to a control.

[0014] In some aspects, the antibody or antigen-binding fragment promotes uptake of pyroGlu3 Abeta by microglia in the CNS.

[0015] In some aspects, the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) that is at least 90%, at least 95%, at least 97%, at least 99%, or 100% identical to an amino acid sequence selected from any one of SEQ ID NOs: 37, 39, 41, 43, 45, 47, 49, 51, and 54.

[0016] In some aspects, the antibody or antigen-binding fragment comprises a light chain variable region (VL) that is at least 90%, at least 95%, at least 97%, at least 99%, or 100% identical to an amino acid sequence selected from any one of SEQ ID NOs: 38, 40, 42, 44, 46, 48, 50, 52, 53, and 55.

[0017] In some aspects, the antibody or antigen-binding fragment comprises a VH and a VL, wherein the VH and VL comprise amino acid sequences at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to the amino acid sequences of: (a) SEQ ID NOs: 37 and 38, respectively; (b) SEQ ID NOs: 39 and 40, respectively; (c) SEQ ID NOs: 41 and 42, respectively; (d) SEQ ID NOs: 43 and 44, respectively; (e) SEQ ID NOs: 45 and 46, respectively; (f) SEQ ID NOs: 47 and 48, respectively; (g) SEQ ID NOs: 49 and 50, respectively; (h) SEQ ID NOs: 51 and 52, respectively; (i) SEQ ID NOs: 43 and 53, respectively; or (j) SEQ ID NOs: 54 and 55, respectively.

[0018] In some aspects, the antibody or antigen-binding fragment comprises a heavy chain variable region and / or a light chain variable region comprising the amino acid sequences of: (a) SEQ ID NOs: 37 and 38, respectively; (b) SEQ ID NOs: 39 and 40, respectively; (c) SEQ ID NOs: 41 and 42, respectively; (d) SEQ ID NOs: 43 and 44, respectively; (e) SEQ ID NOs: 45 and 46, respectively; (f) SEQ ID NOs: 47 and 48, respectively; (g) SEQ ID NOs: 49 and 50, respectively; (h) SEQ ID NOs: 51 and 52, respectively; (i) SEQ ID NOs: 43 and 53, respectively; or (j) SEQ ID NOs: 54 and 55, respectively.

[0019] In some aspects, the antibody or antigen-binding fragment comprises a heavy chain constant region and a light chain constant region.

[0020] In some aspects, the heavy chain constant region is an isotype selected from the group consisting of human IgGl, IgG2, IgG3, and IgG4 isotypes.

[0021] In some aspects, the antibody or antigen-binding fragment comprises a heavy chain constant region and a light chain constant region, In some aspects, the heavy chain constant region is a human IgGi heavy chain constant region, and the light chain constant region is a human IgGK light chain constant region.

[0022] In some aspects, the antibody or antigen-binding fragment is a monoclonal antibody.

[0023] In some aspects, the antibody or antigen-binding fragment thereof is a murine, chimeric, humanized, or human antibody or antigen-binding fragment thereof.

[0024] In some aspects, the antibody or antigen-binding fragment thereof is a full length antibody

[0025] In some aspects, the antibody or antigen-binding fragment thereof is an antigen binding fragment. In some aspects, the antigen-binding fragment is a Fab, Fab’, F(ab’)2, single chain Fv (scFv), disulfide linked Fv, V-NAR domain, IgNar, intrabody, IgGACH2, minibody, F(ab’)3, tetrabody, triabody, diabody, single-domain antibody, DVD-Ig, Fcab, mAb2, (scFv)2, or scFv-Fc.

[0026] In some aspects, the antibody or antigen-binding fragment thereof is monospecific.

[0027] In some aspects, the antibody or antigen-binding fragment thereof is bispecific.

[0028] In some aspects, the antibody or antigen-binding fragment thereof is multispecific.

[0029] In some aspects, the antibody or antigen-binding fragment thereof further comprises an antigen-binding domain that binds to blood brain barrier (BBB) target.

[0030] Provided herein is a complex comprising the antibody or antigen-binding fragment disclosed herein and an antigen-binding domain that binds to a BBB target.

[0031] In some aspects, the antibody or antigen-binding fragment or complex discloses herein, further comprises a detectable label.

[0032] In some aspects, the BBB target is human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc).

[0033] In some aspects, (a) the BBB target is human TfR and wherein the antibody or antigenbinding fragment or complex comprises a human TfR antigen-binding domain comprising heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and light chain variable region (VL) CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of:SEQ ID NOs: 56, 57, 58, 66, 67, and 68, respectively;SEQ ID NOs: 56, 57, 58, 69, 67, and 68, respectively;SEQ ID NOs: 56, 57, 59, 69, 67, and 68, respectively;SEQ ID NOs: 56, 57, 60, 69, 67, and 68, respectively;SEQ ID NOs: 56, 57, 59, 70, 67, and 68, respectively;SEQ ID NOs: 56, 57, 59, 71, 67, and 68, respectively;SEQ ID NOs: 56, 57, 59, 72, 67, and 68, respectively;SEQ ID NOs: 56, 57, 59, 73, 67, and 68, respectively;SEQ ID NOs: 56, 57, 61, 69, 67, and 68, respectively;SEQ ID NOs: 62, 63, 64, 74, 75, and 76, respectively;SEQ ID NOs: 62, 65, 64, 77, 75, and 76, respectively;SEQ ID NOs: 62, 63, 64, 78, 75, and 76, respectively;SEQ ID NOs: 62, 63, 64, 79, 75, and 76, respectively;SEQ ID NOs: 62, 65, 64, 80, 75, and 76, respectively;SEQ ID NOs: 56, 57, 59, 66, 67, and 68, respectively;SEQ ID NOs:62, 275, 64, 276, 75, and 277, respectively; orSEQ ID NOs:62, 275, 64, 77, 75, and 76, respectively; or(b) the BBB target is human CD98hc and wherein the antibody or antigen-binding fragment or complex comprises a human CD98hc antigen-binding domain comprising heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and light chain variable region (VL) CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of:SEQ ID NOs: 98, 99, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 101, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 102, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 103, 100, 122, 123, and 124, respectively;SEQ ID NOs:98, 104, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 105, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 106, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 107, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 108, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 109, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 110, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 125, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 126, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 127, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 128, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 129, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 130, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 131, 123, and 124, respectively;SEQ ID NOs: 98, 106, 100, 127, 123, and 124, respectively;SEQ ID NOs: 98, 111, 100, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 112, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 113, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 114, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 115, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 116, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 117, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 118, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 119, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 120, 122, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 132, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 133, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 134, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 135, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 136, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 137, 123, and 124, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 138, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 139, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 140, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 141, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 142, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 143, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 144, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 145, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 146, respectively;SEQ ID NOs: 98, 99, 100, 122, 123, and 147, respectively;SEQ ID NOs: 121, 99, 100, 122, 123, and 124, respectively; orSEQ ID NOs: 121, 99, 100, 122, 123, and 147, respectively.

[0034] In some aspects, (a) the BBB target is human TfR and wherein the human TfR antigenbinding domain comprises a VH and a VL, wherein the VH and VL comprise amino acidsequences at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to the amino acid sequences of:SEQ ID NOs: 81 and 82, respectively;SEQ ID NOs: 81 and 83, respectively;SEQ ID NOs: 84 and 83, respectively;SEQ ID NOs: 85 and 83, respectively;SEQ ID NOs: 84 and 86, respectively;SEQ ID NOs: 84 and 87, respectively;SEQ ID NOs: 84 and 88, respectively;SEQ ID NOs: 84 and 89, respectively;SEQ ID NOs: 90 and 83, respectively;SEQ ID NOs: 91 and 92, respectively;SEQ ID NOs: 93 and 94, respectively;SEQ ID NOs: 91 and 95, respectively;SEQ ID NOs: 91 and 96, respectively;SEQ ID NOs: 93 and 97, respectively;SEQ ID NOs: 269 and 270, respectively;SEQ ID NOs: 271 and 272, respectively; orSEQ ID NOs: 273 and 274, respectively; or(b) the BBB target is human CD98hc and wherein the human CD98hc antigen-binding domain comprises a VH and a VL, wherein the VH and VL comprise amino acid sequences at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to the amino acid sequences of:SEQ ID NOs: 148 and 149, respectively;SEQ ID NOs: 150 and 149, respectively;SEQ ID NOs: 151 and 149, respectively;SEQ ID NOs: 152 and 149, respectively;SEQ ID NOs: 153 and 149, respectively;SEQ ID NOs: 154 and 149, respectively;SEQ ID NOs: 155 and 149, respectively;SEQ ID NOs: 156 and 149, respectively;SEQ ID NOs: 157 and 149, respectively;SEQ IDNOs: 158 and 149, respectively; SEQ ID NOs: 159 and 149, respectively; SEQ ID NOs: 148 and 160, respectively; SEQ ID NOs: 148 and 161, respectively; SEQ ID NOs: 148 and 162, respectively; SEQ ID NOs: 148 and 163, respectively; SEQ ID NOs: 148 and 164, respectively; SEQ ID NOs: 148 and 165, respectively; SEQ IDNOs: 148 and 166, respectively; SEQ ID NOs: 155 and 162, respectively; SEQ ID NOs: 167 and 149, respectively; SEQ ID NOs: 168 and 149, respectively; SEQ ID NOs: 169 and 149, respectively; SEQ ID NOs: 170 and 149, respectively; SEQ ID NOs: 171 and 149, respectively; SEQ ID NOs: 172 and 149, respectively; SEQ ID NOs: 173 and 149, respectively; SEQ ID NOs: 174 and 149, respectively; SEQ ID NOs: 175 and 149, respectively; SEQ ID NOs: 176 and 149, respectively; SEQ ID NOs: 148 and 177, respectively; SEQ ID NOs: 148 and 178, respectively; SEQ ID NOs: 148 and 179, respectively; SEQ ID NOs: 148 and 180, respectively; SEQ ID NOs: 148 and 181, respectively; SEQ ID NOs: 148 and 182, respectively; SEQ ID NOs: 148 and 183, respectively; SEQ ID NOs: 148 and 184, respectively; SEQ ID NOs: 148 and 185, respectively; SEQ ID NOs: 148 and 186, respectively; SEQ ID NOs: 148 and 187, respectively; SEQ ID NOs: 148 and 188, respectively;SEQ ID NOs: 148 and 189, respectively;SEQ ID NOs: 148 and 190, respectively;SEQ ID NOs: 148 and 191, respectively;SEQ ID NOs: 148 and 192, respectively;SEQ ID NOs: 193 and 194, respectively; orSEQ ID NOs: 193 and 192, respectively

[0035] In some aspects, the human TfR antigen-binding domain comprises a disulfide staple.

[0036] In some aspects, the human TfR antigen-binding domain comprises the amino acid sequence of SEQ ID NO:202 or SEQ ID NO:203.

[0037] In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises the VH CDR 1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; and the human TfR antigen-binding domain comprises VH CDR1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs: 56, 57, 59, 69, 67, and 68, respectively. In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises the VH CDR 1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; and the human TfR antigen-binding domain comprises VH CDR1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs: 56, 57, 59, 72, 67, and 68, respectively. In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises the VH CDR 1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; and the human TfR antigen-binding domain comprises VH CDR1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs: 56, 57, 59, 73, 67, and 68, respectively.

[0038] In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises a VH and a VL comprising the amino acid sequences of SEQ ID NO:43 and 53 respectively; and the human TfR antigen-binding domain comprises a VH and a VL comprisingthe amino acid sequences of SEQ ID NOs: 84 and 83, respectively. In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises a VH and a VL comprising the amino acid sequences of SEQ ID NO:43 and 53 respectively; and the human TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs: 84 and 88, respectively. In some aspects, the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises a VH and a VL comprising the amino acid sequences of SEQ ID NO:47 and 48 respectively; and the human TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs: 84 and 89, respectively.

[0039] In some aspects, the (i) the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta is a full-length antibody, a single-chain fragment variable (scFv), a VHH, or a Fab, and (ii) the anti-BBB antigen-binding domain is a single-chain fragment variable (scFv).

[0040] In some aspects, the (i) the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta is a full-length antibody, a single-chain fragment variable (scFv), a VHH, or a Fab and (ii) the anti-BBB antigen-binding domain is a VHH.

[0041] In some aspects, the (i) the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta is a full-length antibody, a single-chain fragment variable (scFv), a VHH, or a Fab and (ii) the anti-BBB antigen-binding domain is a Fab.

[0042] In some aspects, the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the anti-BBB antigen-binding domain are connected by a linker. In some aspects, the linker connects the C-terminus of the heavy chain Fc of the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the N-terminus of the anti-BBB antigen-binding domain. In some aspects, the linker comprises GGSGG (SEQ ID NO: 214). In some aspects, the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the anti-BBB antigen-binding domain are directly connected. In some aspects, the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the anti-BBB antigen-binding domain comprises the amino acid sequence of SEQ ID NO:280, SEQ ID NO:281, or SEQ ID NO:282.

[0043] In some aspects, the antibody or antigen-binding fragment or complex is bivalent.

[0044] In some aspects, the antibody or antigen-binding fragment or complex is trivalent, optionally the antibody or antigen-binding fragment or complex comprises (i) one antigenbinding domain that binds to the BBB target and two antigen-binding domains that bind topyroGlu3 Abeta or (ii) two antigen-binding domains that bind to the BBB target and one antigenbinding domain that binds to pyroGlu3 Abeta.

[0045] In some aspects, the antibody or antigen-binding fragment or complex is tetravalent, optionally the antibody or antigen-binding fragment or complex comprises two antigen-binding domains that bind to the BBB target and two antigen-binding domains that bind to pyroGlu3 Abeta.

[0046] In some aspects, the antibody or antigen-binding fragment or complex described herein comprise the amino acid seuqences of (a) SEQ ID NOs: 223, 224, and 225, (b) SEQ ID NOs: 226, 227, and 228; (c) SEQ ID NOs: 229, 230, and 231; (d) SEQ ID NOs: 241, 224, and 225; (e) SEQ ID NOs: 242, 224, and 225; (f) SEQ ID NOs: 243, 224, and 225; (g) SEQ ID NOs: 244, 224, and 225; (h) SEQ ID NOs 245, 224, and 225; (i) SEQ ID NOs: 246, 224, and 225; (j) SEQ ID NOs: 247, 224, and 225; (k) SEQ ID NOs: 248, 224, and 225; (1) SEQ ID NOs: 249, 224, and 225; (m) SEQ ID NOs: 250, 224, and 225; (n) SEQ ID NOs: 251, 230, and 231; (o) SEQ ID NOs: 252, 230, and 231; (p) SEQ ID NOs: 253, 230, and 231; (q) SEQ ID NOs: 254, 230, and 231; (r) SEQ ID NOs: 255, 230, and 231; (s) SEQ ID NOs: 256, 230, and 231; (t) SEQ ID NOs: 257, 230, and 231; (u) SEQ ID NOs: 258, 230, and 231; (v) SEQ ID NOs: 259, 230, and 231; (w) SEQ ID NOs: 260, 230, and 231; (x) SEQ ID NOs: 261, 230, and 231; (y) SEQ ID NOs: 262, 230, and 231; (z) SEQ ID NOs: 263, 230, and 231; (aa) SEQ ID NOs: 264, 230, and 231; (bb) SEQ ID NOs: 265, 230, and 231; SEQ ID NOs: 266, 230, and 231; (cc) SEQ ID NOs: 278, 224, and 225; or (dd) SEQ ID NOs: 279, 230, and 231.

[0047] In some aspects, the antibody or antigen-binding fragment or complex enhances brain uptake of the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroGlu3 Abeta in one or more of the frontal cortex, hippocampus, and thalamus of an animal.

[0048] In some aspects, the antibody or antigen-binding fragment or complex increases the concentration of the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroGlu3 Abeta in the CSF of an animal.

[0049] Provided herein is an isolated polynucleotide comprising a nucleic acid molecule encoding the heavy chain variable region or heavy chain of the antibody or antigen-binding fragment thereof described herein.

[0050] In some aspects, the nucleic acid molecule encodes the VH of any one of SEQ ID NOs: 37, 39, 41, 43, 45, 47, 49, 51, and 54.

[0051] Provided herein is an isolated polynucleotide comprising a nucleic acid molecule encoding the light chain variable region or light chain of the antibody or antigen-binding fragment thereof described herein.

[0052] In some aspects, the nucleic acid molecule encodes the VL of any one of SEQ ID NOs: 38, 40, 42, 44, 46, 48, 50, 52, 53, and 55.

[0053] Provided herein is an isolated polynucleotide comprising a nucleic acid molecule encoding the heavy chain variable region or heavy chain of the antibody or antigen-binding fragment thereof described herein and the light chain variable region or light chain of the antibody or antigen-binding fragment thereof described herein.

[0054] Provided herein is an isolated vector comprising the polynucleotide described herein.

[0055] Provided herein is a host cell comprising (a) the polynucleotide described herein (b) the vector described herein, or (c) a first vector comprising the polynucleotide described herein and a second vector comprising the polynucleotide described herein.

[0056] In some aspects, the host cell is selected from the group consisting of E. coli, Pseudomonas, Bacillus, Streptomyces, yeast, CHO, YB / 20, NS0, PER-C6, HEK-293T, NIH-3T3, HeLa, BHK, Hep G2, SP2 / 0, Rl.l, B-W, L-M, COS 1, COS 7, BSC1, BSC40, BMT10 cell, plant cell, insect cell, and human cell in tissue culture, optionally wherein the host cell is a HEK-293 cell.

[0057] Provided herein is a method of producing an antibody or antigen-binding fragment thereof that binds to human pyroGlu3 Abeta comprising culturing the host cell described herein so that the nucleic acid molecule is expressed and the antibody or antigen-binding fragment thereof is produced, optionally the method further comprises isolating the antibody or antigenbinding fragment thereof from the culture.

[0058] Provided herein is an isolated antibody or antigen-binding fragment thereof that specifically binds to human pyroGlu3 Abeta and is encoded by the polynucleotide described herein, encoded by the vector described herein, or produced by the method described herein.

[0059] Provided herein is a pharmaceutical composition comprising the antibody or antigenbinding fragment thereof or complex described herein and a pharmaceutically acceptable excipient.

[0060] Provided herein is a method of promoting phagocytosis of Abeta plaques comprising contacting a composition comprising Abeta plaques and macrophages and / or microglia with theantibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0061] In some aspects, the contacting is in vitro. In some aspects, the contact is in a subject.

[0062] Provided herein is a method of slowing the progression of neurodegeneration in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein. As used herein “slowing the progression” of neurodegeneration refers to decreasing the speed at which degeneration occurs as compared to a subject who did not receive the administration. In some aspects, the administration provided herein can prevent or delay conversion from mild cognitive impairment (MCI) to Alzheimer’s disease.

[0063] Provided herein is a method of reducing neurodegeneration in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0064] Provided herein is a method of decreasing cognitive decline in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0065] In some aspects, the subject has Alzheimer’s disease.

[0066] Provided herein is a method of treating a neurodegenerative disease in a subject, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0067] Provided herein is a method of preventing a neurodegenerative disease in a subject, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0068] In some aspects, the neurodegenerative disease is Alzheimer’s disease, early Alzheimer’s disease, or mild cognitive impairment (MCI).

[0069] Provided herein is a method of treating Alzheimer’s disease in a subject, the method comprising administering to the subject a therapeutically effective amount of the antibody orantigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0070] Provided herein is a method of preventing Alzheimer’s disease in a subject, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical composition described herein.

[0071] In some aspects, the Alzheimer’s disease is familial Alzheimer’s disease.

[0072] In some aspects, a method provided herein comprises administering to the subject a therapeutically effective amount of an antibody or antigen binding fragment thereof or complex described herein or the pharmaceutical described herein, wherein the BBB target is TfR.

[0073] In some aspects, the administration results in minimal hematological adverse events. In some aspects, the administration results in no adverse or long-term reduction in reticulocyte count in the subject. In some aspects, the administration results in no adverse or long-term reduction in red blood cell count in the subject. In some aspects, the administration results in no adverse or long-term reduction in hematocrit in the subject. In some aspects, the administration results in no adverse or long-term reduction in hemoglobin level in the subject. In some aspects, the administration results in rapid clearance or reduction of Abeta in the brain of the subject. In some aspects, the administration requires less frequent monitoring of Abeta levels or no monitoring of Abeta levels in the brain of the subject as compared to the monitoring required of other anti-Abeta antibody therapies. In some aspects, the subject has not been APOE genotyped prior to the administration. In some aspects, the administration is associated with less ARIA as compared to the ARIA associated with other anti-Abeta antibody therapies.

[0074] In some aspects, the administration is intravenous or subcutaneous.

[0075] In some aspects, a method provided herein further comprises administering an additional active agent. In some aspects, the additional active agent is one or more of a tau-targeting agent, an ApoER2 -targeting agent, an ApoE4-targeting agent, an ApoE2 -targeting agent, a NLRP3 -targeting agent, a GSK3B targeting agent, and a GLP-1 targeting agent.

[0076] Provided herein is an antibody or antigen-binding fragment thereof or complex described herein or the pharmaceutical composition described herein for use in the method described herein or for use in the manufacture of a medicament for use in the method described herein.

[0077] Provided herein is the use of the antibody or antigen-binding fragment thereof or complex described herein or the pharmaceutical composition described herein in the method described herein.BRIEF DESCRIPTION OF THE DRAWINGS

[0078] Figure 1A (FIG. 1A) shows an exemplary 2+1 multi-specific binding protein containing an scFv that binds to a BBB target (e.g., TfR or CD98hc) linked to a C-terminus of the heavy chain (with an Fc “hole” mutation) of a full-length antibody that binds to pyroGlu3 AB eta (A0).

[0079] Figure 1B (FIG. 1B) shows an exemplary 2+1 multi-specific binding protein containing an scFv that binds to a BBB target (e.g., TfR or CD98hc) linked to a C-terminus of the heavy chain (with an Fc “knob” mutation) of a full-length antibody that binds to pyroGlu3 Ab eta (A0).

[0080] Figure 2 (FIG. 2) shows an exemplary 2+2 multi-specific binding protein with two copies of an scFv that binds to a BBB target (e.g., TfR or CD98hc), wherein one copy is linked to the C-terminus of each heavy chain of a full-length antibody that binds to pyroGlu3 Abeta (A0).

[0081] Figure 3 (FIG. 3) shows exemplary multi-specific binding proteins with heterodimeric Fc regions (e.g., wherein one polypeptide of the Fc region contains a knob mutation and the other polypeptide of the Fc region contains a hole mutation). FIG. 3 includes (i) an exemplary 2+1 multi-specific binding protein with an scFv that binds to a BBB target (e.g., TfR or CD98hc) linked to the C-terminus of a heavy chain of an antibody that binds to pyroGlu3 Abeta (A0); (ii) an exemplary 1+1 multi-specific binding protein wherein a Fab of a full-length antibody that binds to pyroGlu3 Abeta (A0) is replaced with an scFv that binds to a BBB target (e.g., TfR or CD98hc); and (iii) an exemplary 1+1 multi-specific binding protein where one VHH that binds to a BBB target (e.g., TfR or CD98hc) is linked to the N-terminus of one polypeptide of an Fc region and one VHH that binds to pyroGlu3 Abeta (A0) is linked to the N-terminus of a second polypeptide of the Fc region.

[0082] Figure 4 (FIG. 4) shows ex vivo plaque phagocytosis by human macrophages 24h after addition of 0.1 pg / mL anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure. Anti-pyroGlu3 Abeta (A0) antibodies AB-6, AB-7, AB-8, AB-9, and AB-10 induced a similar extent of plaque phagocytosis as anti-Abeta (A0) antibodies donanemab and remternetug, compared to isotype control. One way ANOVA with Tukey’s multiple comparisons test. ****■. p < 0.0001.

[0083] Figure 5 (FIG. 5) shows uptake of pyroGlu3 Abeta peptide by iPSC-derived microglia following addition of 0.1 pg / mL of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure. Anti-pyroGlu3 Abeta (A0) antibodies AB-6, AB-7, AB-8, AB-9, and AB-10 displayed similar uptake of pyroGlu3-24 Abeta (A0) peptide as observed with anti-Abeta (A0) antibodies donanemab and remternetug. Anti-pyroGlu3 Abeta (A0) antibodies AB-3 and AB-8 showed a smaller increase of pyroGlu3-24 Abeta (A0) peptide uptake compared to other antibodies tested in this assay. One way ANOVA with Tukey’s multiple comparisons test. **: p < 0.005; ***: p < 0.001; ****: p < 0.0001.

[0084] Figure 6 (FIG. 6) shows representative images of target engagement with amyloid plaque of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure in 6-month-old 5XFAD mice 4 days post administration of 50 mg / kg i.v. dose. Anti-pyroGlu3-24 Abeta (A0) antibodies AB-6, AB-7, and AB-8 show binding to amyloid plaques in vivo.

[0085] Figure 7 (FIG. 7) shows quantification of huIgG positive area (i.e., amyloid plaque) in brains after dosing aged 5XFAD mice with 50 mg / kg of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure for 4 days.

[0086] Figure 8 (FIG. 8) shows serum pharmacokinetics of anti-pyroGlu3 Abeta (A0) antibodies (50 mg / kg) of the present disclosure administered to aged 5XFAD mice over the course of 4 days.

[0087] Figure 9 (FIG. 9) sets forth isoaffinity plots showing equilibrium dissociation constants (KD), as a function of the association and dissociation rate constants (k-on and k-off), of anti-pyroGlu3 Abeta (A0) huIgG antibodies of the present disclosure to pyroGlu3 Abeta (A0) peptide (pE3-24).

[0088] Figure 10 (FIG. 10) shows the effect of anti-TfR antibodies of the present disclosure on levels of circulating reticulocytes and TfR expression in huTfR KI mice. Mice were administered 3 mg / kg, 10 mg / kg, or 30 mg / kg of Iso-TfR antibody variants (L7-2 hlgGl WT, LIO-16 hlgGl WT, 39.38-hIgGl WT), and levels of circulating reticulocytes and TfR expression levels were measured 1 day (DI) and 7 days (D7) post-dose. Levels of circulating reticulocytes are shown as percent reticulocytes.

[0089] Figure 11 (FIG. 11) shows serum antibody levels in huTfR KI mice after administration of 3 mg / kg, 10 mg / kg, or 30 mg / kg of Iso-TfR antibody variants (L7-2 hlgGl WT, LIO-16 hlgGl WT, 39.38-hIgGl WT). Serum PK was measured 1 day (DI) and 7 days (D7) post-dose.

[0090] Figure 12 (FIG. 12) shows serum antibody levels in huTfR KI mice administered 3 mg / kg of remternetug and TfR-remternetug antibody variants (L7-2 hlgGl WT, LI 0-16 hlgGl WT, 39.38-hIgGl WT). Mice were dosed twice, 14 days apart. Serum PK was measured at day 1 (DI), day 7 (D7), and day 14 (DI 4) after the first dose and 24h after the second dose (day 15; D15).

[0091] Figure 13 (FIG. 13) shows brain uptake of antibodies in huTfR KI mice dosed with 3 mg / kg of remternetug and TfR-remternetug antibody variants (L7-2 hlgGl WT, LI 0-16 hlgGl WT, 39.38-hIgGl WT). Antibody concentration (ng / mg tissue) in vessel-depleted brain after 24h was measured.

[0092] Figure 14 (FIG. 14) shows ex vivo plaque phagocytosis by primary human macrophages with anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure, showing a similar extent of plaque phagocytosis as donanemab and remternetug, compared to isotype control. One way ANOVA with Tukey’s multiple comparisons test. ****■. p < 0.0001.

[0093] Figures 15A and 15B (FIG. 15A and FIG. 15B) show the uptake of pyroGlu3 Abeta (A0) polypeptide by iPSC-derived microglia following incubation with anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure. FIG. 15A shows data from the 24hr time point (one way ANOVA with Tukey’s multiple comparisons test. **: p < 0.005; ***: p < 0.001; ****: p < 0.0001). FIG. 15B shows the time course of the Abeta (A0) peptide uptake in this study.

[0094] Figure 16 (FIG. 16) shows cell uptake by immunohistochemistry of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure in hCMEC / D3 cell line.

[0095] Figure 17 (FIG. 17) shows in vivo brain uptake of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure in hu-TfR knock-in (KI) mice following a single dose.

[0096] Figure 18 (FIG. 18) shows serum pharmacokinetics of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure following administration of three i.v. doses to hu-TfR knock-in (KI) mice.

[0097] Figure 19 (FIG. 19) shows in vivo brain uptake of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure in hu-TfR knock-in (KI) mice in a multi-dose study.

[0098] Figure 20 (FIG. 20) shows in vivo brain uptake of brain uptake of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure in aged (9-month) 5XFAD mice following a single dose.

[0099] Figure 21 (FIG. 21) shows serum pharmacokinetics of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure following administration of three i.v. doses to aged 5XFAD mice in a multi-dose study.

[0100] Figure 22 (FIG. 22) shows in vivo brain uptake of anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure in aged 5XFAD mice in a multi-dose study.

[0101] Figure 23 (FIG. 23) shows immunofluorescent staining of amyloid plaques in aged 5XFAD mouse brain sections with anti-pyroGlu3 Abeta (A0) antibodies of the present disclosure.

[0102] Figure 24 (FIG. 24) shows anti-pyroGlu3 Abeta (AP)-msTfR antibodies of the present disclosure in brain parenchyma administered to aged 5XFAD mice.

[0103] Figure 25 (FIG. 25) shows reduction in Abetai-42 (AP1-42) levels in mouse brain lysates following administration of anti-pyroGlu3 Abeta (AP)-msTfR antibodies of the present disclosure.

[0104] Figures 26A and 26B (FIG. 26A and FIG. 26B) show enhanced uptake of Abeta (pyroGlu3 AP) by iMicroglia using a human CNS triple culture cell model at antibody concentrations of 1 pg / ml and 10 pg / ml, respectively.

[0105] Figures 27A and 27B (FIG. 27A and FIG. 27B) show uptake of Abeta (pyroGlu3 AP) by anti-Abeta (AP) antibody remternetug-TfR.39.38 having an effectorless Fc mutation LALAPS or a wildtype Fc in microglia in triple cultures and human fetal astrocyte s+neur on cell population, respectively.

[0106] Figure 28 (FIG. 28) shows transcytosis of anti-pyroGlu3 Abeta (AP)-TfR antibodies across human brain endothelial cell line hCMEC / D3.

[0107] Figure 29 (FIG. 29) shows transcytosis of anti-pyroGlu3 Abeta (AP)-huTfR antibodies across primary cynomolgus brain endothelial cells.

[0108] Figure 30 (FIG. 30) shows the level of ADCC induced by anti-pyroGlu3 Abeta (AP)-huTfR antibodies of the present disclosure.

[0109] Figures 31 A to 31D (FIG. 31A-FIG. 31D) show that binding kinetics for 2+1 format antibodies of the present disclosure to human and cynomolgus Fc gamma receptors have kinetic profiles comparable with that of wildtype IgGl Fc binding.

[0110] Figures 32A and 32B (FIG. 32A and FIG. 32B) show binding kinetics of anti-pyroGlu3 Abeta (AP)-TfR antibodies of the present disclosure to human (FIG. 32A) andcynomolgus (FIG. 32B) Fc gamma receptors, the binding of which are not distinguishable from wildtype IgGl Fc.

[0111] Figure 33 (FIG. 33) shows serum pharmacokinetics of antibodies in huTfR KI mice that received two weekly doses of 3 mg / kg or 30 mg / kg of antibody AB-8 huIgGl and various AB-8-TfR variants. Mice were dosed on DI (day 1) and D8 (day 8). Serum PK was measured at D2 (day 2), D8 (day 8) before dosing, and D9 (day 9).

[0112] Figures 34A and 34B (FIG. 34A and FIG. 34B) show reticulocyte percentages in peripheral whole blood assessed at DI (day 1) and D7 (day 7) (FIG. 34A) post first dose and on D8 (day 8) (FIG. 34B) at study termination in huTfR KI mice.

[0113] Figure 35 (FIG. 35) shows brain uptake of antibodies in huTfR KI mice that received two weekly doses of 3 mg / kg or 30 mg / kg of antibody AB-8 huIgGl and various AB-8-TfR variants. Antibody levels (ng / mg protein) in vessel-depleted brain at 24 hours after the second dose was measured.

[0114] Figure 36 (FIG. 36) shows reticulocyte levels as percent change from baseline in whole blood at day 10 from non-human primates (NHPs) dosed twice over 8 days with antibody AB-8 huIgGl and various AB-8-TfR variants dosed at 3 mg / kg and 30 mg / kg.

[0115] Figure 37 (FIG. 37) shows red blood cells (RBCs) levels as a percent change from baseline in whole blood at day 10 from NHPs receiving 2 doses over 8 days of antibody AB-8 huIgGl and various AB-8-TfR variants dosed at 3 mg / kg and 30 mg / kg.

[0116] Figure 38 (FIG. 38) shows serum pharmacokinetic profile from an NHP study with antibody AB-8 huIgGl and AB-8-TfR variants.

[0117] Figure 39 (FIG. 39) shows brain uptake of dosed antibodies from an NHP study with antibody AB-8 huIgGl and AB-8-TfR variants. Enhanced brain uptake was observed at both 3 mg / kg and 30 mg / kg of all TfR antibodies compared to that observed with AB-8 hu IgG naked control antibody.

[0118] Figure 40 (FIG. 40) shows AB-8-TfR42Q and AB-8-TfRL10-8 colocalizes with Abeta (A0) plaques on human Alzheimer's disease (AD) brain sections.

[0119] Figure 41 (FIG. 41) shows quantification of plaque and dosed antibody colocalized area in the hippocampus of 5XFAD mice.

[0120] Figure 42 (FIG. 42) shows TfR enhanced CSF uptake of AB-8-TfR42Q in NHPs.

[0121] Figure 43 (FIG. 43) shows a bar graph of the CSF concentrations after the 3 mg / kg dose of AB-8 huIgG (circle data points) and AB-8-TfR42Q (triangle data points) at 2, 4, and 10 days after dosing.

[0122] Figure 44 (FIG. 44) shows TfR levels in various brain regions from NHP dosed with anti-pyroGlu3 Abeta (A0) antibodies.

[0123] Figure 45 (FIG. 45) shows serum and brain antibody levels after various time points in hu TfR KI mice dosed either intravenously or subcutaneously with 3 mg / kg of anti-pyroGlu3 Abeta (A0) antibodies.DETAILED DESCRIPTION OF THE PRESENT DISCLOSURE

[0124] The present disclosure relates to anti-beta amyloid antibodies that specifically bind N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta (A0)), multi-specific binding proteins comprising an anti-Abeta (pyroGlu3 A0) antigen-binding domain and a blood brain barrier (BBB) antigen-binding domain that specifically binds to a BBB target selected from human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc), and uses (e.g., therapeutic uses) thereof.

[0125] Amyloid beta (Abeta; AP) peptide in circulating form is composed of 38-43 amino acids (mostly 38, 40, or 42 amino acids) resulting from the cleavage of amyloid precursor protein (APP). Conversion of Abeta (AP) from soluble to insoluble forms, and the deposition of these insoluble forms as neuritic and cerebrovascular plaques in the brain has been associated with a number of conditions and diseases, including Alzheimer's disease (AD), Down's syndrome, cerebral amyloid angiopathy (CAA), and other amyloid-beta associated disorders.

[0126] The deposits found in plaques are comprised mainly of a heterogeneous mixture of Abeta (AP) peptides. PyroGlu3 Abeta (AP) (also referred to as N3pE amyloid beta, N3pE Abeta (AP), N3pGlu Abeta (AP), or AbetaP3_42 (ApP3_42)) is a truncated form of Abeta (AP) peptide found only amyloid plaques. PyroGlu3 Abeta (AP) lacks the first two amino acid residues at the N-terminus of Abeta (AP) and has a pyroglutamate derived from the glutamic acid at the third amino acid position. Although PyroGlu3 Abeta (AP) peptide is a minor component of deposited Abeta (AP) in the brain, studies have demonstrated that PyroGlu3 Abeta (AP) peptide has aggressive aggregation properties and accumulates early in the deposition cascade.

[0127] Passive transfer of substances from blood to brain is restricted by the blood brain barrier (BBB). The BBB provides precise control of central nervous system (CNS) homeostasis allowingfor proper neuronal function and also protecting neural tissue from toxins and pathogens.However, the BBB poses a problem with regard to delivering therapeutics to the CNS. While recombinant proteins and antibody therapeutics have shown much success outside the CNS, such biologies do not cross the BBB efficiently. As a result, delivery of certain therapeutics to the CNS has relied on injection of the therapeutic directly into the CNS; an invasive procedure with limited efficacy due to rapid export of cerebral spinal fluid (CSF) containing the therapeutic from the brain to the blood. Alternatively, a therapeutic intended for the CNS may be administered systemically at doses high enough to allow for sufficient penetration of the BBB by the therapeutic. However, this approach may result in unintended effects due to the high dose in the periphery or increased manufacturing and formulation burdens to achieve the high dose.

[0128] Antibodies that target Abeta (A0) have been previously described, as well as polyclonal and monoclonal antibodies that are directed specifically against pyroGlu3 Abeta (A0) (e.g., bapineuzumab, gantenerumab, aducanumab, GSK933776, Alia-1758, BAN 2802, solanezumab, crenezumab, ponezumab, lecanemab and trontinemab; see, e.g., USPN 7,122,374, W02002 / 046237, W02004 / 080419, WO2012 / 021469, WO2012 / 021475, WO2017 / 123517, WO2017 / 160555, WO2018 / 005282, W02018 / 031361, WO2018 / 194951, WO2022 / 150735, WO2022 / 192636, WO2022 / 251048, W02024 / 020470, WO2024 / 107683, W02003 / 070760, WO2014 / 056816, W02015 / 120280, W02014 / 089500, WO2016 / 087944, WO2017 / 211827, WO2019 / 040612, W02021 / 081101, WO2022 / 261026, W02023 / 081194, W02020 / 023530, WO2021 / 186245, WO2023 / 111618, WO2024 / 118665, WO2024 / 206161, WO2023 / 170295, WO2023 / 170291, WO2023 / 170290, W02018 / 011353, and WO2024 / 133925). Despite recent approval of two anti-Abeta (A0) antibodies for the treatment of Alzheimer’s Disease (donanemab-azbt, lecanemab-irmb), there is still a need for therapeutically useful antibody antagonists to pyroGlu3 Abeta (A0) possessing a number of desirable properties, such as, for example, high affinity, plaque-clearance, and the ability to effectively and efficiently cross the blood brain barrier.

[0129] The present disclosure provides antibodies that specifically bind pyroGlu3 Abeta (A0).

[0130] The present disclosure further provides anti-pyroGlu3 Abeta (A0) multi-specific binding proteins. Anti-pyroGlu3 Abeta (A0) multi-specific binding proteins provided herein are capable of crossing the blood brain barrier and capable of transporting an anti-pyroGlu3 Abeta (A0) antigen binding domain, antibody, or antigen-binding fragment thereof across the bloodbrain barrier, thus enhancing or increasing exposure of the anti-pyroGlu3 Abeta (A0) antigen binding domain, antibody, or antigen-binding fragment thereof to the CNS.

[0131] The present disclosure provides various multi-specific binding protein formats comprising an anti-blood brain barrier antigen-binding domain linked to an anti-pyroGlu3 Abeta (A0) antigen -binding domain (e.g., an antibody antigen binding domain or antibody or antigenbinding fragment thereof that binds specifically to pyroGlu3 Abeta (A0)).

[0132] In some aspects, a multi-specific binding protein of the present disclosure with a “1 +1 multi-specific binding protein format” comprises a TfR antigen-binding domain in a bivalent, bispecific format comprising (i) one antigen-binding domain that binds to human TfR; and (ii) one antigen-binding domain that binds to pyroGlu3 Abeta (A0). Such a “1+1” format can also comprise an Fc domain or Fc region. In some aspects, a multi-specific binding protein of the present disclosure with a “1 +1 multi-specific binding protein format” comprises a CD98hc antigen-binding domain in a bivalent, bi-specific format comprising (i) one antigen-binding domain that binds to human CD98hc; and (ii) one antigen-binding domain that binds to pyroGlu3 Abeta (A0). Such a “1+1” format can also comprise an Fc domain or Fc region.

[0133] In some aspects, a multi-specific binding protein of the present disclosure with a “2+1 multi-specific binding protein format” comprises a TfR antigen-binding domain in a trivalent, bispecific format comprising (i) an antigen-binding domain that binds to human TfR and (ii) an antibody comprising two anti-pyroGlu3 Abeta (A0) antigen-binding domains, wherein the antibody comprises two heavy chains and two light chains; wherein the antigen-binding domain that binds to human TfR is linked to the C-terminus of one of the two antibody heavy chains. In some aspects, a multi-specific binding protein of the present disclosure with a “2+1 multispecific binding protein format” comprises a CD98hc antigen-binding domain in a trivalent, bispecific format comprising (i) an antigen-binding domain that binds to human CD98hc, and (ii) an antibody comprising two anti-pyroGlu3 Abeta (A0) antigen-binding domains, wherein the antibody comprises two heavy chains and two light chains; wherein the antigen-binding domain that binds to human CD98hc is linked to the C-terminus of one of the two antibody heavy chains.

[0134] In some embodiments, the multi-specific protein comprises a “1+2” multi-specific protein format, wherein the multi-specific protein is trivalent and bi-specific. In some embodiments, the multi-specific protein comprises: (i) two antigen-binding domains that specifically bind to human TfR and (ii) an antibody or antigen-binding fragment thereof that specifically binds to pyroGlu3 Abeta (A0), wherein the antibody or antigen-binding fragmentthereof is a scFv, VHH, or Fab, optionally comprising an Fc domain. In some embodiments, the multi-specific protein comprises a “1+2” multi-specific protein format, wherein the multispecific protein is trivalent and bi-specific. In some embodiments, the multi-specific protein comprises: (i) two antigen-binding domains that specifically binds to human CD98hc and (ii) an antibody or antigen-binding fragment thereof that specifically binds to pyroGlu3 Abeta (A0), wherein the antibody or antigen-binding fragment thereof is a scFv, VHH, or Fab, optionally comprising an Fc domain.

[0135] In some aspects, a multi-specific binding protein of the present disclosure with a “2+2 multi-specific binding protein format” comprises a TfR antigen-binding domain in a tetravalent, bi-specific format comprising (i) two antigen-binding domains that bind to human TfR and (ii) an antibody comprising two anti-pyroGlu3 Abeta (A0) antigen-binding domains, wherein the antibody comprises two heavy chains and two light chains; wherein one antigen-binding domain that binds to human TfR is linked to the C-terminus of one of the two antibody heavy chains, and the other antigen-binding domain that binds to human TfR is linked to the C-terminus of the other of the two antibody heavy chains. The two antigen-binding domains that bind to human TfR can comprise the same amino acid sequence. The two antigen-binding domains that bind to human TfR can comprise different amino acid sequences. In some aspects, a multi-specific binding protein of the present disclosure with a “2+2 multi-specific binding protein format” comprises a CD98hc antigen-binding domain in a tetravalent, bi-specific format comprising (i) two antigen-binding domains that bind to human CD98hc and (ii) an antibody comprising two anti-pyroGlu3 Abeta (A0) antigen-binding domains, wherein the antibody comprises two heavy chains and two light chains; wherein one antigen-binding domain that binds to human CD98hc is linked to the C-terminus of one of the two antibody heavy chains, and the other antigen-binding domain that binds to human CD98hc is linked to the C-terminus of the other of the two antibody heavy chains. In some aspects, a multi-specific binding protein of the present disclosure with a “2+2 multi-specific binding protein format” comprises a TfR antigen-binding domain in a tetravalent, bi-specific format comprising (i) two antigen-binding domains that bind to human pyroGlu3 Abeta (A0) and (ii) an antibody comprising two anti-TfR antigen-binding domains, wherein the antibody comprises two heavy chains and two light chains; wherein one antigenbinding domain that binds to human pyroGlu3 Abeta (A0) is linked to the C-terminus of one of the two antibody heavy chains, and the other antigen-binding domain that binds to human pyroGlu3 Abeta (A0) is linked to the C-terminus of the other of the two antibody heavy chains.The two antigen-binding domains that bind to human pyroGlu3 Abeta (A0) can comprise the same amino acid sequence. The two antigen-binding domains that bind to human pyroGlu3 Abeta (AP) can comprise different amino acid sequences. In some aspects, a multi-specific binding protein of the present disclosure with a “2+2 multi-specific binding protein format” comprises a CD98hc antigen-binding domain in a tetravalent, bi-specific format comprising (i) two antigen-binding domains that bind to human pyroGlu3 Abeta (AP) and (ii) an antibody comprising two anti-CD98hc antigen-binding domains, wherein the antibody comprises two heavy chains and two light chains; wherein one antigen-binding domain that binds to human pyroGlu3 Abeta (AP) is linked to the C-terminus of one of the two antibody heavy chains, and the other antigen-binding domain that binds to human pyroGlu3 Abeta (AP) is linked to the C-terminus of the other of the two antibody heavy chains.

[0136] The techniques and procedures described or referenced herein are generally well understood and commonly employed using conventional methodology by those skilled in the art, such as, for example, the widely utilized methodologies such as those described in Sambrook et al. Molecular Cloning: A Laboratory Manual 3d edition (2001) Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N. Y.; Current Protocols in Molecular Biology (F. M. Ausubel, et al. eds., (2003); Monoclonal Antibodies: A Practical Approach (P. Shepherd and C. Dean, eds., Oxford University Press, 2000).Definitions

[0137] The terms “amyloid beta,” “Abeta,” “AP,” and “A-beta,” used interchangeably herein, refer to the peptide formed by the proteolytic cleavage of a larger glycoprotein called amyloid precursor protein (APP).

[0138] The terms “pyroGlu3 AP,” “N3pE Ap,” or “N3pGlu AP” peptide, polypeptide, or protein, used interchangeably herein, refer to any native Ap peptide or protein bearing aminoterminal pyroglutamate at amino acid position 3 from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys (cynos)) and rodents (e.g., mice and rats), unless otherwise indicated. In some aspects, the term encompasses both wild-type sequences and naturally occurring variant sequences, e.g., splice variants or allelic variants. In some aspects, the pyroGlu3 AP is human pyroGlu3 Ap.

[0139] The terms “anti-pyroGlu3 AP antibody,” and “antibody that binds to pyroGlu3 AP,” and “antibody that specifically binds pyroGlu3 AP” refer to an antibody that is capable ofbinding pyroGlu3 A0 with sufficient affinity such that the antibody is useful as a diagnostic and / or therapeutic agent in targeting pyroGlu3 A0.

[0140] The terms “central nervous system” or “CNS” refer to the complex of nerve tissues that control bodily function and includes the brain and spinal cord.

[0141] The terms “blood brain barrier” or “BBB” refer to a network of brain capillary endothelial cells that are closely sealed by tight junctions.

[0142] A “central nervous system antigen” or “CNS antigen” is an antigen expressed in the CNS, including the brain.

[0143] A “brain antigen” is a CNS antigen expressed in the brain.

[0144] The terms “BBB target,” “BBB protein,” “BBB receptor,” and “BBB antigen” refer to a target / protein / receptor / antigen expressed on blood brain barrier cells (e.g., TfR or CD98hc). In some aspects, an antigen-binding domain (e.g., in an antibody, scFv, or Fab) that binds to the BBB target / protein / receptor / antigen allows transportation of a molecule or compound associated with said antigen-binding domain across the BBB.

[0145] A “neurological disorder” as used herein refers to a disease or disorder which affects the CNS and / or which has an etiology in the CNS. Exemplary CNS diseases or disorders include, but are not limited to, neuropathy, amyloidosis, Alzheimer’s disease, and neurodegenerative diseases and disorders.

[0146] The terms “Transferrin receptor,” “TfR,” “TfR polypeptide,” and “TfR protein” are used interchangeably herein to refer to any native TfR from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys (cynos)) and rodents (e.g., mice and rats), unless otherwise indicated. TfR is also referred to as transferrin receptor protein 1, TR, tfRl, Trfr, T9, and p90. In some aspects, the term encompasses both wild-type sequences and naturally occurring variant sequences, e.g., splice variants or allelic variants. In some aspects, the term encompasses “full-length,” unprocessed TfR, as well as any form of TfR that results from processing in the cell. Full-length transferrin receptor protein includes a short N-terminal intracellular region, a transmembrane region, and a large extracellular domain. The extracellular domain is characterized by three domains: a protease-like domain, a helical domain, and an apical domain. In some aspects, the TfR is human TfR. As used herein, the term “human TfR” refers to a polypeptide with the following amino acid sequence:MMDQ ARS AF SNLFGGEPLS YTRF SLARQ VDGDNSHVEMKL AVDEEENADNNTKANVT KPKRCSGSICYGTIAVIVFFLIGFMIGYLGYCKGVEPKTECERLAGTESPVREEPGEDFPAARRLYWDDLKRKLSEKLDSTDFTGTIKLLNENSYVPREAGSQKDENLALYVENQFREFK LSKVWRDQHFVKIQVKDSAQNSVIIVDKNGRLVYLVENPGGYVAYSKAATVTGKLVHA NFGTKKDFEDLYTPVNGSIVIVRAGKITFAEKVANAESLNAIGVLIYMDQTKFPIVNAELS FFGHAHLGTGDPYTPGFPSFNHTQFPPSRSSGLPNIPVQTISRAAAEKLFGNMEGDCPSD WKTDSTCRMVTSESKNVKLTVSNVLKEIKILNIFGVIKGFVEPDHYVVVGAQRDAWGPG AAKSGVGTALLLKLAQMFSDMVLKDGFQPSRSIIFASWSAGDFGSVGATEWLEGYLSSL HLKAFTYINLDKAVLGTSNFKVSASPLLYTLIEKTMQNVKHPVTGQFLYQDSNWASKVE KLTLDNAAFPFLAYSGIPAVSFCFCEDTDYPYLGTTMDTYKELIERIPELNKVARAAAEV AGQFVIKLTHDVELNLDYERYNSQLLSFVRDLNQYRADIKEMGLSLQWLYSARGDFFR ATSRLTTDFGNAEKTDRFVMKKLNDRVMRVEYHFLSPYVSPKESPFRHVFWGSGSHTLP ALLENLKLRKQNNGAFNETLFRNQLALATWTIQGAANALSGDVWDIDNEF (SEQ ID NO: 2).

[0147] As used herein, the terms “CD98hc,” “CD98hc polypeptide,” and “CD98hc protein” are used interchangeably herein to refer to any native CD98hc from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys (cynos)) and rodents (e.g., mice and rats), unless otherwise indicated. CD98hc is also referred to as 4F2 cell-surface antigen heavy chain, 4F2hc, 4F2 heavy chain antigen, lymphocyte activation antigen 4F2 large subunit, solute carrier family 3 member 2, and CD98. CD98hc protein is encoded by the SLC3A2 gene and is part of the large amino acid transporter (LAT) complex. In some aspects, the term encompasses both wild-type sequences and naturally occurring variant sequences, e.g., splice variants or allelic variants. In some aspects, the term encompasses “full-length,” unprocessed CD98hc, as well as any form of CD98hc that results from processing in the cell. In some aspects, the CD98hc is human CD98hc. As used herein, the term “human CD98hc” refers to a polypeptide with the following amino acid sequence:MELQPPEASIAVVSIPRQLPGSHSEAGVQGLSAGDDSELGSHCVAQTGLELLASGDPLPS ASQNAEMIETGSDCVTQAGLQLLASSDPPALASKNAEVTGTMSQDTEVDMKEVELNEL EPEKQPMNAASGAAMSLAGAEKNGLVKIKVAEDEAEAAAAAKFTGLSKEELLKVAGSP GWVRTRWALLLLFWLGWLGMLAGAVVIIVRAPRCRELPAQKWWHTGALYRIGDLQAF QGHGAGNLAGLKGRLDYLSSLKVKGLVLGPIHKNQKDDVAQTDLLQIDPNFGSKEDFD SLLQSAKKKSIRVILDLTPNYRGENSWFSTQVDTVATKVKDALEFWLQAGVDGFQVRDI ENLKDASSFLAEWQNITKGFSEDRLLIAGTNSSDLQQILSLLESNKDLLLTSSYLSDSGST GEHTKSL VTQYLNATGNRWC SWSLSQ ARLLTSFLP AQLLRLYQLMLFTLPGTPVF S YGDEIGLDAAALPGQPMEAPVMLWDESSFPDIPGAVSANMTVKGQSEDPGSLLSLFRRLSDQ RSKERSLLHGDFHAF S AGPGLF S YIRHWDQNERFL VVLNFGD VGLS AGLQ ASDLPAS AS LPAKADLLLSTQPGREEGSPLELERLKLEPHEGLLLRFPYAA (SEQ ID NO: 3).

[0148] As used herein, the terms “antibody” and “immunoglobulin” are used interchangeably and refer to an antibody molecule that recognizes and specifically binds to a target, such as a protein, polypeptide, peptide, carbohydrate, polynucleotide, lipid, or combinations of the foregoing (e.g., a glycoprotein), through at least one antigen recognition site within the variable region of the immunoglobulin molecule. The term “antibody” encompasses monoclonal antibodies, chimeric antibodies, humanized antibodies, human antibodies, multi-specific (e.g., bispecific) antibodies, and any other immunoglobulin molecule so long as the antibodies exhibit the desired biological activity. An antibody can be of any the five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, or subclasses (isotypes) thereof (e.g., IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2), based on the identity of their heavy-chain constant regions referred to as alpha, delta, epsilon, gamma, and mu, respectively. The different classes of antibodies have different and well-known subunit structures and three-dimensional configurations. For the structure and properties of the different classes of antibodies, see, e.g., Basic and Clinical Immunology, 8th Ed., Daniel P. Stites, Abba I. Terr and Tristram G. Parslow (eds.), Appleton & Lange, Norwalk, CT, 1994, page 71 and Chapter 6.

[0149] The terms “full-length antibody,” “intact antibody,” or “whole antibody” are used interchangeably to refer to an antibody, such as an anti-pyroGlu3 A0 antibody, in its substantially intact form, as opposed to an antibody fragment. Specifically, full-length antibodies include those with 2 light chains and 2 heavy chains including an Fc region. The constant domains may be native sequence constant domains e.g., human native sequence constant domains) or amino acid sequence variants thereof. In some cases, the intact antibody may have one or more effector functions.

[0150] The terms “monovalent antibody” or “monoarm antibody” refers to an antibody having a single antigen-binding recognition domain that is specific to a target antigen (i.e., the antibody comprises no more than one antigen-binding domain). In some embodiments, a single antigenbinding domain comprises a single variable region heavy chain polypeptide and a single variable region light chain polypeptide. An antibody that is “monovalent” for a target comprises no more than one antigen-binding domain for that target.

[0151] The terms “anti-TfR antibody,” “antibody that binds to TfR,” and “antibody that specifically binds TfR” refer to an antibody that is capable of binding TfR with sufficient affinity such that the antibody is useful as a diagnostic and / or therapeutic agent in targeting TfR. In some aspects, the anti-TfR antibody is capable of transporting another diagnostic and / or therapeutic agent into the brain. In certain aspects, an anti-TfR antibody binds to an epitope of TfR that is conserved among TfR from different species.

[0152] The terms “anti-TfR antigen-binding domain,” “antigen-binding domain that binds to TfR,” “anti-TfR antigen-binding region,” “antigen-binding region that binds to TfR,” and “TfR binding domain” refer to an antigen-binding domain that binds to TfR with sufficient affinity such that the antigen-binding domain is useful as a diagnostic and / or therapeutic agent in targeting TfR. In one aspect, the extent of binding of an anti-TfR antigen-binding domain to an unrelated, non-TfR polypeptide is less than about 10% of the binding of the antigen-binding domain to TfR as measured, e.g., by a radioimmunoassay (RIA). In certain embodiments, an anti-TfR antigen-binding domain binds to an epitope of TfR that is conserved among TfR from different species.

[0153] The terms “anti-CD98hc antibody,” “antibody that binds to CD98hc,” and “antibody that specifically binds CD98hc” refer to an antibody that is capable of binding CD98hc with sufficient affinity such that the antibody is useful as a diagnostic and / or therapeutic agent in targeting CD98hc. In some aspects, the anti-CD98hc antibody is capable of transporting another diagnostic and / or therapeutic agent into the brain. In certain aspects, an anti-CD98hc antibody binds to an epitope of CD98hc that is conserved among CD98hc from different species.

[0154] The terms “anti-CD98hc antigen-binding domain,” “antigen-binding domain that binds to CD98hc,” “anti-CD98hc antigen-binding region,” “antigen-binding region that binds to CD98hc,” and “CD98hc binding domain” refer to an antigen-binding domain that binds to CD98hc with sufficient affinity such that the antigen-binding domain is useful for targeting CD98hc and / or useful as a diagnostic agent, a therapeutic agent, or for transporting a molecule or compound across the BBB. In one aspect, the extent of binding of an anti-CD98hc antigenbinding domain to an unrelated, non-CD98hc polypeptide is less than about 10% of the binding of the antigen-binding domain to CD98hc as measured, e.g., by a radioimmunoassay (RIA). In certain aspects, an anti-CD98hc antigen-binding domain binds to an epitope of CD98hc that is conserved among CD98hc from different species.

[0155] An “antigen-binding domain” or “antigen-binding region” refers to a monovalent portion of an antibody that binds to an antigen. An “antigen-binding domain” can comprise the antigenic determining regions of an antibody (e.g., the complementarity determining regions (CDRs)). An antibody or antigen-binding fragment thereof (including mono-specific and multispecific (e.g., bi-specific) antibodies or antigen-binding fragments thereof can comprise an antigen-binding domain. In some aspects, an antigen-binding domain is not present in the context of an antibody. In some aspects, provided herein is an antibody, a VHH, a Fab, a Fab’, a Fab’-SH, a F(ab’)2, a Fv, or a scFv comprising an antigen-binding domain.

[0156] The terms “full-length antibody,” “intact antibody” or “whole antibody” are used interchangeably to refer to an antibody in its substantially intact form, as opposed to an antibody fragment. Specifically, whole antibodies include those with heavy and light chains including an Fc region. The constant regions can be native sequence constant regions (e.g., human native sequence constant regions) or amino acid sequence variants thereof. In some cases, the intact antibody can have one or more effector functions. The C-terminal lysine (residue 447 according to the EU numbering system) of an intact antibody can be removed, for example, during production or purification of the antibody, or by recombinantly engineering the nucleic acid encoding a heavy chain of the antibody. Accordingly, a composition of “full-length antibodies,” “intact antibodies,” or “whole antibodies” can comprise antibody populations with all K447 residues removed, antibody populations with no K447 residues removed, and antibody populations having a mixture of antibodies with and without the K447 residue.

[0157] The term “native IgG antibodies” refers to heterotetrameric glycoproteins of about 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 one covalent disulfide bond, while the number of disulfide linkages varies among the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intra-chain disulfide bridges. Each heavy chain has at one end a variable domain (VH) followed by a number of constant domains. Each light chain has a variable domain at one end (VL) and a constant domain at its other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the light chain variable domain is aligned with the variable domain of the heavy chain.Particular amino acid residues are believed to form an interface between the light chain and heavy chain variable domains.

[0158] The terms “VH” and “VH domain” are used interchangeably to refer to the heavy chain variable region of an antibody.

[0159] As used herein, the term “heavy chain” when used in reference to an antibody can refer to any distinct type, e.g., alpha (a), delta (8), epsilon (a), gamma (y), and mu (p), based on the amino acid sequence of the constant region, which give rise to IgA, IgD, IgE, IgG, and IgM classes of antibodies, respectively, including subclasses of IgG, e.g., IgGi, IgG2, IgGs, and IgG4. Heavy chain amino acid sequences are well known in the art. In some aspects, the heavy chain is a human heavy chain.

[0160] The terms “VL” and “VL domain” are used interchangeably to refer to the light chain variable region of an antibody.

[0161] As used herein, the term “light chain” when used in reference to an antibody can refer to any distinct type, e.g., kappa (K) or lambda (1) based on the amino acid sequence of the constant regions. Light chain amino acid sequences are well known in the art. In some aspects, the light chain is a human light chain.

[0162] The terms “variable region” or “variable domain” refers to the amino-terminal domains of the heavy or light chain of the antibody. The variable domains of the heavy chain and light chain may be referred to as “VH” and “VL”, respectively. These domains are generally the most variable parts of the antibody (relative to other antibodies of the same class) and contain the antigen binding sites. Generally, the variable region or variable domain is typically about the amino-terminal 110 to 120 amino acids or 110 to 125 amino acids in the mature heavy chain and about 90 to 115 amino acids in the mature light chain.

[0163] The term “Fv” or “variable fragment” refers to the minimum antibody fragment which comprises a complete antigen-binding site and consists of a dimer of one heavy-chain variable region (VH) and one light-chain variable region (VL). From the folding of these two domains emanate six hypervariable loops (3 loops each from the H and L chain) that contribute the amino acid residues for antigen binding and confer antigen binding specificity to the antibody.

[0164] As used herein, the term “constant region” is a region of an antibody that is not the variable region of the antibody, e.g., a carboxyl terminal portion of a light and / or heavy chain which is not directly involved in binding of an antibody to antigen, but which can exhibit various effector functions, such as interaction with the Fc receptor. The constant region of an immunoglobulin molecule generally has a more conserved amino acid sequence relative to an immunoglobulin variable domain. In certain aspects, an antibody or antigen-binding fragmentcomprises a constant region or portion thereof that is sufficient for antibody-dependent cell-mediated cytotoxicity (ADCC).

[0165] A “constant domain” means a domain within a constant region that is capable of forming an immunoglobulin fold. Constant domains include the CHI, CH2, CH3, and CL domains.

[0166] The term “antibody fragment” refers to a portion of an antibody. An “antigen-binding fragment” of an antibody refers to a portion of an antibody that binds to an antigen. An antigenbinding fragment of an antibody can comprise the antigenic determining regions of an antibody (e.g., the complementarity determining regions (CDRs)). Examples of antigen-binding fragments of antibodies include, but are not limited to Fab, Fab’, F(ab’)2, and Fv fragments, linear antibodies, and single chain antibodies. An antigen-binding fragment of an antibody can be monovalent or multi-valent (e.g., bi-valent). An antigen-binding fragment of an antibody can be monospecific or multi-specific (e.g., bi-specific.) An antigen-binding fragment of an antibody can be derived from any animal species, such as rodents (e.g., mouse, rat, or hamster) and humans or can be artificially produced.

[0167] The term “Fab” or “fragment antigen-binding region” refers to a region on an antibody that binds to antigens. It is composed of one constant domain, one variable domain of the heavy chain and one variable domain of the light chain. Each Fab fragment is monovalent with respect to antigen binding, i.e., it has a single antigen-binding site.

[0168] The term “F(ab’)2 fragment” refers to antibody fragments that are generated by pepsin digestion of whole IgG antibodies to remove most of the Fc region while leaving intact some of the hinge region. F(ab’)2 fragments have two antigen-binding F(ab) portions linked together by disulfide bonds, and therefore are divalent with a molecular weight of about 110 kDa. Fab’ fragments differ from Fab fragments by having a few additional residues at the carboxy terminus of the CHI domain including one or more cysteines from the antibody hinge region. Fab’-SH is the designation herein for Fab’ in which the cysteine residue(s) of the constant domains bear a free thiol group.

[0169] As used herein, a “Fc fragment,” “fragment crystallizable region,” or “Fc region” is composed of two or more polypeptides, each being an antibody heavy chain fragment and each containing at least one (e.g., two or three) heavy chain constant domains. In some aspects, an Fc region is composed of two heavy chain fragments, each containing a CH2 domain and a CH3 domain. Although the boundaries of the Fc region of an immunoglobulin heavy chain might vary,the human IgG heavy-chain Fc region is usually defined to stretch from an amino acid residue at position Cys226, or from Pro230, to the carboxyl-terminus thereof. The C-terminal lysine (residue 447 according to the EU numbering system) of the Fc region can be removed, for example, during production or purification of the antibody, or by recombinantly engineering the nucleic acid encoding a heavy chain of the antibody. Accordingly, an Fc region may not contain any K447 residues, may contain at least one polypeptide containing a K447 residue and at least one polypeptide that does not contain a K447 residue, or may only contain polypeptides that include a K447 residue. Suitable native- sequence Fc regions for use in the present disclosure include human IgGl, IgG2, IgG3 and IgG4. In a native antibody, an Fc region refers to the region of an antibody that interacts with cell surface receptors called Fc receptors and some proteins of the complement system. However, as used herein, an Fc region can be modified to increase, decrease, or eliminate interaction with Fc receptors and / or proteins of the compliment system. In native IgG, IgA and IgD antibody isotypes, the Fc region is composed of two identical protein fragments, derived from the second and third constant domains of the antibody’s two heavy chains. However, as used herein, the two or more polypeptides in an “Fc region” do not need to have identical sequences. In some aspects, an “Fc region” comprises a first polypeptide comprising an Fc domain (e.g., IgGl Fc domain) with a knob mutation and a second polypeptide comprising an Fc domain (e.g., IgGl Fc domain) with a hole mutation. In native IgM and IgE antibody isotypes, the Fc region contains three heavy chain constant domains (CH domains 2-4) in each polypeptide chain.

[0170] The term “Fc domain” refers to one or more constant region domains within an Fc region, such as a CH2 or CH3 domain, in a single polypeptide. In some aspects, the Fc domain includes at least one amino acid deletion, addition, or substitution as compared to the amino acid sequence of a native Fc domain, such as by including a set of “knob-into-hole” deletions, additions, or substitutions or including amino acid deletions, additions, or substitutions to effect electrostatic steering of the Fc domain to favor attractive interactions among different polypeptide chains. In some aspects, the Fc domain is in a “knob” format. In some aspects, the Fc domain is in a “hole” format.

[0171] The term “single-chain Fv”, also abbreviated as “sFv” or “scFv”, refers to antibody fragments that comprise the VH and VL antibody domains that form a single polypeptide chain. In some aspects, the scFv polypeptide comprises a polypeptide linker between the VH and VL domains, which enables the scFv to form the desired structure for antigen binding.

[0172] The term “diabodies” refers to small antibody fragments prepared by constructing scFv fragments with short linkers (about 5-10 residues) between the VH and VL domains, such that inter-chain but not intra-chain pairing of the variable domains is achieved, thereby resulting in a bivalent fragment (i.e., a fragment having two antigen-binding sites). Bispecific diabodies are heterodimers of two “crossover” scFv fragments in which the VH and VL domains of the two antibodies are present on different polypeptide chains.

[0173] The term “CDR” or “complementarity determining region” refers to hypervariable regions in the variable region of an immunoglobulin that determine antibody diversity and antigen specificity.

[0174] The term “Kabat numbering” and like terms are recognized in the art and refer to a system of numbering amino acid residues in the heavy and light chain variable regions of an antibody or an antigen-binding fragment thereof. In certain aspects, CDRs can be determined according to the Kabat numbering system (see, e.g., Kabat EA & Wu TT (1971) Ann NY Acad Sci 190: 382-391 and Kabat EA et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U. S. Department of Health and Human Services, NIH Publication No. 91-3242). Using the Kabat numbering system, CDRs within an antibody heavy chain molecule are typically present at amino acid positions 31 to 35, which optionally can include one or two additional amino acids, following 35 (referred to in the Kabat numbering scheme as 35 A and 35B) (CDRH1), amino acid positions 50 to 65 (CDRH2), and amino acid positions 95 to 102 (CDRH3). Using the Kabat numbering system, CDRs within an antibody light chain molecule are typically present at amino acid positions 24 to 34 (CDRL1), amino acid positions 50 to 56 (CDRL2), and amino acid positions 89 to 97 (CDRL3).

[0175] The term “Chothia” refers to the location of the structural loops (see, e.g., Chothia C & Lesk AM, (1987), J Mol Biol 196: 901-917; Al-Lazikani B et al., (1997) J Mol Biol 273: 927-948; Chothia C et al., (1992) J Mol Biol 227: 799-817; Tramontane A et al., (1990) J Mol Biol 215(1): 175-82; and U. S. Patent No. 7,709,226). In some aspects, the Chothia residues are numbered as shown in the table below.

[0176] The AbM hypervariable regions represent a compromise between the Kabat CDRs and Chothia structural loops and are used by Oxford Molecular’s AbM antibody modeling software.

[0177] In some aspects, the CDRs can be “contact” CDRs. The “contact” CDRs are based on an analysis of the available complex crystal structures.

[0178] The residues from each of these CDRs are noted below.Loop _ Kabat _ AbM _ Chothia _ ContactLI L24-L34 L24-L34 L26-L32 L30-L36L2 L50-L56 L50-L56 L50-L52 L46-L55L3 L89-L97 L89-L97 L91-L96 L89-L96Hl H31-H35B H26-H35B H26-H32 H30-H35B (Kabat numbering) Hl H31-H35 H26-H35 H26-H32 H30-H35 (Chothia numbering) H2 H50-H65 H50-H58 H52-H56 H47-H58H3 H95-H102 H95-H102 H96-H101 H93-H101

[0179] CDRs can comprise “extended CDRs” as follows: 24-36 or 24-34 (LI), 46-56 or 50-56 (L2), and 89-97 or 89-96 (L3) in the VL, and 26-35 (Hl), 50-65 or 49-65 (H2), and 93-102, 94-102, or 95-102 (H3) in the VH. The variable-domain residues are numbered according to Kabat et al., supra, for each of these extended-CDR definitions.

[0180] CDRs can also be identified according to the IMGT numbering system as described in Lefranc M-P, (1999) The Immunologist 7: 132-136 and Lefranc M-P et al., (1999) Nucleic Acids Res 27: 209-212. According to the IMGT numbering scheme, VH-CDR1 is at positions 26 to 35, VH-CDR2 is at positions 51 to 57, VH-CDR3 is at positions 93 to 102, VL-CDR1 is at positions 27 to 32, VL-CDR2 is at positions 50 to 52, and VL-CDR3 is at positions 89 to 97.

[0181] The term “monoclonal” when referring to an antibody or antigen-binding fragment thereof refers to a homogeneous antibody or antigen-binding fragment population involved in the highly specific recognition and binding of a single antigenic determinant, or epitope. This is in contrast to polyclonal antibodies that typically include different antibodies directed against different antigenic determinants. The term “monoclonal” antibody or antigen-binding fragment thereof encompasses both intact and full-length monoclonal antibodies as well as antibody fragments (such as Fab, Fab’, F(ab’)2, Fv), single chain (scFv) mutants, fusion proteins or complexes comprising an antibody or antibody portion, and any other modified immunoglobulin molecule comprising an antigen recognition site. Furthermore, a “monoclonal” antibody or antigen-binding fragment thereof refers to such antibodies and antigen-binding fragments thereof made in any number of manners including but not limited to by hybridoma, phage selection, recombinant expression, and transgenic animals.

[0182] The term “chimeric” antibodies or antigen-binding fragments thereof refers to antibodies or antigen-binding fragments thereof wherein the amino acid sequence is derived from two or more species. Typically, the variable region of both light and heavy chains corresponds tothe variable region of antibodies or antigen-binding fragments thereof derived from one species of mammals (e.g., mouse, rat, rabbit, etc.) with the desired specificity, affinity, and capability, while the constant regions are homologous to the sequences in antibodies or antigen-binding fragments thereof derived from another (usually human) to avoid eliciting an immune response in that species.

[0183] The term “humanized” antibody or antigen-binding fragment thereof refers to forms of non-human (e.g., murine) antibodies or antigen-binding fragments that are specific immunoglobulin chains, chimeric immunoglobulins, or fragments thereof that contain minimal non-human (e.g., murine) sequences. Typically, humanized antibodies or antigen-binding fragments thereof are human immunoglobulins in which residues from the complementarity determining regions (CDRs) are replaced by residues from the CDRs of a molecule originating from a non-human species (e.g., mouse, rat, rabbit, hamster) that have the desired specificity, affinity, and capability (“CDR grafted”) (Jones et al., Nature 321:522-525 (1986); Riechmann et al., Nature 332:323-327 (1988); Verhoeyen et al., Science 239:1534-1536 (1988)). The humanized antibody or antigen-binding fragment thereof can be further modified by the substitution of additional residues either in the Fv framework region and / or within the replaced non-human residues to refine and optimize the specificity, affinity, and / or capability of the antibody or antigen-binding fragment thereof. In general, the humanized antibody or antigenbinding fragment thereof will comprise VH and VL that comprise substantially all of at least one, and typically two or three, of the CDR regions that correspond to the non-human immunoglobulin, whereas all or substantially all of the FR regions are those of a human immunoglobulin consensus sequence. The humanized antibody or antigen-binding fragment thereof can also comprise at least a portion of an immunoglobulin constant region or Fc region, typically that of a human immunoglobulin. Examples of methods used to generate humanized antibodies are described in U. S. Pat. 5,225,539; Roguska et al., Proc. Natl. Acad. Sci., USA, 91(3):969-973 (1994), and Roguska et al., Protein Eng. 9(10):895-904 (1996). In some aspects, a “humanized antibody” is a resurfaced antibody.

[0184] The term “human” antibody or antigen-binding fragment thereof means an antibody or antigen-binding fragment thereof having an amino acid sequence derived from a human immunoglobulin gene locus, where such antibody or antigen-binding fragment is made using any technique known in the art. This definition of a human antibody or antigen-binding fragment thereof includes intact or full-length antibodies and fragments thereof.

[0185] “Framework” or “FR” residues are those variable-domain residues other than the CDR residues as herein defined.

[0186] An “acceptor human framework” as used herein is a framework comprising the amino acid sequence of a VL or VH framework derived from a human immunoglobulin framework or a human consensus framework. An acceptor human framework “derived from” a human immunoglobulin framework or a human consensus framework can comprise the same amino acid sequence thereof, or it can comprise pre-existing amino acid sequence changes. In some aspects, the number of pre-existing amino acid changes are 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less. Where pre-existing amino acid changes are present in a VH, in some aspects those changes occur at only three, two, or one of positions 71H, 73H and 78H; for instance, the amino acid residues at those positions can be 71A, 73T and / or 78A. In some aspects, the VL acceptor human framework is identical in sequence to the VL human immunoglobulin framework sequence or human consensus framework sequence.

[0187] A “human consensus framework” is a framework that represents the most commonly occurring amino acid residues in a selection of human immunoglobulin VL or VH framework sequences. Generally, the selection of human immunoglobulin VL or VH sequences is from a subgroup of variable domain sequences. Generally, the subgroup of sequences is a subgroup as in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991). Examples include for the VL, the subgroup can be subgroup kappa I, kappa II, kappa III or kappa IV as in Kabat et al., supra. Additionally, for the VH, the subgroup can be subgroup I, subgroup II, or subgroup III as in Kabat et al., supra.

[0188] An “amino-acid modification” at a specified position, e.g., of an antibody of the present disclosure, refers to the substitution or deletion of the specified residue, or the insertion of at least one amino acid residue adjacent the specified residue. Insertion “adjacent” to a specified residue means insertion within one to two residues thereof. The insertion can be N-terminal or C-terminal to the specified residue. In some aspects, an amino acid modification is a substitution.

[0189] Antibody “effector functions” refer to those biological activities attributable to the Fc region (a native sequence Fc region or amino acid sequence variant Fc region) of an antibody and vary with the antibody isotype.

[0190] A “native sequence Fc region” comprises an amino acid sequence identical to the amino acid sequence of an Fc region found in nature. Native sequence human Fc regions include a native sequence human IgGl Fc region (non-A and A allotypes); native sequence human IgG2 Fcregion; native sequence human IgG3 Fc region; and native sequence human IgG4 Fc region as well.

[0191] A “variant Fc region” comprises an amino acid sequence which differs from that of a native sequence Fc region by virtue of at least one amino acid modification, in some aspects two or more amino acid substitution(s). In some aspects, the variant Fc region has at least one amino acid substitution compared to a native sequence Fc region or to the Fc region of a parent polypeptide, e.g., from about one to about ten amino acid substitutions, and in some aspects from about one to about five amino acid substitutions compared to a native sequence Fc region or in the Fc region of the parent polypeptide. In some aspects, the variant Fc region possesses at least 80% homology with a native sequence Fc region and / or with an Fc region of a parent polypeptide, at least 90% homology therewith, or at least 95% homology therewith.

[0192] “Fc receptor” or “FcR” describes a receptor that binds to the Fc region of an antibody. In some aspects, an FcR is a native sequence human FcR. In some aspects, a FcR is one which binds an IgG antibody (a gamma receptor) and includes receptors of the FcyRI, FcyRII, and FcyRIII subclasses, including allelic variants and alternatively spliced forms of these receptors, FcyRII receptors include FcyRIIA (an “activating receptor”) and FcyRIIB (an “inhibiting receptor”), which have similar amino acid sequences that differ primarily in the cytoplasmic domains thereof. Activating receptor FcyRIIA contains an immunoreceptor tyrosine-based activation motif (“IT AM”) in its cytoplasmic domain. Inhibiting receptor FcyRIIB contains an immunoreceptor tyrosine-based inhibition motif (“ITIM”) in its cytoplasmic domain. Other FcRs, including those to be identified in the future, are encompassed by the term “FcR” herein. FcRs can also increase the serum half-life of antibodies.

[0193] “Binding affinity” generally refers to the strength of the sum total of non-covalent interactions between a single binding site of a molecule (e.g., an antibody or antigen -binding fragment thereof) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, “binding affinity” refers to intrinsic binding affinity which reflects a 1: 1 interaction between members of a binding pair (e.g., antibody or antigen-binding fragment thereof and antigen). The affinity of a molecule X for its partner Y can generally be represented by the equilibrium dissociation constant (KD). Affinity can be measured and / or expressed in a number of ways known in the art, including, but not limited to, equilibrium dissociation constant (KD), and equilibrium association constant (KA). The KD is calculated from the quotient of koff / kon, whereas KA is calculated from the quotient of kon / koff. konrefers to the association rate constantof, e.g., an antibody or antigen-binding fragment thereof to an antigen, and koffrefers to the dissociation rate constant of, e.g., an antibody or antigen-binding fragment thereof from an antigen. The konand koffcan be determined by techniques known to one of ordinary skill in the art, such as BIAcore® or KinExA. Dissociation constants may also be determined through any analytical technique, including any biochemical or biophysical technique such as ELISA, surface plasmon resonance (SPR), bio-layer interferometry (see, e.g., Octet System by ForteBio), isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), circular dichroism (CD), stopped-flow analysis, and colorimetric or fluorescent protein melting analyses. (See, e.g., Estep etal, (2013) MAbs 5(2):270-8.)

[0194] With regard to the binding of an antibody to a target molecule, the term “specific binding” or “specifically binds” or is “specific for” a particular polypeptide or an epitope on a particular polypeptide target means binding that is measurably different from a non-specific interaction. Specific binding can be measured, for example, by determining binding of a molecule compared to binding of a control molecule. For example, specific binding can be determined by competition with a control molecule that is similar to the target, for example, an excess of non-labeled target. In this case, specific binding is indicated if the binding of the labeled target to a probe is competitively inhibited by excess unlabeled target. The term “specific binding” or “specifically binds” or is “specific for” a particular polypeptide or an epitope on a particular polypeptide target as used herein can be exhibited, for example, by a molecule having a KD for the target of about any of 10'4M or lower, 10'5M or lower, 10'6M or lower, 10'7M or lower, 10'8M or lower, 10'9M or lower, IO'10M or lower, 10'11M or lower, 10'12M or lower or a KD in the range of 10'4M to 10'6M or 10'6M to IO'10M or 10'7M to 10'9M. As will be appreciated by the skilled artisan, affinity and KD values are inversely related. A high affinity for an antigen is measured by a low KD value. In some aspects, the term “specific binding” refers to binding where a molecule binds to a particular polypeptide or epitope on a particular polypeptide without substantially binding to any other polypeptide or polypeptide epitope.

[0195] The term “linker” or “linked” refers to the covalent linkage between two polypeptides or two heterologous molecules. In some aspects, a linker is a chemical linker. In some aspects, the linker comprises a peptide bond, and the two polypeptides or two heterologous molecules are linked to each other either directly to or via one or more additional amino acids. A glycine linker is one that comprises one or more glycines, but no other amino acids, e.g., GGGG (SEQ ID NO: 204). A glycine-rich linker is one that comprises one or more glycines and can contain otheramino acids as long as glycine is the predominant species in the linker e.g., GGGNGG (SEQ ID NO: 205), wherein N is any amino acid. A glycine-serine linker is one which contains both glycine and serine in any proportion, e.g., GGGS (SEQ ID NO: 206). Similarly, a proline linker is one that comprises one or more prolines but no other amino acids. A proline-rich linker is one that comprises one or more prolines and can contain other amino acids so long as proline is the predominant species in the linker.

[0196] As used herein, “percent (%) amino acid sequence identity” and “homology” with respect to a peptide, polypeptide or antibody sequence refers to the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the specific peptide or polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as identical matches. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, ALIGN or MEGALIGN™ (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms known in the art needed to achieve maximal alignment over the full-length of the sequences being compared.

[0197] A polypeptide, antibody, polynucleotide, vector, cell, or composition which is “isolated” is a polypeptide, antibody, polynucleotide, vector, cell, or composition which is in a form not found in nature. Isolated polypeptides, antibodies, polynucleotides, vectors, cells or compositions include those which have been purified to a degree that they are no longer in a form in which they are found in nature. In some aspects, an antibody, polynucleotide, vector, cell, or composition which is isolated is substantially pure.

[0198] As used herein, “substantially pure” refers to material which is at least 50% pure (i.e., free from contaminants), at least 90% pure, at least 95% pure, at least 98% pure, or at least 99% pure.

[0199] The term “expression system” refers to one or more nucleic acid molecules comprising coding sequence and control sequence(s) in operable linkage, along with a host cell and / or other in vitro transcription and translation machinery, such that one or more proteins encoded by the nucleic acid molecule(s) are capable of being produced.

[0200] The term “vector,” as used herein, is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. One type of vector is a“plasmid,” which refers to a circular double stranded DNA into which additional DNA segments can be ligated. Another type of vector is a phage vector. Another type of vector is a viral vector, wherein additional DNA segments can be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can be integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operatively linked. Such vectors are referred to herein as “recombinant expression vectors,” or simply, “expression vectors.” In general, expression vectors of utility in recombinant DNA techniques are often in the form of plasmids. In the present specification, “plasmid” and “vector” can be used interchangeably as the plasmid is the most commonly used form of vector.

[0201] “Polynucleotide,” or “nucleic acid,” as used interchangeably herein, refer to polymers of nucleotides of any length, and include DNA and RNA. The nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and / or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase or by a synthetic reaction.

[0202] A “host cell” includes an individual cell or cell culture that can be or has been a recipient for vector(s) for incorporation of polynucleotide inserts. Host cells include progeny of a single host cell, and the progeny may not necessarily be completely identical (in morphology or in genomic DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation. A host cell includes cells transfected in vivo with a polynucleotide(s) of this disclosure. In some aspects, the host cell is an isolated host cell.

[0203] “ Carriers” as used herein include pharmaceutically acceptable carriers, excipients, or stabilizers that are nontoxic to the cell or mammal being exposed thereto at the dosages and concentrations employed.

[0204] As used herein, the term “treatment” refers to clinical intervention designed to alter the natural course of the individual being treated during the course of clinical pathology. Desirable effects of treatment include decreasing the rate of progression, ameliorating or palliating the pathological state, and remission or improved prognosis of a particular disease, disorder, or condition. An individual is successfully “treated”, for example, if one or more symptoms associated with a particular disease, disorder, or condition are mitigated or eliminated.

[0205] The terms “administer,” “administering,” “administration,” and the like, as used herein, refer to methods that can be used to deliver a drug, e.g., an anti-human antibody or antigenbinding fragment thereof, to the desired site of biological action.

[0206] An “effective amount” refers to at least an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic result. An effective amount can be provided in one or more administrations. An effective amount is also one in which any toxic or detrimental effects of the treatment are outweighed by the therapeutically beneficial effects. For therapeutic use, beneficial or desired results include clinical results such as decreasing one or more symptoms resulting from the disease, increasing the quality of life of those suffering from the disease, decreasing the dose of other medications required to treat the disease, enhancing effect of another medication such as via targeting, delaying the progression of the disease, and / or prolonging survival. An effective amount of drug, compound, or pharmaceutical composition is an amount sufficient to accomplish therapeutic treatment either directly or indirectly. As is understood in the clinical context, an effective amount of a drug, compound, or pharmaceutical composition may or may not be achieved in conjunction with another drug, compound, or pharmaceutical composition. Thus, an “effective amount” can be considered in the context of administering one or more therapeutic agents, and a single agent can be considered to be given in an effective amount if, in conjunction with one or more other agents, a desirable result can be or is achieved.

[0207] As used herein, the terms “subject” and “patient” are used interchangeably. The subject can be a mammal such as a non-human animal (e.g., cow, pig, horse, cat, dog, rat, mouse, monkey or other primate, etc.). In some aspects, the subject is a cynomolgus monkey. In some aspects, the subject is a human.

[0208] As used herein, administration “in conjunction” or “in combination” with another compound or composition includes simultaneous administration and / or administration at different times. Administration in conjunction also encompasses administration as a coformulation or administration as separate compositions, including at different dosing frequencies or intervals, and using the same route of administration or different routes of administration. In some aspects, administration in conjunction is administration as a part of the same treatment regimen.

[0209] As used herein, a “complex” refers to one or more proteins comprising connected parts. The parts can be connected e.g., via a peptide bond (e.g., in a fusion protein or complex), a linker(e.g., a peptide linker), or via noncovalent protein-protein interactions such as disulfide bonds (e.g., in an antibody). Exemplary parts that can be included in a complex include an antigenbinding domain that specifically binds to N3pE Ap and an antigen-binding domain that specifically binds to CD98hc or TfR, an Fc region, and / or an Fc domain. Accordingly, nonlimiting examples of a “complex” comprising an anti-N3pE Ap antigen-binding domain and an anti-CD98hc or anti-TfR antigen-binding domain include (a) a fusion protein or complex comprising the anti-pyroGlu3 AP antigen-binding domain and the anti-CD98hc or anti-TfR antigen-binding domain in a single polypeptide chain and (b) four proteins connected via noncovalent protein-protein interactions, wherein the first protein contains a heavy chain of anti-pyroGlu3 AP antibody comprising an anti-pyroGlu3 AP antigen-binding domain, the second protein contains a heavy chain of the anti-pyroGlu3 AP fused to an scFv containing an anti-CD98hc or anti-TfR scFv containing an anti-CD98hc or anti-TfR antigen-binding domain, and the third and fourth proteins contain light chains of the anti-pyroGlu3 AP antibody (e.g., as shown in Figure 1). Formats of other exemplary complexes are provided in Figures 1-3.

[0210] As used herein, the term “composition” refers to pharmaceutical combination of components. The components can be connected to each other via covalent bonds or non-covalent bonds or can merely be contained in the same mixture or solution. Accordingly, a “composition” comprising an antigen-binding domain can refer, e.g., to an antibody, scFv, or Fab comprising the antigen-binding domain, can refer to a fusion protein or complex comprising the antigenbinding domain, or can refer to a mixture of solution comprising such an antigen-binding domain, antibody, scFv, Fab, fusion protein or complex. In some aspects, a composition is a “pharmaceutical composition.”

[0211] As used herein the term “pharmaceutical composition” refers to a combination comprising an active agent (e.g., an antigen-binding domain, an antibody, a scFv, or Fab comprising the antigen-binding domain disclosed herein) with at least one inert pharmaceutically acceptable agent (e.g., an excipient or a carrier).

[0212] As used herein, the terms “about” and “approximately,” when used to modify a numeric value or numeric range, indicate that deviations of up to 10% above and down to 10% below the value or range remain within the intended meaning of the recited value or range. It is understood that wherever aspects are described herein with the language “about” or “approximately” a numeric value or range, otherwise analogous aspects referring to the specific numeric value or range are also provided.

[0213] As used herein and in the appended claims, the singular forms “a,” “an,” and “the” include plural reference unless the context clearly indicates otherwise. For example, reference to an “antibody” is a reference to from one to many antibodies, such as molar amounts, and includes equivalents thereof known to those skilled in the art, and so forth.

[0214] It is understood that wherever aspects are described herein with the language “comprising,” otherwise analogous aspects described in terms of “consisting of’ and / or “consisting essentially of’ are also provided. In this disclosure, “comprises,” “comprising,” “containing” and “having” and the like can mean “includes,” “including,” and the like;“consisting essentially of’ or “consists essentially of’ are open-ended, allowing for the presence of more than that which is recited so long as basic or novel characteristics of that which is recited is not changed by the presence of more than that which is recited, but excludes prior art aspects.

[0215] All references cited herein, including patent applications and publications, are hereby incorporated by reference in their entirety.

[0216] The present disclosure will be more fully understood by reference to the following Examples. They should not, however, be construed as limiting the scope of the present disclosure. All citations throughout the disclosure are hereby expressly incorporated by reference.Antigen-Binding Domains and Compositions Comprising Antigen-Binding Domains

[0217] In some aspects, compositions provided herein may comprise an antigen-binding domain.

[0218] In some aspects, compositions provided herein may comprise a fusion protein or complex. In some aspects, the fusion protein or complex comprises an antigen-binding domain.

[0219] In some aspects, compositions provided herein may comprise a multi-specific protein. In some aspects, the multi-specific protein comprises an antigen-binding domain.

[0220] In some aspects, compositions provided herein may comprise an antibody or an antibody fragment thereof.

[0221] In some aspects, compositions provided herein may comprise an antigen-binding fragment. Antigen-binding fragments of antibodies include, but are not limited to, Fab, Fab’, Fab’-SH, F(ab’)2, Fv, and scFv fragments, and other fragments described below. For a review of certain antibody fragments, see Hudson et al. Nat. Med. 9:129-134 (2003). For a review of scFv fragments, see, e.g., WO 93 / 16185; and U. S. Patent Nos. 5,571,894 and 5,587,458. Fordiscussion of Fab and F(ab’)2 fragments comprising salvage receptor binding epitope residues and having increased in vivo half-life, see U. S. Patent No. 5,869,046.

[0222] Diabodies are antibody fragments with two antigen-binding sites that can be bivalent and / or bispecific. See, for example, EP404097; WO 1993 / 01161; Hudson et al. Nat. Med. 9:129- 134 (2003). Triabodies and tetrabodies are also described in Hudson et al. Nat. Med. 9:129-134 (2003). Single-domain antibodies are antibody fragments comprising all or a portion of the heavy chain variable domain or all or a portion of the light chain variable domain of an antibody. In some aspects, a single-domain antibody is a human single-domain antibody (see, e.g., U. S. Patent No. 6,248,516).

[0223] Antibody fragments can be made by various techniques, including but not limited to proteolytic digestion of an intact antibody as well as production by recombinant host cells (e.g., E. coli or phage), as described herein.

[0224] As provided herein, a composition comprising an antigen-binding domain provided herein can be chimeric. Certain chimeric antibodies are described, e.g., in U. S. Patent No.4,816,567. In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate, such as a monkey) and a human constant region. In a further example, a chimeric antibody is a “class switched” antibody in which the class or subclass has been changed from that of the parent antibody.

[0225] As provided herein, a composition comprising an antigen-binding domain provided herein can be humanized. Typically, a non-human antibody is humanized to reduce immunogenicity to humans, while retaining the specificity and affinity of the parental non-human antibody. In some aspects, a humanized antibody is substantially non-immunogenic in humans. In some aspects, a humanized antibody has substantially the same affinity for a target as an antibody from another species from which the humanized antibody is derived. See, e.g., U. S. Pat. No. 5,530,101; 5,693,761; 5,693,762; and 5,585,089. In some aspects, amino acids of an antibody variable domain that can be modified without diminishing the native affinity of the antigen-binding domain while reducing its immunogenicity are identified. See, e.g., U. S. Pat. Nos. 5,766,886 and 5,869,619. Generally, a humanized antibody comprises one or more variable domains in which CDRs (or portions thereof) are derived from a non-human antibody, and framework regions (FRs) (or portions thereof) are derived from human antibody sequences. A humanized antibody can comprise at least a portion of a human constant region. In some aspects,some FR residues in a humanized antibody are substituted with corresponding residues from a non-human antibody (e.g., the antibody from which the CDR residues are derived), for example, to restore or improve antibody specificity or affinity.

[0226] Humanized antibodies and methods of making them are reviewed, for example, in Almagro et al. Front. Biosci. 13:161 9-1633 (2008), and are further described, e.g., in U. S. Patent Nos. 5,821,337; 7,527,791; 6,982,321; and 7087409. Human framework regions that can be used for humanization include but are not limited to: framework regions selected using the “best- fit” method (see, e.g., Sims et al. J. Immunol. 151:2296 (1993)); framework regions derived from the consensus sequence of human antibodies of a particular subgroup of light or heavy chain variable regions (see, e.g., Carter et al. Proc. Natl. Acad. Set. USA 89:4285 (1992); and Presta et al., J. Immunol. 151:2623 (1993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson Front. Biosci. 13: 1619-1633 (2008)); and framework regions derived from screening FR libraries (see, e.g., Baca et al. J. Biol. Chem. 272:10678-10684 (1997) and Rosok et al. J. Biol. Chem. 271:22611-22618 (1996)).

[0227] As provided herein, a composition comprising an antigen-binding domain provided herein can be human. Human antibodies can be produced using various techniques known in the art. Human antibodies are described generally in van Dijk et al. Curr. Opin. Pharmacol. 5:368-74 (2001) andLonberg Curr. Opin. Immunol. 20:450-459 (2008).

[0228] Human antibodies can be prepared by administering an immunogen to a transgenic animal that has been modified to produce intact human antibodies or intact antibodies with human variable regions in response to antigenic challenge. One can engineer mouse strains deficient in mouse antibody production with large fragments of the human Ig loci in anticipation that such mice would produce human antibodies in the absence of mouse antibodies. Large human Ig fragments can preserve the large variable gene diversity as well as the proper regulation of antibody production and expression. By exploiting the mouse machinery for antibody diversification and selection and the lack of immunological tolerance to human proteins, the reproduced human antibody repertoire in these mouse strains can yield high affinity fully human antibodies against any antigen of interest, including human antigens. Using the hybridoma technology, antigen-specific human mAbs with the desired specificity can be produced and selected. Certain exemplary methods are described in U. S. Pat. No. 5,545,807, EP546073, andEP546073. See also, for example, U. S. Patent Nos. 6,075,181 and 6,150,584 describing XENOMOUSE™ technology; U. S. Patent No. 5,770,429 describing HUMAB®technology; U. S. Patent No. 7,041,870 describing K-M MOUSE® technology, and U. S. Patent Application Publication No. US 2007 / 0061900, describing VELOCIMOUSE® technology.Human variable regions from intact antibodies generated by such animals can be further modified, e.g., by combining with a different human constant region.

[0229] Human antibodies can also be made by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, e.g., Kozbor J. Immunol. 133:3001 (1984) and Boemer et al. J.Immunol. 147:86 (1991)). Human antibodies generated via human B-cell hybridoma technology are also described in Li et al. Proc. Natl. Acad. Sci. USA, 1 03:3557-3562 (2006). Additional methods include those described, for example, in U. S. Patent No. 7,189,826 (describing production of monoclonal human IgM antibodies from hybridoma cell lines). Human hybridoma technology (Trioma technology) is also described in Vollmers et al. Histology and Histopathology 20(3):927-937 (2005) and Vollmers et al. Methods and Findings in Experimental and Clinical Pharmacology 27(3): 185-91 (2005). Human antibodies can also be generated by isolating Fv clone variable domain sequences selected from human-derived phage display libraries. Such variable domain sequences can then be combined with a desired human constant domain. Techniques for selecting human antibodies from antibody libraries are described below.

[0230] In some aspects, an antibody is a human antibody isolated by in vitro methods and / or screening combinatorial libraries for antibodies with the desired activity or activities. Suitable examples include but are not limited to phage display (CAT, Morphosys, Dyax, Biosite / Medarex, Xoma, Symphogen, Alexion (formerly Proliferon), Affimed) ribosome display (CAT), yeast display (Adimab), and the like. In certain phage display methods, repertoires of VH and VL genes are separately cloned by polymerase chain reaction (PCR) and recombined randomly in phage libraries, which can then be screened for antigen-binding phage as described in Winter et al. Ann. Rev. Immunol. 12: 433-455 (1994). For example, a variety of methods are known in the art for generating phage display libraries and screening such libraries for antibodies possessing the desired binding characteristics. See also Sidhu et al. J. Mol. Biol. 338(2): 299-310, 2004; Lee et al. J. Mol. Biol. 340(5): 1073-1093, 2004; Fellouse Proc. Natl. Acad. Sci. USA 101(34): 12467- 12472 (2004); and Lee et al. J. Immunol. Methods 284(2): 1 19-132 (2004). Phage typically display antibody fragments, either as single-chain Fv (scFv) fragments or as Fab fragments. Libraries from immunized sources provide high-affinity antibodies to the immunogen without the requirement of constructing hybridomas. Alternatively, the naive repertoire can be cloned (e.g.,from human) to provide a single source of antibodies to a wide range of non-self and also selfantigens without any immunization as described by Griffiths et al. EMBO J. 12: 725-734 (1993). Finally, naive libraries can also be made synthetically by cloning unrearranged V-gene segments from stem cells, and using PCR primers comprising random sequence to encode the highly variable CDR3 regions and to accomplish rearrangement in vitro, as described by Hoogenboom et al. J. Mol. Biol., 227: 381-388, 1992. Patent publications describing human antibody phage libraries include, for example: U. S. Patent No. 5,750,373, and U. S. Patent Publication Nos. 2007 / 0292936 and 2009 / 0002360. Antibodies isolated from human antibody libraries are considered human antibodies or human antibody fragments herein.Fc Domains and Regions

[0231] A composition provided herein can comprise an Fc domain or region or fragment thereof. In some aspects, an Fc domain or region is of IgG class, the IgM class, or the IgA class. In some aspects, an Fc domain or region or fragment thereof is an IgG Fc domain or region or fragment thereof. In some aspects, an Fc domain or region or fragment thereof is a human IgG Fc domain or region or fragment thereof. In some aspects, an Fc domain or region or fragment thereof is a human IgGl Fc domain or region or fragment thereof. In some aspects, an Fc domain or region or fragment thereof is a human IgG2 Fc domain or region or fragment thereof. In some aspects, an Fc domain or region or fragment thereof is a human IgG4 Fc domain or region or fragment thereof.

[0232] In some aspects provided herein, a composition provided herein comprises a modified Fc domain or region or fragment thereof. In some aspects, the modified Fc domain or region or fragment thereof is a modified IgGl Fc comprising one or more modifications. For example, in some aspects, the IgGl modified Fc comprises one or more amino acid substitutions (e.g., relative to a wild-type Fc domain of the same isotype).

[0233] In some aspects, the Fc domain or region or fragment thereof is a wildtype IgG2. In some aspects, the modified Fc domain or region or fragment thereof is a modified IgG2 Fc comprising one or more modifications. For example, in some aspects, the modified IgG2 Fc comprises one or more amino acid substitutions (e.g., relative to a wild-type IgG2 Fc).

[0234] In some aspects, the Fc domain or region or fragment thereof is a wildtype IgG4. In some aspects, the modified Fc domain or region or fragment thereof is a modified IgG4 Fc comprising one or more modifications. For example, in some aspects, the modified IgG4 Fc comprises one or more amino acid substitutions (e.g., relative to a wild-type IgG4 Fc). In someaspects, the one or more amino acid substitutions in the modified IgG4 Fc are IgG4-S228P or IgG4-S228P / L235E, where the amino acid position is according to the EU numbering convention.

[0235] In some aspects, the Fc domain or region or fragment thereof is a human IgGl and the one or more amino acid substitutions are selected from N297A (Bolt S et al. (1993) Eur J Immunol 23:403-411), D265A (Shields et al. (2001) A. J. Biol. Chem. 276, 6591-6604), L234A, L235A (Hutchins et al. (1995) Proc Natl Acad Sci USA, 92: 11980-11984; Alegre et al., (1994) Transplantation 57:1537-1543. 31; Xu et al., (2000) Cell Immunol, 200:16-26), G237A (Alegre et al. (1994) Transplantation 57:1537-1543. 31; Xu et al. (2000) Cell Immunol, 200:16-26), C226S, C229S, E233P, L234V, L234F, L235E (McEarchern et al., (2007) Blood, 109: 1185- 1192), P331S (Sazinsky et al., (2008) Proc Natl Acad Sci USA 2008, 105:20167-20172), K322A (Hezareh et al. (2001) J Virol. 75(24) 12161-12168), S267E, L328F, A330L, M252Y, S254T, E430G, and / or T256E, where the amino acid position is according to the EU numbering convention. In some aspects, the IgGl Fc comprises the amino acid substitutions L234A, L235A, and P329S (LALAPS) according to EU numbering. In some aspects, the IgGl Fc comprises the amino acid substitutions L234A, L235A, and P331S (LALAPSv2) according to EU numbering. In some aspects, the IgGl Fc comprises the amino acid substitutions L234A, L235A, and P329G (LALAPG) according to EU numbering. In some aspects, the IgGl Fc comprises the amino acid substitutions L234A, L235A, and P331G (LALAPGv2) according to EU numbering. In some aspects, the IgGl Fc comprises N325S and L328F mutations according to EU numbering. In some aspects, the IgGl Fc comprises P329G or P329S according to EU numbering. In some aspects, the IgGl Fc comprises K322A according to EU numbering.TABLE 1: Exemplary Fc DomainsFc mutations Description(according to EUnumbering)huIgGl WT Human IgGl Fc ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSW NSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYIC NVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVF LFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDG VEVHNAI< TI< PREEQYNSTYRVVSVLTVLHQDWLNGI< EYI< CKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQ VSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGS FFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 211)Fc mutations Description(according to EUnumbering)huIgGl NSLF (N325S, Both polypeptides of huIgGl Fc contain N325S and L328F L328F) mutations, which decreases binding to FcGR3a and increase binding to FcGR2ahuIgGl LALAPS (L234A, Both polypeptides of huIgGl Fc contain L234A, L235A, and L235A, P329S) P329S mutations, which minimize effector function (FcGR2a, FcgR3a and complement binding)huIgGl LALAPSv2 (L234A, Both polypeptides of huIgGl Fc contain L234A, L235A, and L235A, P331S) P331S mutations, which minimize effector function (FcGR2a, FcgR3a and complement binding)huIgGl LALA (L234A, Both polypeptides of huIgGl Fc contain L234A, L235A L235A) mutations, which minimize effector function (FcGR2a, FcgR3a binding)huIgGl K322A Both polypeptides of huIgGl Fc contain K332A mutations, which minimizes complement activated effector function.huIgGl YTE (M252Y, Both polypeptides of huIgGl Fc contain M252Y, S254T, T256E S254T, T256E) mutations, which enhance affinity to FcRn to extend antibody halflifehuIgGl LS (M428L, Both huIgGl Fc contain M428L, N434S mutations.N434S) This enhances affinity to FcRn to extend antibody half-life huIgGl LALAPS (L234A, One huIgGl Fc polypeptide contains L234A, L235A, and P329S L235A, P329S) / huIgGl mutations. The other huIgGl Fc polypeptide is wild-type.WT This tunes effector function activity (FcgR2, FcgR3a and complement binding)huIgGl LALAPSv2 (L234A, One huIgGl Fc polypeptide contains L234A, L235A, and P331S L235A, P331S) / huIgGl mutations. The other huIgGl Fc polypeptide is wild-type.WT This tunes effector function activity (FcgR2, FcgR3a and complement binding)huIgGl LALA (L234A, One huIgGl Fc polypeptide contains L234A and L235A L235A) / huIgGl WT mutations.The other huIgGl Fc polypeptide is wild-type.This tunes effector function activity (FcgR2, and FcgR3a binding) HuIgGl LALA (L234A, One huIgGl Fc polypeptide contains L234A and L235A L235A) / hu IgGl PS mutations.(P329S) The other huIgGl Fc polypeptide contains a P329S mutation.This tunes effector function activity (FcgR2, FcgR3a and complement binding).HuIgGl LALA (L234A, One huIgGl Fc polypeptide contains L234A and L235A L235A) / huIgGl PSv2 mutations.(P331S) The other huIgGl Fc polypeptide contains a P329S mutation.This tunes effector function activity (FcgR2, FcgR3a and complement binding).huIgG2 WT Human IgG2 FcASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFGTQTYTFc mutations Description(according to EUnumbering)CNVDHKPSNTKVDKTVERKCCVECPPCPAPPVAGPSVFLFP PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEV HNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKV SNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSL TCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 212)huIgG4 WT Human IgG4 Fc ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSW NSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTC NVDHKPSNTKVDKRVESKYGPPCPSCPAPEFLGGPSVFLFPP KPKDTLMISRTPEVTCWVDVSQEDPEVQFNWYVDGVEVH NAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVS NKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLT CLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLY SRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK(SEQ ID NO: 213)huIgG4 S228P Both human IgG4 Fc polypeptides contain S228P mutations.This minimizes Fab arm exchange present in wt IgG4 Fc huIgG4 S228P / L235E Both human IgG4 Fc polypeptides contain S228P and L235E mutations.This minimizes Fab arm exchange present in wt IgG4 Fc andminimizes effector function (FcGR2a, FcgR3a binding).

[0236] In some aspects provided herein, a composition provided herein comprises one or more mutations to promote heterodimerization of Fc domains. In some aspects, a Fc region of a bispecific complex provided herein is formed by Fc domains that contain amino acid mutations, substitutions, additions, or deletions to promote heterodimerization in which different polypeptides comprising different Fc domains can dimerize to yield a heterodimer configuration. In some aspects, a bispecific of the present disclosure comprises a first Fc sequence comprising a first CH3 region, and a second Fc sequence comprising a second CH3 region, wherein the sequences of the first and second CH3 regions are different and are such that the heterodimeric interaction between said first and second CH3 regions is stronger than each of the homodimeric interactions of said first and second CH3 regions.

[0237] Methods to promote heterodimerization of Fc domains include amino acid deletions, additions, or substitutions of the amino acid sequence of the Fc domain, such as by including a set of “knob-into-hole” deletions, additions, or substitutions or including amino acid deletions,additions, or substitutions to effect electrostatic steering of the Fc to favor attractive interactions among different polypeptide chains. Methods for promoting heterodimerization of complementary Fc polypeptides have been previously described in, for example, Ridgway et al, 1996, Protein Eng, 9:617-621; Merchant et al, 1998, Nature Biotechnol, 16:677-681; Moore et al, 2011, MAbs, 3:546-557; Von Kreudenstein et al, 2013, 5:646-654; Gunasekaran et al, 2010, J Biol Chem, 285:19637-19464; Leaver-Fay et al, 2016, Structure, 24:641-651; Ha et al, 2016, Frontiers in Immunology, 7:1; Davis et al, 2010, Protein Eng Des Sei, 23:195-202; PCT Pub. Nos. W01996 / 027011; WO 1998 / 050431; W02006 / 028936; W02009 / 089004;WO2011 / 143545; WO2014 / 067011; WO2012 / 058768; WO2018 / 027025; US Pub. Nos.US2014 / 0363426; US2015 / 0307628; US2018 / 0016354; US2015 / 0239991; US2017 / 0058054; U. S. Pat. Nos. 5,731,168; 7,183,076; 9,701,759; 9,605,084; 9,650,446; 8,216,805; 8,765,412; and 8,258,268.

[0238] In some aspects, complementary Fc polypeptides of an Fc heterodimer include a mutation to alter charge polarity across the Fc dimer interface such that co-expression of electrostatically matched Fc domains support favorable attractive interactions, thereby promoting desired Fc heterodimer formation; whereas unfavorable repulsive charge interactions suppress unwanted Fc homodimer formation (Guneskaran et al, 2010, J Biol Chem, 285:19637-19646). When co-expressed in a cell, association between the polypeptide chains is possible but the chains do not substantially self-associate due to charge repulsion.

[0239] “Knob-hole” or “knob-into-hole” configurations are complementary Fc polypeptides of an Fc heterodimer that promote heterodimerization of two Fc polypeptides. “Knob-into-hole” technology is described in U. S. Pat. Nos. 5,731,168; 7,695,936; 8,216,805; 8,765,412; Ridgway et al., ProtEng 9, 617-621 (1996); and Carter, J Immunol Meth 248, 7-15 (2001). Generally, the method involves introducing a protuberance (“knob”) at the interface of a first polypeptide and a corresponding cavity (“hole”) in the interface of a second polypeptide, such that the protuberance can be positioned in the cavity so as to promote heterodimer formation and hinder homodimer formation. Protuberances are constructed by replacing small amino acid side chains from the interface of the first polypeptide with larger side chains (e.g., tyrosine or tryptophan).Compensatory cavities of identical or similar size to the protuberances are created in the interface of the second polypeptide by replacing large amino acid side chains with smaller ones (e.g., alanine or threonine). The protuberance and cavity can be made by altering the nucleic acid encoding the polypeptides, e.g., by site-specific mutagenesis, or by peptide synthesis.

[0240] In some aspects, a composition provided herein comprises a Fc dimer comprising a “knob” mutation in one Fc domain and a “hole” mutation in the other Fc domain. In some aspects, the “knob” mutation comprises the amino acid substitution T366W according to EU numbering. In some aspects, the “hole” mutation comprises the amino acids substitutions T366S, L368A, and Y407V according to EU numbering. In some aspects, the “knob” mutation comprises the amino acid substitution T366W in one of the two subunits of the Fc dimer, and the “hole” mutation comprises the amino acid substitutions T366S, L368A and Y407V in the other subunit of the Fc dimer. In some aspects, the subunit of the Fc dimer comprising the “knob” mutation additionally comprises the amino acid substitution S354C, and the subunit of the Fc dimer comprising the “hole” mutation additionally comprises the amino acid substitution Y349C. Introduction of these two cysteine residues results in the formation of a disulfide bridge between the two subunits of the Fc dimer, thus further stabilizing the dimer (Carter, J Immunol Methods 248, 7-15 (2001)). Thus, in such configurations, a first Fc polypeptide comprises amino acid modifications to form the “knob” and a second Fc polypeptide comprises amino acid modifications to form the “hole” thus forming an Fc heterodimer comprising complementary Fc polypeptides.

[0241] Exemplary paired amino acid modifications of complementary Fc polypeptides of an Fc heterodimeric configuration are set forth below in Table 2 (EU numbering). In some aspects, the Fc polypeptide is a wildtype human IgGl Fc having one Fc polypeptide comprising a knob configuration and one Fc polypeptide comprising a hole configuration, e.g., as set forth:ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS GLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGG PSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQY NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSR DELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (knob: SEQ ID NO:267) and ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS GLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGG PSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQY NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSR DELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (hole; SEQ ID NO: 268), respectively.TABLE 2: Exemplary paired Fc modifications for heterodimeric Fc domains First Fc polypeptide Second Fc polypeptideT366Y Y407TT366W T366S / L368W / Y407VT366W T366S / L368A / Y407VT366W / S354C T366S / L368A / Y407V / Y349CT350 V / L351 Y / F405 A / Y407 V T350 V / T366L / K392L / T394WK360D / D399M / Y407 A E345R / Q347R / T366V / K409V K409D / K392D D399K / E356KK360E / K409W Q347R / D399V / F405T L360E / K409W / Y349C Q347R / D399V / F405T / S354C K370E / K409W E357N / D399V / F405TExemplary Anti-PyroGlu3 Abeta (A ) Antibodies

[0242] The terms “anti-pyroGlu3 Abeta (A0) antibody,” “anti-pyroGlu3 Abeta (AP),” “antibody that binds to pyroGlu3 Abeta (AP),” used interchangeably herein, refer to an antibody that is capable of binding pyroGlu3 Abeta (AP)with sufficient affinity such that the antibody is useful as a diagnostic and / or therapeutic agent in targeting Abeta (AP).

[0243] Provided herein are antibodies to pyroGlu3 Abeta (AP). Anti-pyroGlu3 Abeta (AP) antibodies of the present disclosure provided herein are useful, e.g., for the treatment of disorders associated with amyloid beta, such as, for example, Alzherimer’s disease.

[0244] Table 3 below lists the heavy chain CDR amino acid sequences (according to Kabat numbering) of anti-pyroGlu3 Abeta (AP) antibodies of the present disclosure. Table 4 below lists the light chain CDR amino acid sequences (according to Kabat numbering) of anti-pyroGlu3 Abeta (AP) antibodies of the present disclosure. Table 5 below lists the heavy chain variable region and light chain variable region amino acid sequences of anti-pyroGlu3 Abeta (AP) antibodies of the present disclosure.TABLE 3. Heavy Chain CDR Sequences of Anti-pyroGlu3 Abeta (A ) AntibodiesAntibody CDR-H1 CDR-H2 CDR-H3AB-1 SGSYYWG SFYYSGTIYYNPSLKS QREYS S S SNYYYYYMD V(SEQ ID NO: 4) (SEQ ID NO: 5) (SEQ ID NO: 6)AB-2 SSSYYWG(SE SIYFSGSTYYNPSLKS HREMATIRDYHYYYMDV Q ID NO: 7) (SEQ ID NO: 8) (SEQ ID NO: 9)AB-5 DYAMH (SEQ GISWNNNNIDYADSVK DREYSGSYYDAFDI (SEQ ID ID NO: 10) G (SEQ ID NO: 11) NO: 12)AB-3 SNYMS (SEQ VIYSGGTKHYADSVKG SYYYGSGSSDAFDI (SEQ ID ID NO: 13) (SEQ ID NO: 14) NO: 15)AB -4 NFGMS (SEQ GIYWNGVSTSYPDSVK GGSPYGDYGAFDI (SEQ ID ID NO: 16) G (SEQ ID NO: 17) NO: 18)AB-6 SGSYYWG SFYYSGTIYYNPSLKS QREYS S S SNYYYYYMD V (SEQ ID NO: 4) (SEQ ID NO: 5) (SEQ ID NO: 6)AB-7 SSSYYWG SIYFSGSTYYNPSLKS HREMATIRDYHYYYMDV (SEQ ID NO: 7) (SEQ ID NO: 8) (SEQ ID NO: 9)AB-10 DYAMH (SEQ GISWNQGNIDYADSVK DREYSGSYYDAFDI (SEQ ID ID NO: 10) G (SEQ ID NO: 19) NO: 12)AB-8 SNYMS (SEQ VIYSGGTKHYADSVKG SYYYGSGSSDAFDI (SEQ ID ID NO: 13) (SEQ ID NO: 14) NO: 15)AB-9 NFGMS (SEQ GIYWNAVSTSYPDSVK GGSPYGDYGAFDI (SEQ IDID NO: 16) G (SEQ ID NO: 20) NO: 18)TABLE 4: Light Chain CDR Sequences of Anti-pyroGlu3 Abeta (A0) Antibodies Antibody CDR-L1 CDR-L2 CDR-L3 AB-1 RASQSIRFYLN (SEQ GASSLQS (SEQ ID QQTYSTPIT ID NO: 21) NO: 22) (SEQ ID NO: 23) AB-2 RASQSISNYLN (SEQ AASSLQS (SEQ ID QQSYSTPLT ID NO: 24) NO: 25) (SEQ ID NO: 26) AB-5 RASQSISSWLA (SEQ KASSLEN (SEQ ID QEYNSYNT ID NO: 27) NO: 28) (SEQ ID NO: 29) AB-3 RASQSISNYLN (SEQ DAYSLQI (SEQ ID QQSYSIPPT ID NO: 24) NO: 30) (SEQ ID NO: 31) AB -4 RSSQSLVHSDGNTYLS KISNRFS (SEQ ID MQTTQFPYT (SEQ ID NO: 32) NO: 33) (SEQ ID NO: 34) AB-6 RASQSIRFYLN (SEQ GASSLQS (SEQ ID QQTYSTPIT ID NO: 21) NO: 22) (SEQ ID NO: 23) AB-7 RASQSISNYLN (SEQ AASSLQS (SEQ ID QQSYSTPLT ID NO: 24) NO: 25) (SEQ ID NO: 26) AB-10 RASQSISSWLA (SEQ KASSLES (SEQ ID QEYNSYNT ID NO: 27) NO: 35) (SEQ ID NO: 29) AB-8 RASQSISNYLN (SEQ DAYSLQI (SEQ ID QQSYSIPPTID NO: 24) NO: 30) (SEQ ID NO: 31)Antibody CDR-L1 CDR-L2 CDR-L3 AB-9 RSSQSLVHSEGNTYLS KISNRFS (SEQ ID MQTTQFPYT(SEQ ID NO: 36) NO: 33) (SEQ ID NO: 34)TABLE 5: VH and VL Sequences of Anti-pyroGlu3 Abeta (A0) Antibodies Antibody VH sequence VL sequenceAB-1 QLQLQESGPGLVKPSETLSLTCT DIQMTQSPSSLSASVGDRVTIT VSGGSISSGSYYWGWIRQPPGK CRASQSIRFYLNWYQQKPGKA GLEWIGSFYYSGTIYYNPSLKSR PKVLIYGASSLQSGVPSRFSGS VTISVDTSKNQFSLKLRSVTAAD GSGTDFTLTISSLQPEDFATYH T AVYYC ARQRE YS S S SNYYYYY CQQTYSTPITFGQGTRLEIK MDVWGKGTTVTVSS (SEQ ID (SEQ ID NO: 38)NO: 37)AB-2 QLHLQESGPGLVKPSETLSLTCT DIQMTQSPSSLSASVGDRVTIT VSGGSTS S S S YYWGWIRQPPGK CRASQSISNYLNWYQQKPGKA GLEWIGSIYFSGSTYYNPSLKSR PKLLIYAASSLQSGVPSRFSGS VTISVDTSKNQFSLKLKTVTAA GSGTDFTLTISSLQPEDFATYY DT AVYYC VRHREMATIRDYHY CQQSYSTPLTFGGGTRVEIK YYMDVWGKGTTVTVSS (SEQ (SEQ ID NO: 40)ID NO: 39)AB-5 EVQLVESGGGLVQPGRSLRLSC DIQMTQSPSTLSASVGDRVTIT AASGFTFDDYAMHWVRQAPGK CRASQSISSWLAWYQQKPGK GLEWVSGISWNNNNIDYADSVK APKVLIYKAS SLENGVPSRF SG GRFTISRDNAKNSLYLQMNSLR SGSGTEFTLTISSLQPDDFATY AEDTALYYCVKDREYSGSYYD YCQEYNSYNTFGQGTKLEIK AFDIWGQGTMVTVSS (SEQ ID (SEQ ID NO: 42)NO: 41)AB-3 EVQLVESGGGLIQPGGSLRLSCA DIQMTQSPSSLSASVGDRVTIT ASGFTVS SNYMSWVRQAPGKG CRASQSISNYLNWYQQKPGKA LEWVSVIYSGGTKHYADSVKGR PNLLIYDAYSLQIGVPSRFSGS FTISRDNSKNTLYLQMNSLRAE GSGTDFTLTISSLQPEDFATYS DT AVYYC ARSYYYGSGSSDAFD CQQSYSIPPTFGGGTKVEIK IWGQGTMVTVSS (SEQ ID NO: (SEQ ID NO: 44)43)AB-4 EVQLVESGGGVVRPGGSLRLSC DIVMTQTPLSSPVTLGQPASIS AASGFTFDNFGMSWVRQAPGK CRSSQSLVHSDGNTYLSWLQQ GLEWVSGIYWNGVSTSYPDSVK RPGQPPRLLIYKISNRFSGVPD GRFTISRDNAKNSLYLQMNSLR RF SGSGAGTDFTLKISRVEAED GEDTALYYCARGGSPYGDYGA VGVYYCMQTTQFPYTFGQGTKLEIK (SEQ ID NO: 46)Antibody VH sequence VL sequence FDIWGQGTVVTVSS (SEQ IDNO: 45)AB-6 QLQLQESGPGLVKPSETLSLTCT DIQMTQSPSSLSASVGDRVTIT VSGGSISSGSYYWGWIRQPPGK CRASQSIRFYLNWYQQKPGKA GLEWIGSFYYSGTIYYNPSLKSR PKVVIYGASSLQSGVPSRFSGS VTISVDTSKNQFSLKLSSVTAAD GSGTDFTLTISSLQPEDFATYY T AVYYC ARQRE YS S S SNYYYYY CQQTYSTPITFGQGTRLEIK MDVWGKGTTVTVSS (SEQ ID (SEQ ID NO: 48)NO: 47)AB-7 QLQLQESGPGLVKPSETLSLTCT DIQMTQSPSSLSASVGDRVTIT VSGGSTS S S S YYWGWIRQPPGK CRASQSISNYLNWYQQKPGKA GLEWIGSIYFSGSTYYNPSLKSR PKLLIYAASSLQSGVPSRFSGS VTISVDTSKNQFSLKLSSVTAAD GSGTDFTLTISSLQPEDFATYY T AVYYC VRHREMATIRDYHYY CQQSYSTPLTFGGGTKVEIK YMDVWGKGTTVTVSS (SEQ ID (SEQ ID NO: 50)NO: 49)AB-10 EVQLVESGGGLVQPGRSLRLSC DIQMTQSPSTLSASVGDRVTIT AASGFTFDDYAMHWVRQAPGK CRASQSISSWLAWYQQKPGK GLEWVSGISWNQGNIDYADSVK APKVVIYKASSLESGVPSRFSG GRFTISRDNAKNSLYLQMNSLR SGSGTEFTLTISSLQPDDFATY AEDTALYYCVKDREYSGSYYD YCQEYNSYNTFGQGTKLEIK AFDIWGQGTMVTVSS (SEQ ID (SEQ ID NO: 52)NO: 51)AB-8 EVQLVESGGGLIQPGGSLRLSCA DIQMTQSPSSLSASVGDRVTIT ASGFTVS SNYMSWVRQ APGKG CRASQSISNYLNWYQQKPGKA LEWVSVIYSGGTKHYADSVKGR PKLLIYDAYSLQIGVPSRFSGS FTISRDNSKNTLYLQMNSLRAE GSGTDFTLTISSLQPEDFATYY DT AVYYC ARSYYYGSGSSDAFD CQQSYSIPPTFGGGTKVEIK IWGQGTMVTVSS (SEQ ID NO: (SEQ ID NO: 53)43)AB-9 EVQLVESGGGVVRPGGSLRLSC DIVMTQTPLSSPVTLGQPASIS AASGFTFDNFGMSWVRQAPGK CRSSQSLVHSEGNTYLSWLQQ GLEWVSGIYWNAVSTSYPDSVK RPGQPPRLLIYKISNRFSGVPD GRFTISRDNAKNSLYLQMNSLR RF SGSGAGTDFTLKISRVEAED AEDTALYYCARGGSPYGDYGA VGVYYCMQTTQFPYTFGQGT FDIWGQGTMVTVSS (SEQ ID KLEIK (SEQ ID NO: 55)NO: 54)

[0245] In some aspects, provided herein are anti-pyroGlu3 Abeta (AP) antibodies comprising: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:4; (b) HVR-H2 comprising theamino acid sequence of SEQ ID NO: 5; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:6; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:21; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:22; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:23; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 8; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:9; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:24; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:25; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:26; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 10; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 11; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 12; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:27; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:28; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:29; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 13; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 14; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 15; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:24; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:30; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:31; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 16; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 17; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 18; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:32; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:33; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:34; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:4; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:5; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:6; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:21; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:22; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:23; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:8; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:9; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:24; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:25; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:26; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 10; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 19; (c) HVR-H3 comprising the aminoacid sequence of SEQ ID NO: 12; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:27; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:35; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:29; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 13; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 14; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 15; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:24; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:30; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:31; and (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 16; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:20; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 18; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:36; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:33; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:34.

[0246] In another aspect, an anti-pyroGlu3 Abeta (AP) antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:37, 39, 41, 43, 45, 47, 49, 51, and 54. In certain aspects, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 37, 39, 41, 43, 45, 47, 49, 51, and 54 contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-pyroGlu3 A Abeta (P) antibody comprising that sequence retains the ability to bind to pyroGlu3 Abeta (AP). In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted in SEQ ID NO: 37, 39, 41, 43, 45, 47, 49, 51, or 54. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 37, 39, 41, 43, 45, 47, 49, 51, or 54. In certain aspects, substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs).Optionally, the anti-pyroGlu3 Abeta (AP) antibody comprises the VH sequence of SEQ ID NO: 37, 39, 41, 43, 45, 47, 49, 51, and 54, including post- post-translational modifications of that sequence. In a particular aspect, the VH comprises one, two or three HVRs selected from: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:4, 7, 10, 13, and 16; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:5, 8, 11, 14, 17, 19, and 20; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:6, 9, 12, 15, and 18.

[0247] In another aspect, an anti-pyroGlu3 Abeta (AP) antibody is provided, wherein the antibody comprises a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:38, 40, 42, 44, 46, 48, 50, 52, 53, and 55. In certain aspects, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 38, 40, 42, 44, 46, 48, 50, 52, 53, and 55, and contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-pyroGlu3 Abeta (AP) antibody comprising that sequence retains the ability to bind to pyroGlu3 Abeta (AP). In some aspects, a total of 1 to 10 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 38, 40, 42, 44, 46, 48, 50, 52, 53, or 55. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 38, 40, 42, 44, 46, 48, 50, 52, 53, or 55. In certain aspects, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the anti-pyroGlu3 Abeta (AP) antibody comprises the VL sequence of SEQ ID NO: 38, 40, 42, 44, 46, 48, 50, 52, 53, and 55, including post-post-translational modifications of that sequence. In a particular aspect, the VL comprises one, two or three HVRs selected from (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:21, 24, 27, 32, and 36; (b) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:22, 25, 28, 30, 33, and 35; and (c) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:23, 26, 29, 31, and 34.

[0248] In some aspects, an anti-pyroGlu3 Abeta (AP) antibody is provided, wherein the antibody comprises a VH as in any of the aspects provided above, and a VL as in any of the aspects provided above. In some aspects, provided herein are anti-pyroGlu3 Abeta (AP) antibodies, wherein the antibody comprises a VH as in any of the aspects provided above, and a VL as in any of the aspects provided above. In one aspect, the antibody comprises the VH and VL sequences in SEQ ID NOs:37, 39, 41, 43, 45, 47, 49, 51, and 54, and SEQ ID NOs: 38, 40, 42, 44, 46, 48, 50, 52, 53, and 55, respectively, including post-post-translational modifications of those sequences.

[0249] In some aspects, provided herein are anti-pyroGlu3 Abeta (AP) antibodies comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), wherein the VH and VL are selected from the group consisting of: VH comprising the amino acid sequence of SEQ IDNO:37 and VL comprising the amino acid sequence of SEQ ID NO:38; VH comprising the amino acid sequence of SEQ ID NO:39 and VL comprising the amino acid sequence of SEQ ID NO:40; VH comprising the amino acid sequence of SEQ ID NO:41 and VL comprising the amino acid sequence of SEQ ID NO:42; VH comprising the amino acid sequence of SEQ ID NO:43 and VL comprising the amino acid sequence of SEQ ID NO:44; VH comprising the amino acid sequence of SEQ ID NO:45 and VL comprising the amino acid sequence of SEQ ID NO:46; VH comprising the amino acid sequence of SEQ ID NO:47 and VL comprising the amino acid sequence of SEQ ID NO:48; VH comprising the amino acid sequence of SEQ ID NO:49 and VL comprising the amino acid sequence of SEQ ID NO: 50; VH comprising the amino acid sequence of SEQ ID NO:51 and VL comprising the amino acid sequence of SEQ ID NO:52; VH comprising the amino acid sequence of SEQ ID NO:43 and VL comprising the amino acid sequence of SEQ ID NO:53; and VH comprising the amino acid sequence of SEQ ID NO:54 and VL comprising the amino acid sequence of SEQ ID NO: 55.

[0250] In some aspects, an anti-Abeta (pyroGlu3 AP) antibody of the present disclosure competitively inhibits binding of at least one reference antibody selected from anti-pyroGlu3 Abeta (AP) antibody AB-1, AB-2, AB-3, AB-4, AB-5, AB-6, AB-7, AB-8, AB-9, and AB-10, and any combination thereof, for binding to pyroGlu3 Abeta (AP).

[0251] In some aspects, an anti-pyroGlu3 Abeta (AP) antibody of the present disclosure binds to an epitope of human pyroGlu3 Abeta (AP) that is the same as or overlaps with the pyroGlu3 Abeta (AP) epitope bound by at least one reference antibody selected from anti-pyroGlu3 Abeta (AP) antibody AB-1, AB-2, AB-3, AB-4, AB-5, AB-6, AB-7, AB-8, AB-9, and AB-10. Detailed exemplary methods for mapping an epitope to which an antibody binds are provided in Morris (1996) “Epitope Mapping Protocols,” in Methods in Molecular Biology vol. 66 (Humana Press, Totowa, NJ).

[0252] In some aspects, an anti-pyroGlu3 Abeta (AP) antibody of the present disclosure competitively inhibits binding of at least one reference antibody, or binds to an epitope of human pyroGlu3 Abeta (AP) that is the same as or overlaps with the pyroGlu3 Abeta (AP) epitope bound by at least one reference antibody, wherein the reference antibody is an anti-pyroGlu3 Abeta (AP) antibody comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), wherein the VH and VL are selected from the group consisting of: VH comprising the amino acid sequence of SEQ ID NO:37 and VL comprising the amino acid sequence of SEQ ID NO:38; VH comprising the amino acid sequence of SEQ ID NO:39 and VL comprising theamino acid sequence of SEQ ID NO: 40; VH comprising the amino acid sequence of SEQ ID NO:41 and VL comprising the amino acid sequence of SEQ ID NO:42; VH comprising the amino acid sequence of SEQ ID NO:43 and VL comprising the amino acid sequence of SEQ ID NO:44; VH comprising the amino acid sequence of SEQ ID NO:45 and VL comprising the amino acid sequence of SEQ ID NO:46; VH comprising the amino acid sequence of SEQ ID NO:47 and VL comprising the amino acid sequence of SEQ ID NO:48; VH comprising the amino acid sequence of SEQ ID NO:49 and VL comprising the amino acid sequence of SEQ ID NO:50; VH comprising the amino acid sequence of SEQ ID NO:51 and VL comprising the amino acid sequence of SEQ ID NO:52; VH comprising the amino acid sequence of SEQ ID NO:43 and VL comprising the amino acid sequence of SEQ ID NO:53; and VH comprising the amino acid sequence of SEQ ID NO:54 and VL comprising the amino acid sequence of SEQ ID NO:55.

[0253] In some aspects, the anti-pyroGlu3 Abeta (AP) antibody according to any of the above aspects is a monoclonal antibody, including a humanized and / or human antibody. In some aspects, the anti-pyroGlu3 Abeta (AP) antibody is an antibody fragment, e.g., a Fv, Fab, Fab', scFv, diabody, or F(ab')2 fragment. In some aspects, the anti-pyroGlu3 Abeta (AP) antibody is a substantially full-length antibody, e.g., an IgGl antibody, IgG2a antibody, IgG4 antibody or other antibody class or isotype as defined herein.

[0254] In some aspects, an anti-pyroGlu3 Abeta (AP) antibody according to any of the above aspects may incorporate any of the features, singly or in combination, as described below.Multi-specific Binding Proteins that Bind to Blood-Brain Barrier Receptors or Proteins

[0255] Provided herein are multi-specific binding proteins that specifically bind to pyroGlu3 Abeta (AP) and specifically bind to human receptors or proteins of the blood-brain barrier. Such multi-specific binding proteins are capable of crossing the blood brain barrier (BBB) and capable of transporting other agents (e.g., therapeutically active agents, such as anti-pyroGlu3 Abeta (AP) antibodies, antigen-binding domains, or fragments thereof) with the composition across the blood brain barrier. Exemplary human receptors or proteins of the blood-brain barrier are transferrin receptor (TfR) and CD98 heavy chain (CD98hc).

[0256] In some aspects, the compositions provided herein comprise a pyroGlu3 Abeta (AP) antigen-binding domain and a TfR antigen-binding domain. In some aspects, the compositions provided herein comprise a pyroGlu3 Abeta (AP) antigen-binding domain and a CD98hc antigenbinding domain.Antigen-Binding Domains That Bind to TfR

[0257] Provided herein are antigen-binding domains that specifically bind to human TfR. Such anti-TfR antigen-binding domains are capable of crossing the blood brain barrier (BBB) and capable of transporting other agents (e.g., therapeutically active agents) associated with the antigen-binding domain across the blood brain barrier. Accordingly, in some aspects, provided herein are antigen-binding domains that specifically bind to human TfR and are capable of being internalized in blood brain barrier epithelial cells.

[0258] Antibodies (e.g., antigen-binding domains) to human transferrin receptor were obtained, characterized, and recombinantly expressed as described previously in WO2024 / 026472 (filed July 28, 2023) and WO2025 / 166045 (International Application No. PCT / US2025 / 013846; filed January 30, 2025), each of which is incorporated herein by reference in its entirety.

[0259] Table 6 below lists the heavy chain CDR amino acid sequences (according to Kabat numbering) of anti-TfR antigen-binding domains of the present disclosure. Table 7 below lists the light chain CDR amino acid sequences (according to Kabat numbering) of anti-TfR antigenbinding domains of the present disclosure. Table 8 below lists the heavy chain variable region and light chain variable region amino acid sequences of anti-TfR antigen-binding domains of the present disclosure.TABLE 6: Heavy Chain CDR Sequences of Anti-TfR Antigen Binding DomainsAntibody CDR-H1 CDR-H2 CDR-H3 TfR.15. WH8.1.24A SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SAYYGNFDTMDN (SEQ ID NO: 56) ID NO: 57) ID NO: 58) TfR.15. WH8.1.24A.42Q SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SAYYGNFDTMDN (SEQ ID NO: 56) ID NO: 57) ID NO: 58) TfR.15. WH8.1.42Q SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGNFDTMDN (SEQ ID NO: 56) ID NO: 57) ID NO: 59) TfR.15. WH8.1.42Q. H6- SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGRFDTMDN (SEQ 4 ID NO: 56) ID NO: 57) ID NO: 60) TfR.15. WH8.1.42Q. L7- SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGNFDTMDN (SEQ 2 ID NO: 56) ID NO: 57) ID NO: 59) TfR.15. WH8.1.42Q. L10- SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGNFDTMDN (SEQ 1 ID NO: 56) ID NO: 57) ID NO: 59) TfR.15. WH8.1.42Q. L10- SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGNFDTMDN (SEQ 8 ID NO: 56) ID NO: 57) ID NO: 59) TfR.15. WH8.1.42Q. L10- SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGNFDTMDN (SEQ16 ID NO: 56) ID NO: 57) ID NO: 59)Antibody CDR-H1 CDR-H2 CDR-H3 TfR.15. WH8.1.42Q. H3- SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSNYGNFDTMDN (SEQ 7 ID NO: 56) ID NO: 57) ID NO: 61) TfR.9.1B.39.38 EYAMH (SEQ GIAPNTGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ ID ID NO: 62) ID NO: 63) NO: 64) TfR.9.1B.39.27. L-35 EYAMH (SEQ GIAPGTGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ ID ID NO: 62) ID NO: 65) NO: 64) TfR.9.1B.39.38. L-21 EYAMH (SEQ GIAPNTGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ ID ID NO: 62) ID NO: 63) NO: 64) TfR.9.1B.39.38. L-6 EYAMH (SEQ GIAPNTGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ ID ID NO: 62) ID NO: 63) NO: 64) TfR.9.1B.39.27. L-19 EYAMH (SEQ GIAPGTGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ ID ID NO: 62) ID NO: 65) NO: 64) TfR.15. WH8.1 SYYMH (SEQ NIYPGSGSSNYAQKFQG (SEQ SSYYGNFDTMDN (SEQ ID NO:56) ID NO: 57) ID NO: 59) TfR.9.1B.35 EYAMH (SEQ GINPNDGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ ID ID NO: 62) ID NO: 275) NO: 64) TfR.9.1B.39 EYAMH (SEQ GINPNDGGTSYNQKFKG (SEQ QGWLLRGGDY (SEQ IDID NO: 62) ID NO: 275) NO: 64)TABLE 7: Light Chain CDR Sequences of Anti-TfR Antigen Binding Domains Antibody CDR-L1 CDR-L2 CDR-L3 TfR.15. WH8.1.24A RASSSIRYMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 66) NO: 67) ID NO: 68) TfR.15. WH8.1.24A.42Q RASSSIRQMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 69) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q RASSSIRQMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 69) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q. H6-4 RASSSIRQMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 69) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q. L7-2 RASSSIDQMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 70) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q. L10-l RASSSIRQMA (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 71) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q. L10-8 RASSSIRQMS (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 72) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q. L10-16 RASSSIRQMH (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 73) NO: 67) ID NO: 68) TfR.15. WH8.1.42Q. H3-7 RASSSIRQMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 69) NO: 67) ID NO: 68) TfR.9.1B.39.38 KASQSLLDEGGKTYLN LVSKLDS (SEQ ID WQGTHFPQT (SEQ (SEQ ID NO: 74) ID NO: 76)NO: 75)Antibody CDR-L1 CDR-L2 CDR-L3 TfR.9.1B.39.27. L-35 KASQSLLDEDGKTYLN LVSKLDS (SEQ ID WQGTHFPQT (SEQ (SEQ ID NO: 77) NO: 75) ID NO: 76) TfR.9.1B.39.38. L-21 KASQSLLDEDNKTYLN LVSKLDS (SEQ ID WQGTHFPQT (SEQ (SEQ ID NO: 78) NO: 75) ID NO: 76) TfR.9.1B.39.38. L-6 KASQSLLDEDKKTYLN LVSKLDS (SEQ ID WQGTHFPQT (SEQ (SEQ ID NO: 79) NO: 75) ID NO: 76) TfR.9.1B.39.27. L-19 KASQSLLDEDAKTYLN LVSKLDS (SEQ ID WQGTHFPQT (SEQ (SEQ ID NO: 80) NO: 75) ID NO: 76) TfR.15. WH8.1 RASSSIRYMN (SEQ ID DTSNRAT (SEQ ID HQRSSYPWT (SEQ NO: 66) NO: 67) ID NO: 68) TfR.9.1B.35 KSSQSLLDSDGKTYLN LVSKLDS (SEQ ID WQGTHFPQS (SEQ (SEQ ID NO: 276) NO: 75) ID NO: 277) TfR.9.1B.39 KASQSLLDEDGKTYLN LVSKLDS (SEQ ID WQGTHFPQT (SEQ (SEQ ID NO: 77) ID NO: 76)NO: 75)TABLE 8: VH and VL Sequences of Anti-TfR Antigen Binding Domains Antibody VH sequence VL sequence TfR.15. WH8.1.24A QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIRYMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSAYYGNFDTMDN TFGGGTKVEIK (SEQ ID NO: 82) WGQGTTVTVSS (SEQ ID NO: 81)TfR.15. WH8.1.24A.42Q QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIRQMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSAYYGNFDTMDN TFGGGTKVEIK (SEQ ID NO: 83) WGQGTTVTVSS (SEQ ID NO: 81)TfR.15. WH8.1.42Q QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIRQMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 83) GQGTTVTVSS (SEQ ID NO: 84)TfR.15. WH8.1.42Q. H6-4 QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIRQMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGRFDTMDNW TFGGGTKVEIK (SEQ ID NO: 83)GQGTTVTVSS (SEQ ID NO: 85)Antibody VH sequence VL sequenceTfR.15. WH8.1.42Q. L7-2 QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIDQMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 86) GQGTTVTVSS (SEQ ID NO: 84)TfR.15. WH8.1.42Q. L10- QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR 1 KASGYTFTSYYMHWVRQAPGQG ASSSIRQMAWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 87) GQGTTVTVSS (SEQ ID NO: 84)TfR.15. WH8.1.42Q. L10- QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR 8 KASGYTFTSYYMHWVRQAPGQG ASSSIRQMSWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 88) GQGTTVTVSS (SEQ ID NO: 84)TfR.15. WH8.1.42Q. L10- QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR 16 KASGYTFTSYYMHWVRQAPGQG ASSSIRQMHWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 89) GQGTTVTVSS (SEQ ID NO: 84)TfR.15. WH8.1.42Q. H3-7 QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIRQMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSNYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 83) GQGTTVTVSS (SEQ ID NO: 90)TfR.9.1B.39.38 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCK KASGYTFTEYAMHWVRQAPGQG ASQSLLDEGGKTYLNWLQQRPG LEWMGGIAPNTGGTSYNQKFKG QSPRRLIYLVSKLDSGVPDRFSGS RVTMTVDTSTSTAYMELSSLRSE GSGTDFTLKISRVEAEDVGVYYC DTAVYYCASQGWLLRGGDYWG WQGTHFPQTFGGGTKVEIK (SEQ QGTLVTVSS (SEQ ID NO: 91) ID NO: 92)TfR.9. IB.39.27. L-35 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCK KASGYTFTEYAMHWVRQAPGQG ASQSLLDEDGKTYLNWLQQRPG LEWMGGIAPGTGGTSYNQKFKG QSPRRLIYLVSKLDSGVPDRFSGS RVTMTVDTSTSTAYMELSSLRSE GSGTDFTLKISRVEAEDVGVYYC DTAVYYCASQGWLLRGGDYWG WQGTHFPQTFGGGTKVEIK (SEQ QGTLVTVSS (SEQ ID NO: 93) ID NO: 94)TfR.9.1B.39.38. L-21 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCKKASGYTFTEYAMHWVRQAPGQG ASQSLLDEDNKTYLNWLQQRPGAntibody VH sequence VL sequence LEWMGGIAPNTGGTSYNQKFKG QSPRRLIYLVSKLDSGVPDRFSGS RVTMTVDTSTSTAYMELSSLRSE GSGTDFTLKISRVEAEDVGVYYC DTAVYYCASQGWLLRGGDYWG WQGTHFPQTFGGGTKVEIK (SEQ QGTLVTVSS (SEQ ID NO: 91) ID NO: 95)TfR.9.1B.39.38. L-6 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCK KASGYTFTEYAMHWVRQAPGQG ASQSLLDEDKKTYLNWLQQRPG LEWMGGIAPNTGGTSYNQKFKG QSPRRLIYLVSKLDSGVPDRFSGS RVTMTVDTSTSTAYMELSSLRSE GSGTDFTLKISRVEAEDVGVYYC DTAVYYCASQGWLLRGGDYWG WQGTHFPQTFGGGTKVEIK (SEQ QGTLVTVSS (SEQ ID NO: 91) ID NO: 96)TfR.9. IB.39.27. L-19 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCK KASGYTFTEYAMHWVRQAPGQG ASQSLLDEDAKTYLNWLQQRPG LEWMGGIAPGTGGTSYNQKFKG QSPRRLIYLVSKLDSGVPDRFSGS RVTMTVDTSTSTAYMELSSLRSE GSGTDFTLKISRVEAEDVGVYYC DTAVYYCASQGWLLRGGDYWG WQGTHFPQTFGGGTKVEIK (SEQ QGTLVTVSS (SEQ ID NO: 93) ID NO: 97)TfR.15. WH8.1 QVQLVQSGAEVKKPGASVKVSC EIVLTQSPATLSLSPGERATLSCR KASGYTFTSYYMHWVRQAPGQG ASSSIRYMNWYQQKPGQAPRRLI LEWMGNIYPGSGSSNYAQKFQG YDTSNRATGIPARFSGSGSGTDFT RVTMTRDTSTSTVYMELS SLRSE LTIS SLEPEDFA VYYCHQRS SYPW DTAVYYCTRSSYYGNFDTMDNW TFGGGTKVEIK (SEQ ID NO: 270) GQGTTVTVSS (SEQ ID NO: 269)TfR.9. IB.35 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCK KASGYTFTEYAMHWVRQAPGQG SSQSLLDSDGKTYLNWLQQRPGQ LEWMGGINPNDGGTSYNQKFKG SPRRLIYLVSKLDSGVPDRFSGSG RVTMTVDTSTSTAYMELSSLRSE SGTDFTLKISRVEAEDVGVYHCW DTAVYYCASQGWLLRGGDYWG QGTHFPQSFGGGTKVEIK (SEQ ID QGTLVTVSS (SEQ ID NO:271) NO: 272)TfR.9. IB.39 QVQLVQSGAEVKKPGASVKVSC DVVMTQSPLSLPVTLGQPASISCK KASGYTFTEYAMHWVRQAPGQG ASQSLLDEDGKTYLNWLQQRPG LEWMGGINPNDGGTSYNQKFKG QSPRRLIYLVSKLDSGVPDRFSGS RVTMTVDTSTSTAYMELSSLRSE GSGTDFTLKISRVEAEDVGVYYC DTAVYYCASQGWLLRGGDYWG WQGTHFPQTFGGGTKVEIK (SEQ QGTLVTVSS (SEQ ID NO:273) ID NO: 274)

[0260] In the present disclosure, “TfRL10-16” or “TfR10-16” or “10-16” refers to TfR.15. WH8.1.42Q. L10-16;

[0261] “TfRL10-8” or“TfR10-8” or “10-8” refer to TfR.15. WH8.1.42Q. L10-8;

[0262] “TfR7-2” or “TfRL7-2” or “L7-2” refers to TfR.15. WH8.1.42Q. L7-2;

[0263] “TfR42Q” or “42Q” refers to TfR.15. WH8.1.42Q;

[0264] “TfR39.38” or “39.38” refers to TfR.9.1B.39.38; and

[0265] “TfR24A” or “24A” refers to TfR.15. WH8.1.24A.

[0266] In some aspects, the CDRs of an antigen-binding domain that specifically binds to human TfR can be determined according to MacCallum RM et al., (1996) J Mol Biol 262: 732-745. See also, e.g., Martin A. “Protein Sequence and Structure Analysis of Antibody Variable Domains,” in Antibody Engineering, Kontermann and Diibel, eds., Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001). In some aspects, provided herein are antigen-binding domains that specifically bind to human TfR and comprise VH and VL CDRs of an antibody listed in Tables 6, 7, and 8 as determined by the method in MacCallum RM et al.

[0267] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the six CDRs of an antibody listed in Tables 6, 7, and 8 above (i.e. the three VH CDRs of the antibody listed in Table 6 and the three VL CDRs of the same antibody listed in Table 7) as determined by Kabat numbering. In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the six Chothia CDRs of an antibody listed in Table 8.

[0268] In some aspects, the CDRs of an antigen-binding domain that specifically binds to human TfR can be determined according to the AbM numbering scheme, which refers to AbM hypervariable regions, which represent a compromise between the Kabat CDRs and Chothia structural loops, and are used by Oxford Molecular’s AbM antibody modeling software (Oxford Molecular Group, Inc.). In some aspects, provided herein are antigen-binding domains that specifically bind to human TfR and comprise VH and VL CDRs of an antibody listed in Table 8 as determined by the AbM numbering scheme.

[0269] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the six IMGT CDRs of an antibody listed in listed in Table 8 according to the IMGT numbering system as described in Lefranc M-P, (1999) The Immunologist 7: 132-136 and Lefranc M-P et al., (1999) Nucleic Acids Res 27: 209-212. According to the IMGT numbering scheme, VH-CDR1 is at positions 26 to 35, VH-CDR2 is at positions 51 to 57, VH-CDR3 is at positions 93 to 102, VL-CDR1 is at positions 27 to 32, VL-CDR2 is at positions 50 to 52, and VL-CDR3 is at positions 89 to 97.

[0270] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the VH of an antibody listed in Table 8.

[0271] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the VL of antibody listed in Table 8.

[0272] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the VH and the VL of an antibody listed in Table 8.

[0273] In some aspects, an antigen-binding domain of the present disclosure that specifically binds to human TfR comprises: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:58; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:66; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:58; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:69; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:59; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:69; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:60; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:69; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:59; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:70; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:59; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:71; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3comprising the amino acid sequence of SEQ ID NO:59; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:72; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:59; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:73; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:61; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:69; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:63; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:74; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:76; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:65; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:77; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:76; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:63; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:78; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:76; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:63; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:79; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:76; and (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:65; (c) HVR-H3 comprising the amino acid sequence of SEQ IDNO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:80; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:76.

[0274] In some aspects, an antigen-binding domain of the present disclosure that specifically binds to human TfR comprises: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:56; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:57; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:59; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:66; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:67; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:68; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:275; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:276; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:277; or (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:62; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:275; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:64; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:77; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:76.

[0275] In another aspect, an antigen-binding domain of the present disclosure that specifically binds to human TfR comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:81, 84, 85, 90, 91, and 93. In certain aspects, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 81, 84, 85, 90, 91, and 93 contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an antigen-binding domain that specifically binds to human TfR comprising that sequence retains the ability to bind to human TfR. In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted in SEQ ID NO: 81, 84, 85, 90, 91, or 93. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 81, 84, 85, 90, 91, or 93. In certain aspects, substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the antigen-binding domain that specifically binds to human TfR comprises the VHsequence of SEQ ID NO: 81, 84, 85, 90, 91, and 93, including post- post-translational modifications of that sequence. In a particular aspect, the VH comprises one, two or three HVRs selected from: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:56 and 62; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:57, 63, and 65; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:58, 59, 60, 61, and 64.

[0276] In another aspect, an antigen-binding domain of the present disclosure that specifically binds to human TfR comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 269, 271, and 273. In certain aspects, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 269, 271, and 273 contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an antigen-binding domain that specifically binds to human TfR comprising that sequence retains the ability to bind to human TfR. In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted in SEQ ID NO: 269, 271, or 273. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 269, 271, or 273. In certain aspects, substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the antigen-binding domain that specifically binds to human TfR comprises the VH sequence of SEQ ID NO: 269, 271, and 273 including post- post-translational modifications of that sequence. In a particular aspect, the VH comprises one, two or three HVRs selected from: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 56 and 62; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:57, 63, 65, and 275; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ IDNOs:58, 59, 60, 61, and 64.

[0277] In another aspect, an antigen-binding domain of the present disclosure that specifically binds to human TfR comprises comprises a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, and 97. In certain aspects, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to an amino acid sequence selected from the groupconsisting of SEQ ID NOs: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, and 97, and contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an antigen-binding domain of the present disclosure that specifically binds to human TfR comprising that sequence retains the ability to bind to human TfR. In some aspects, a total of 1 to 10 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, or 97. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, or 97. In certain aspects, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the antigen-binding domain of the present disclosure that specifically binds to human TfR comprises the VL sequence of SEQ ID NO: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, and 97, including post-post-translational modifications of that sequence. In a particular aspect, the VL comprises one, two or three HVRs selected from (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:66, 69, 70, 71, 72, 73, 74, 77, 78, 79, and 80; (b) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:67 and 75; and (c) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:68 and 76.

[0278] In another aspect, an antigen-binding domain of the present disclosure that specifically binds to human TfR comprises comprises a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:270, 272, and 274. In certain aspects, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 270, 272, and 274, and contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an antigen-binding domain of the present disclosure that specifically binds to human TfR comprising that sequence retains the ability to bind to human TfR. In some aspects, a total of 1 to 10 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 270, 272, or 274. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 270, 272, or 274. In certain aspects, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the antigen-binding domain of the present disclosure that specifically binds to human TfR comprises the VL sequence of SEQ ID NO: 270, 272 and 274, including post-post-translational modifications of that sequence. In a particular aspect, the VL comprises one,two or three HVRs selected from (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:66, 69, 70, 71, 72, 73, 74, 77, 78, 79, 80, and 276; (b) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:67 and 75; and (c) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs:68, 76 and 277.

[0279] In some aspects, an antigen-binding domain of the present disclosure that specifically binds to human TfR is provided, wherein the antibody comprises a VH as in any of the aspects provided above, and a VL as in any of the aspects provided above. In some aspects, the antigenbinding domain comprises the VH and VL sequences in SEQ ID NOs: 81, 84, 85, 90, 91, and 93, and SEQ ID NOs: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, and 97, respectively, including post-post-translational modifications of those sequences.

[0280] In some aspects, an antigen-binding domain of the present disclosure that specifically binds to human TfR is provided, wherein the antibody comprises a VH as in any of the aspects provided above, and a VL as in any of the aspects provided above. In some aspects, the antigenbinding domain comprises the VH and VL sequences in SEQ ID NOs: 81, 84, 85, 90, 91, 93, 269, 271, and 273 and SEQ ID NOs: 82, 83, 86, 87, 88, 89, 92, 94, 95, 96, 97, 270, 272, and 274, respectively, including post-post-translational modifications of those sequences.

[0281] In some aspects, provided herein are antigen-binding domains that specifically bind to human TfR comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), wherein the VH and VL are selected from the group consisting of: VH comprising the amino acid sequence of SEQ ID NO:81 and VL comprising the amino acid sequence of SEQ ID NO:82; VH comprising the amino acid sequence of SEQ ID NO:81 and VL comprising the amino acid sequence of SEQ ID NO:83; VH comprising the amino acid sequence of SEQ ID NO:84 and VL comprising the amino acid sequence of SEQ ID NO:83; VH comprising the amino acid sequence of SEQ ID NO:85 and VL comprising the amino acid sequence of SEQ ID NO:83; VH comprising the amino acid sequence of SEQ ID NO:84 and VL comprising the amino acid sequence of SEQ ID NO:86; VH comprising the amino acid sequence of SEQ ID NO:84 and VL comprising the amino acid sequence of SEQ ID NO: 87; VH comprising the amino acid sequence of SEQ ID NO:84 and VL comprising the amino acid sequence of SEQ ID NO:88; VH comprising the amino acid sequence of SEQ ID NO:84 and VL comprising the amino acid sequence of SEQ ID NO:89; VH comprising the amino acid sequence of SEQ ID NO:90 and VL comprising the amino acid sequence of SEQ ID NO:83; VH comprising the amino acid sequence of SEQ ID NO:91 and VLcomprising the amino acid sequence of SEQ ID NO:92; VH comprising the amino acid sequence of SEQ ID NO:93 and VL comprising the amino acid sequence of SEQ ID NO:94; VH comprising the amino acid sequence of SEQ ID NO:91 and VL comprising the amino acid sequence of SEQ ID NO:95; VH comprising the amino acid sequence of SEQ ID NO:91 and VL comprising the amino acid sequence of SEQ ID NO:96; and VH comprising the amino acid sequence of SEQ ID NO:93 and VL comprising the amino acid sequence of SEQ ID NO:97.

[0282] In some aspects, provided herein are antigen-binding domains that specifically bind to human TfR comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), wherein the VH and VL are selected from the group consisting of: VH comprising the amino acid sequence of SEQ ID NO:269 and VL comprising the amino acid sequence of SEQ ID NO:270; VH comprising the amino acid sequence of SEQ ID NO:271 and VL comprising the amino acid sequence of SEQ ID NO:272; and VH comprising the amino acid sequence of SEQ ID NO:273 and VL comprising the amino acid sequence of SEQ ID NO:274. In some aspects, provided herein is an antigen-binding domain that competitively inhibits binding to human TfR of an antibody comprising a VH amino acid sequence of an antibody in Table 8 and a VL amino acid sequence of the same antibody in Table 8.

[0283] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises a VH and a VL on a single polypeptide chain (e.g., a VH and VL in Table 8).

[0284] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises an scFv. In some aspects, an scFv comprises a VH and a VL of an antibody listed in Table 8. The scFv can comprise a VH that is N-terminal to a VL or a VL that is N-terminal to a VH. The scFv can comprise a linker, e.g., between a VH and a VL. Accordingly, the scFv can be in the orientation VH-linker-VL or VL-linker-VH. Such a linker can be about 5 to about 25 amino acids in length. Such a linker can be about 5 to about 20 amino acids in length. Such a linker can be about 10 to about 25 amino acids in length. Such a linker can be about 10 to about 20 amino acids in length. Such a linker can be, e.g., a glycine linker, a glycine-rich linker, or a glycine-serine linker. Such a linker can comprise the amino acid sequence of GGSEGKSSGSGSESKSTGGS (SEQ ID NO: 209). Such a linker can comprise the amino acid sequence of GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 210).

[0285] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises a VH on a first polypeptide and a VL on a second polypeptide e.g., a Fab). The Fab comprises one constant domain, one VH, and one VL. In some aspects, the antigen-bindingdomain comprises a Fab comprising a constant domain, a VH of an antibody listed in Table 8 and a VL of the same antibody listed in Table 8. In some aspects, the antigen-binding domain that comprises a Fab comprises the CDRs of the antibody in Tables 6, 7, and 8 (e.g., the nonidentical amino acids in the VH and / or VL are outside of the CDRs).

[0286] In some aspects, an antigen-binding domain that specifically binds to human TfR comprises the antigen-binding fragment of a heavy chain only antibody (e.g., a VHH or nanobody). In some aspects, an antigen-binding domain comprises a VHH comprising the VH of an antibody listed in Table 8. In other aspects, the antigen-binding domain comprises a VHH comprises the heavy chain CDRs of an antibody listed in Tables 6 and 8.

[0287] In some aspects, an antigen-binding domain that specifically binds to human TfR is a murine antigen-binding domain. In some aspects, an antigen-binding domain that specifically binds to human TfR is a chimeric antigen-binding domain. In some aspects, an antigen-binding domain that specifically binds to human TfR is a humanized antigen-binding domain. In some aspects, an antigen-binding domain that specifically binds to human TfR is a human antigenbinding domain.

[0288] In some aspects, an antigen-binding domain provided herein that specifically binds to human TfR also binds to cynomolgus monkey TfR.

[0289] In some aspects, an anti-TfR antigen-binding domain of the present disclosure comprises a heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3, and a light chain variable region (VL) CDR1, VL CDR2, and VL CDR3 sequences comprising the amino acid sequences of SEQ ID NOs:56, 57, 59, 70, 67, and 68, respectively.

[0290] In some aspects, the anti-TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs:84 and 86, respectively.

[0291] In some aspects, the anti-TfR antigen-binding domain comprises a heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and a light chain variable region (VL) CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs:56, 57, 59, 73, 67, and 68, respectively.

[0292] In some aspects, the anti-TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs:84 and 89, respectively.

[0293] In some aspects, the anti-TfR antigen-binding domain comprises a heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and a lightchain variable region (VL) CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs:62, 63, 64, 74, 75, and 76, respectively.

[0294] In some aspects, the anti-TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs:91 and 92, respectively.

[0295] In some aspects, the antigen-binding domain provided herein binds human TfR with an affinity between about 500 nM and about 10,000 nM. In some aspects, the antigen-binding domain binds human TfR with an affinity between about 500 nM and about 5,000 nM, between about 500 nM and about 2,500 nM, between about 500 nM and about 1,000 nM, between about 500 nM and about 750 nM, between about 1,000 nM and about 2,5000 nM, between about 2,500 nM and about 5,000 nM, between about 2,500 nM and about 10,000 nM, between about 5,000 nM and about 10,000 nM, and values and ranges there between.

[0296] In some aspects, an antigen-binding domain provided herein binds human TfR with an affinity between about 10 nM and about 500 nM. In some aspects, the antigen-binding domain binds human TfR with an affinity between about 10 nM and about 250 nM, between about 10 nM and about 100 nM, between about 10 nM and about 50 nM, between about 50 nM and about 500 nM, between about 50 nM and about 250 nM, between about 100 nM and about 250 nM, between about 250 nM and about 500 nM, and values and ranges there between.

[0297] In some aspects, the antigen-binding domain binds human TfR with an affinity of less than about 10 nM. In some aspects, the antigen -binding domain binds human TfR with an affinity between about 1 nM and about 10 nM, between about 1 nM and about 5 nM, between about 0.1 nM and about 1 nM, between about 0.01 nM and about 0.1 nM, and values and ranges there between.

[0298] In some aspects, the affinity of anti-TfR antibody TfR.15. WH8.1.42Q. L10-16 (i.e., anti-TfRL10-16) to human TfR is approximately 2010 nM. In some aspects, the affinity of anti-TfR antibody TfR.15. WH8.1.42Q. L7-2 (i.e., anti-TfRL7-2) is approximately 309 nM. In some aspects, the affinity of anti-TfR antibody TfR.9.1B.39.38 (i.e., anti-TfR39.38) is approximately 3810 nM. Without being bound by this theory, use of an anti-TfR antibody in the multispecific binding proteins described herein, wherein the anti-TfR antibody displays lower affinity binding to human TfR (e.g., -2000 nM), minimizes interference with iron uptake by reticulocytes, thereby reducing the likelihood of anemia. In some aspects, administration of a composition comprising an anti-pyroGlu3 A0 antigen-binding domain and an anti-TfR antigen binding domain of the present disclosure does not result in anemia.

[0299] In some aspects, the binding of an antigen-binding domain provided herein that specifically binds to human TfR is measured using surface plasmon resonance. Surface plasmon resonance can be measured, e.g., using the Carterra LSA platform or BIACORE®. In some aspects, an antigen-binding domain provided herein that specifically binds to human TfR binds to human TfR as measured by, for example, radioimmunoassay (RIA), Western blot, or ELISA OD450.

[0300] Also provided herein are antigen-binding domains that bind to the same epitope of TfR as a TfR antigen-binding domain provided herein. Also provided herein are antigen-binding domains that competitively inhibit binding to TfR of TfR antigen-binding domain provided herein.Antigen-Binding Domains That Bind to CD98hc

[0301] Provided herein are antigen-binding domains that specifically bind to human CD98hc.

[0302] Such antigen-binding domains can be capable of crossing the blood brain barrier (BBB) and capable of transporting other agents (e.g., therapeutically active agents) associated with the antigen-binding domain across the blood brain barrier. Accordingly, in some aspects, provided herein are antigen-binding domains that specifically bind to human CD98hc that are capable of being internalized in blood brain barrier epithelial cells.

[0303] Antibodies (e.g., antigen-binding domains) to human CD98hc were obtained, characterized, and recombinantly expressed as described previously in WO2024 / 026471 (filed July 28, 2023) and in WO2025 / 166042 (International Application Serial No.PCT / US2025 / 013843; filed January 30, 2025), both of which are incorporated herein by reference in their entireties.

[0304] Table 9 below lists the heavy chain CDR amino acid sequences (according to Kabat numbering) of anti-CD98hc antigen-binding domains of the present disclosure. Table 10 below lists the light chain CDR amino acid sequences (according to Kabat numbering) of anti-CD98hc antigen-binding domains of the present disclosure. Table 11 below lists the heavy chain variable region and light chain variable region amino acid sequences of anti-CD98hc antigen-binding domains of the present disclosure.TABLE 9: Heavy Chain CDR Sequences of Anti-CD98hc AntibodiesAntibody CDR-H1 CDR-H2 CDR-H3 CD98hc.04.048. WHl AYDMS (SEQ ID YISSGGGSTYYADSVKG QGGLPAWFVY NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.15 AYDMS YIS SGGGSTYYADEVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 101) (SEQ ID NO: 100) CD98hc.04.048. WH1.20 AYDMS YIS SGGGSTYYADEVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 102) (SEQ ID NO: 100) CD98hc.04.048. WH1.23 AYDMS YIS SGGGSTYYADYVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 103) (SEQ ID NO: 100) CD98hc.04.048. WH1.078 AYDMS YKSSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 104) (SEQ ID NO: 100) CD98hc.04.048. WH1.083 AYDMS YRSSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 105) (SEQ ID NO: 100) CD98hc.04.048. WH1.105 AYDMS YISSQGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 106) (SEQ ID NO: 100) CD98hc.04.048. WH1.118 AYDMS YISSGKGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 107) (SEQ ID NO: 100) CD98hc.04.048. WH1.119 AYDMS YISSGHGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 108) (SEQ ID NO: 100) CD98hc.04.048. WH1.121 AYDMS YIS SGRGSTYYAD SVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 109) (SEQ ID NO: 100) CD98hc.04.048. WH1.133 AYDMS YISSGGGHTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 110) (SEQ ID NO: 100) CD98hc.04.048. WH1.193 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.217 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.218 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.222 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.224 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.232 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.236 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.296 AYDMS YISSQGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 106) (SEQ ID NO: 100) CD98hc.04.048. WH1.151 AYDMS YISSGGGSDYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 111) (SEQ ID NO: 100) CD98hc.04.048. WH1.158 AYDMS YISSGGGSTYYADSVKG QGSLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 112) CD98hc.04.048. WH1.162 AYDMS YISSGGGSTYYADSVKG QGGHPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 113) CD98hc.04.048. WH1.165 AYDMS YISSGGGSTYYADSVKG QGGAPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 114) CD98hc.04.048. WH1.166 AYDMS YISSGGGSTYYADSVKG QGGTPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 115) CD98hc.04.048. WH1.167 AYDMS YISSGGGSTYYADSVKG QGGQPAWFVY(SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 116)Antibody CDR-H1 CDR-H2 CDR-H3 CD98hc.04.048. WH1.168 AYDMS YISSGGGSTYYADSVKG QGGLPALFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 117) CD98hc.04.048. WH1.170 AYDMS YISSGGGSTYYADSVKG QGGLPAKFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 118) CD98hc.04.048. WH1.171 AYDMS YISSGGGSTYYADSVKG QGGLPATFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 119) CD98hc.04.048. WH1.174 AYDMS YISSGGGSTYYADSVKG QGGLPARFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 120) CD98hc.04.048. WH1.185 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.208 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.231 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.235 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.250 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.253 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.262 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.269 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.271 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.274 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.276 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.279 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.281 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.282 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.283 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.39 AYDMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 98) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.55 AYAMS YISSGGGSTYYADSVKG QGGLPAWFVY (SEQ ID NO: 121) (SEQ ID NO: 99) (SEQ ID NO: 100) CD98hc.04.048. WH1.76 AYAMS YISSGGGSTYYADSVKG QGGLPAWFVY(SEQ ID NO: 121) (SEQ ID NO: 99) (SEQ ID NO: 100)TABLE 10: Heavy Chain CDR Sequences of Anti-CD98hc Antibodies Antibody CDR-L1 CDR-L2 CDR-L3 CD98hc.04.048. WHl KSSQSLLYSNTQKNYLA WASTRES (SEQ QQYSYYPPT(SEQ ID NO: 122) ID NO: 123) (SEQ ID NO: 124)Antibody CDR-L1 CDR-L2 CDR-L3 CD98hc.04.048. WH1.15 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.20 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.23 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.078 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.083 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.105 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.118 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.119 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.12I KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.133 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.193 KSSQSLKYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 125) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.2I7 KSSQSLLYQNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 126) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.2I8 KSSQSLLYENTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 127) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.222 KSSQSLLYSHTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 128) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.224 KSSQSLLYSTTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 129) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.232 KSSQSLLYSNSQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 130) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.236 KSSQSLLYSNKQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 131) (SEQ ID NO: (SEQ ID NO: 124)123)Antibody CDR-L1 CDR-L2 CDR-L3 CD98hc.04.048. WH1.296 KSSQSLLYENTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 127) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.15I KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.158 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.162 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.165 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.166 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.167 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.168 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.170 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.171 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.174 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.185 KSSQSSLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 132) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.208 KSSQSLLGSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 133) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.231 KSSQSLLYSNIQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 134) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.235 KSSQSLLYSNHQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 135) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WHI.250 KSSQSLLYSNTQDNYLA WASTRES QQYSYYPPT (SEQ ID NO: 136) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.253 KSSQSLLYSNTQGNYLA WASTRES QQYSYYPPT (SEQ ID NO: 137) (SEQ ID NO: (SEQ ID NO: 124)123)Antibody CDR-L1 CDR-L2 CDR-L3 CD98hc.04.048. WH1.262 KSSQSLLYSNTQKNYLA WASTRES QQKSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 138)123)CD98hc.04.048. WH1.269 KSSQSLLYSNTQKNYLA WASTRES QQYKYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 139)123)CD98hc.04.048. WH1.271 KSSQSLLYSNTQKNYLA WASTRES QQYTYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 140)123)CD98hc.04.048. WH1.274 KSSQSLLYSNTQKNYLA WASTRES QQYEYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 141)123)CD98hc.04.048. WH1.276 KSSQSLLYSNTQKNYLA WASTRES QQYIYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 142)123)CD98hc.04.048. WH1.279 KSSQSLLYSNTQKNYLA WASTRES QQYSVYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 143)123)CD98hc.04.048. WH1.281 KSSQSLLYSNTQKNYLA WASTRES QQYSEYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 144)123)CD98hc.04.048. WH1.282 KSSQSLLYSNTQKNYLA WASTRES QQYSLYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 145)123)CD98hc.04.048. WH1.283 KSSQSLLYSNTQKNYLA WASTRES QQYSQYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 146)123)CD98hc.04.048. WH1.39 KSSQSLLYSNTQKNYLA WASTRES AQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 147)123)CD98hc.04.048. WH1.55 KSSQSLLYSNTQKNYLA WASTRES QQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 124)123)CD98hc.04.048. WH1.76 KSSQSLLYSNTQKNYLA WASTRES AQYSYYPPT (SEQ ID NO: 122) (SEQ ID NO: (SEQ ID NO: 147)123)TABLE 11: VH and VL Sequences of Anti-CD98hc Antibodies Antibody VH sequence VL sequence CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.15 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADEVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGTDNSKNTLYLQMNSLRAEDTAVYYAntibody VH sequence VL sequence CARQGGLPAWFVYWGQGTLVTVS DFTLTISSLQAEDVAVYYCQQYSYYS (SEQ ID NO: 150) PPTFGGGTKVEIK (SEQ ID NO: 149) CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.20 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADFVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 151)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.23 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADYVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 152)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.078 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYKSSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 153)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.083 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYKSSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 154)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.105 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSQGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 155)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.118 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGKGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 156)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.119 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGKGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 157)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.121 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGKGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149)S (SEQ ID NO: 158)Antibody VH sequence VL sequence CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.133 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGHTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 159)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.193 SGFGISAYDMSWVRQAPGKGLEW QSLKYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 160) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.217 SGFGISAYDMSWVRQAPGKGLEW QSLLYQNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 161) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.218 SGFGISAYDMSWVRQAPGKGLEW QSLLYENTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 162) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.222 SGFGISAYDMSWVRQAPGKGLEW QSLLYSHTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 163) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.224 SGFGISAYDMSWVRQAPGKGLEW QSLLYSTTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 164) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.232 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNSQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 165) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.236 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNKQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 166) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.296 SGFGISAYDMSWVRQAPGKGLEW QSLLYENTQKNYLAWYQQKPGQPPVSYISSQGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGTAntibody VH sequence VL sequence DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 162) S (SEQ ID NO: 155)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.151 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSDYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 167)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.158 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGSLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 168)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.162 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGHPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 169)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.165 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGAPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 170)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.166 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGTPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 171)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.167 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGQPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 172)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.168 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPALFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 173)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.170 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAKFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149)S (SEQ ID NO: 174)Antibody VH sequence VL sequence CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.171 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPATFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 175)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.174 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPARFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 149) S (SEQ ID NO: 176)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.185 SGFGISAYDMSWVRQAPGKGLEW QSSLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 177) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.208 SGFGISAYDMSWVRQAPGKGLEW QSLLGSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 178) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.231 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNIQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 179) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.235 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNHQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 180) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.250 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQDNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 181) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.253 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQGNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 182) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.262 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPPVSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGTAntibody VH sequence VL sequence DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQKSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 183) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.269 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYKYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 184) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.271 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYTYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 185) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.274 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYEYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 186) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.276 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYIYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 187) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.279 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSVY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 188) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.281 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSEY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 189) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.282 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSEY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 190) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.283 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCQQYSQY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 191)S (SEQ ID NO: 148)Antibody VH sequence VL sequence CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.39 SGFGISAYDMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCAQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 192) S (SEQ ID NO: 148)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMSQSPSSLAVSVGEKVTMSCKS WH1.55 SGFGISAYAMSWVRQAPGKGLEW SQSLLYSNTQKNYLAWYQQKPGQS VSYISSGGGSTYYADSVKGRFTISR PKLLIYWASTRESGVPDRFTGSGSG DNSKNTLYLQMNSLRAEDTAVYY TDFTLTISSVKAEDLAIYYCQQYSY CARQGGLPAWFVYWGQGTLVTVS YPPTFGGGTKLEIK (SEQ ID NO: 194) S (SEQ ID NO: 193)CD98hc.04.048. EVQLLESGGGLVQPGGSLRLSCAA DIVMTQSPDSLAVSLGERATINCKSS WH1.76 SGFGISAYAMSWVRQAPGKGLEW QSLLYSNTQKNYLAWYQQKPGQPP VSYISSGGGSTYYADSVKGRFTISR KLLIYWASTRESGVPDRFSGSGSGT DNSKNTLYLQMNSLRAEDTAVYY DFTLTISSLQAEDVAVYYCAQYSYY CARQGGLPAWFVYWGQGTLVTVS PPTFGGGTKVEIK (SEQ ID NO: 192)S (SEQ ID NO: 193)

[0305] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the six CDRs of an antibody listed in Tables 9, 10, and 11 (i.e. the three VH CDRs of the antibody listed in Table 9 and the three VL CDRs of the same antibody listed in Table 10).In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the six CDRs of an antibody listed in Tables 9, 10, and 11 as determined by Kabat numbering. In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the six Chothia CDRs of an antibody listed in Table 11.

[0306] In some aspects, the CDRs of an antigen-binding domain that specifically binds to human CD98hc can be determined according to MacCallum RM et al., (1996) J Mol Biol 262: 732-745. See also, e.g., Martin A. “Protein Sequence and Structure Analysis of Antibody Variable Domains,” in Antibody Engineering, Kontermann and Diibel, eds., Chapter 31, pp. 422- 439, Springer-Verlag, Berlin (2001). In some aspects, provided herein are antigen-binding domains that specifically bind to human CD98hc and comprise VH and VL CDRs of an antibody listed in Table 11 as determined by the method in MacCallum RM et al.

[0307] In some aspects, the CDRs of an antigen-binding domain that specifically binds to human CD98hc can be determined according to the AbM numbering scheme (as described above). In some aspects, provided herein are antigen-binding domains that specifically bind to human CD98hc and comprise VH and VL CDRs of an antibody listed in Table 11 as determined by the AbM numbering scheme.

[0308] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the six IMGT CDRs of an antibody listed in Table 11 according to the IMGT numbering system as described above.

[0309] In some aspects, an antigen-binding domain that specifically binds to human CD98hc provided herein is described by its VL domain alone, or its VH domain alone, or by its 3 VL CDRs alone, or its 3 VH CDRs alone.

[0310] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the VH of an antibody listed in Table 11.

[0311] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the VL of antibody listed in Table 11.

[0312] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the VH and the VL of an antibody listed in Table 11.

[0313] In another aspect, an antigen-binding domain of the present disclosure that specifically binds to human CD98hc comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:148, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, and 193. In certain aspects, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 148, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, and 193 contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an antigen-binding domain that specifically binds to human CD98hc comprising that sequence retains the ability to bind to human CD98hc. In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted in SEQ ID NO: 148, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, or 193. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 148, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, or 193. In certain aspects, substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the antigen-binding domain that specifically binds to human CD98hc comprises the VH sequence of SEQ ID NO: 148, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, and 193, including post- post-translational modifications of that sequence.

[0314] In another aspect, an antigen-binding domain of the present disclosure that specifically binds to human CD98hc comprises comprises a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:149, 160, 161, 162, 163, 164, 165, 166, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, and 194. In certain aspects, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to an amino acid sequence selected from the group consisting of SEQ IDNOs: 149, 160, 161, 162, 163, 164, 165, 166, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, and 194 and contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an antigen-binding domain of the present disclosure that specifically binds to human CD98hc comprising that sequence retains the ability to bind to human CD98hc. In some aspects, a total of 1 to 10 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 149, 160, 161, 162, 163, 164, 165, 166, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, or 194. In certain aspects, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 149, 160, 161, 162, 163, 164, 165, 166, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, or 194. In certain aspects, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the antigen-binding domain of the present disclosure that specifically binds to human CD98hc comprises the VL sequence of SEQ ID NO: 149, 160, 161, 162, 163, 164, 165, 166, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, and 194, including post-post-translational modifications of that sequence.

[0315] In some aspects, an antigen-binding domain of the present disclosure that specifically binds to human CD98hc is provided, wherein the antibody comprises a VH as in any of the aspects provided above, and a VL as in any of the aspects provided above. In some aspects, the antigen-binding domain comprises the VH and Vrsequences in SEQ ID NOs: 148, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, and 193, and SEQ ID NOs: 149, 160, 161, 162, 163, 164, 165, 166, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, and 194, respectively, including post-post-translational modifications of those sequences.

[0316] In some aspects, provided herein are antigen-binding domains that specifically bind to human TfR comprising a heavy chain variable domain (VH) and a light chain variable domain(VL), wherein the VH and VL are selected from the group consisting of: VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 150 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 151 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 152 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 153 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 154 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 155 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 156 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 157 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 158 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 159 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 160; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 161; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 162; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 163; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 164; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 165; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 166; VH comprising the amino acid sequence of SEQ ID NO: 155 and VL comprising the amino acid sequence of SEQ ID NO: 162; VH comprising the amino acid sequence of SEQ ID NO: 167 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 168 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 169 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 170 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ IDNO: 171 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 172 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 173 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 174 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 175 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 176 and VL comprising the amino acid sequence of SEQ ID NO: 149; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 177; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 178; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 179; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 180; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 181; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 182; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 183; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 184; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 185; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 186; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 187; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 188; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 189; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 190; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 191; VH comprising the amino acid sequence of SEQ ID NO: 148 and VL comprising the amino acid sequence of SEQ ID NO: 192; VH comprising the amino acid sequence of SEQ ID NO: 193 and VL comprising the amino acid sequence of SEQ ID NO: 194; and VH comprising the amino acid sequence of SEQ ID NO: 193 and VL comprising the amino acid sequence of SEQ ID NO: 192.

[0317] In some aspects, provided herein is an antigen-binding domain that competitively inhibits binding to CD98hc as an antibody comprising a VH amino acid sequence of an antibody in Table 11 and comprising a VL amino acid sequence of the same antibody in Table 11.

[0318] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises a VH and a VL on a single polypeptide chain (e.g., a VH and VL in Table 11). In some aspects, the antigen-binding domain comprises an scFv. The scFv can comprise a VH that is N-terminal to a VL or a VL that is N-terminal to a VH. The scFv can comprise a linker, e.g., between a VH and a VL. Accordingly, the scFv can be in the orientation VH-linker-VL or VL-linker-VH. Such a linker can be about 5 to about 25 amino acids in length. Such a linker can be about 5 to about 20 amino acids in length. Such a linker can be about 10 to about 25 amino acids in length. Such a linker can be about 10 to about 20 amino acids in length. Such a linker can be, e.g., a glycine linker, a glycine-rich linker, or a glycine-serine linker. Such a linker can comprise the amino acid sequence of GGSEGKSSGSGSESKSTGGS (SEQ ID NO: 209). Such a linker can comprise the amino acid sequence of GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 210).

[0319] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises a VH on a first polypeptide and a VL on a second polypeptide (e.g., a Fab).

[0320] In some aspects, an antigen-binding domain that specifically binds to human CD98hc comprises the antigen-binding fragment of a heavy chain only antibody (e.g., a VHH or nanobody).

[0321] In some aspects, an antigen-binding domain that specifically binds to human CD98hc is a murine antigen-binding domain. In some aspects, an antigen-binding domain that specifically binds to human CD98hc is a chimeric antigen-binding domain. In some aspects, an antigenbinding domain that specifically binds to human CD98hc is a humanized antigen-binding domain. In some aspects, an antigen-binding domain that specifically binds to human CD98hc is a human antigen-binding domain.

[0322] In some aspects, an antigen-binding domain provided herein that specifically binds to human CD98hc also binds to cynomolgus monkey CD98hc.

[0323] In some aspects, the antigen-binding domain provided herein binds human CD98hc with an affinity between about 500 nM and about 10,000 nM. In some aspects, the antigenbinding domain binds human CD98hc with an affinity between about 500 nM and about 5,000 nM, between about 500 nM and about 2,500 nM, between about 500 nM and about 1,000 nM, between about 500 nM and about 750 nM, between about 1,000 nM and about 2,5000 nM,between about 2,500 nM and about 5,000 nM, between about 2,500 nM and about 10,000 nM, between about 5,000 nM and about 10,000 nM, and values and ranges there between.

[0324] In some aspects, an antigen-binding domain provided herein binds human CD98hc with an affinity between about 10 nM and about 500 nM. In some aspects, the antigen-binding domain binds human CD98hc with an affinity between about 10 nM and about 250 nM, between about 10 nM and about 100 nM, between about 10 nM and about 50 nM, between about 50 nM and about 500 nM, between about 50 nM and about 250 nM, between about 100 nM and about 250 nM, between about 250 nM and about 500 nM, and values and ranges there between.

[0325] In some aspects, the antigen-binding domain binds human CD98hc with an affinity less than about 10 nM. In some aspects, the antigen -binding domain binds human CD98hc with an affinity between about 1 nM and about 10 nM, between about 1 nM and about 5 nM, between about 0.1 nM and about 1 nM, between about 0.01 nM and about 0.1 nM, and values and ranges there between.

[0326] In some aspects, an antigen-binding domain provided herein that specifically binds to human CD98hc binds to human CD98hc is measured using surface plasmon resonance. Surface plasmon resonance can be measured, e.g., using the Carterra LSA platform. In some aspects, an antigen-binding domain provided herein that specifically binds to human CD98hc binds to human CD98hc as measured by, for example, radioimmunoassay (RIA), Western blot, or ELISA OD450.

[0327] Also provided herein are antigen-binding domains that bind to the same epitope of CD98hc as a CD98hc antigen-binding domain provided herein. Also provided herein are antigenbinding domains that competitively inhibit binding to CD98hc of CD98hc antigen-binding domain provided herein.Multi-specific Binding Proteins that Comprise an Anti-pyroGlu3 Abeta (A0) Antigen-Binding Domain and an Antigen-Binding Domain that Binds to human TfR

[0328] Provided herein are multi-specific binding proteins that comprise an antigen-binding domain that specifically binds to human anti-pyroGlu3 Abeta (A0) and an antigen-binding domain that specifically binds to human TfR.

[0329] In some aspects, an antibody or antigen-binding fragment thereof provided herein comprises an antigen-binding domain that specifically binds to human TfR. In some aspects, an antibody or antigen-binding fragment thereof comprises an antigen-binding domain thatspecifically binds to human TfR linked to a second antigen-binding domain that specifically binds to human pyroGlu3 Abeta (A0). Also provided herein are antibodies or antigen-binding fragments thereof that bind to the same epitope of TfR as a TfR antigen-binding domain provided herein. Also provided herein are antibodies or antigen-binding fragments thereof that competitively inhibit binding to TfR of a TfR antigen-binding domain provided herein.

[0330] In some aspects, a multi-specific binding protein provided herein comprises a first antigen-binding domain that specifically binds to human TfR linked to a second antigen-binding domain that specifically binds to human pyroGlu3 Abeta (A0). The first antigen-binding domain that specifically binds to human TfR can be any antigen-binding domain that binds to human TfR provided herein. The second antigen-binding domain can be any antigen-binding domain that specifically binds to human pyroGlu3 Abeta (A0) provided herein.

[0331] In some aspects, a multi-specific binding protein provided herein comprises an antigenbinding domain that specifically binds to human TfR that is linked to a second antibody or antigen-binding fragment thereof. The second antibody or antigen-binding fragment thereof specifically binds to human pyroGlu3 Abeta (A0). In some aspects, the second antibody or antigen-binding fragment does not bind to TfR.

[0332] In some aspects, a multi-specific binding protein provided herein is bi-specific. In some aspects, a multi-specific binding protein provided herein is bivalent, trivalent, or tetravalent.

[0333] In some aspects, a multi-specific binding protein provided herein comprises a 2+1 multi-specific protein format (as shown in Figure 1A and Figure IB) or a 2+2 multispecificprotein format (as shown in Figure 2). As used herein, “2+1” refers to the multi-specific protein format in Figure 1A or Figure IB.

[0334] In some aspects, a multi-specific binding protein provided herein comprises a TfR antigen-binding domain that is an scFv linked to an antibody that binds to human pyroGlu3 Abeta (A0), wherein the antibody that binds to human pyroGlu3 Abeta (A0) comprises two heavy chains and two light chains. In some aspects, the scFv is linked to the C-terminus of one of the two antibody heavy chains, e.g., via a protein linker.

[0335] In some aspects, the multi-specific binding protein comprises 1) an antigen-binding domain that specifically binds TfR, 2) a second antigen-binding domain that specifically binds to human pyroGlu3 Abeta (A0), and 3) an Fc region, wherein the TfR antigen-binding domain and the second antigen-binding domain are connected or linked to the Fc region of the multi-specific binding protein. In other aspects, the multi-specific binding protein comprises 1) an antigen-binding domain that comprises a heavy chain variable region and specifically binds TfR, 2) a second antigen binding domain that comprises a heavy-chain variable region and specifically binds to human pyroGlu3 Abeta (A0), and 3) an Fc region, wherein the TfR antigen-binding domain and the second antigen-binding domain are connected or linked to the Fc region of the multi-specific binding protein.

[0336] In some aspects, the multi-specific binding protein comprises 1) an antigen-binding domain that specifically binds TfR, 2) a second antigen-binding domain that binds to human pyroGlu3 Abeta (A0), and 3) an Fc region, wherein the antigen-binding domain that specifically binds TfR and the second antigen-binding domain are both connected or linked to the N-terminus of the Fc portion of the multi-specific binding protein. In other aspects, the antigen-binding domain that specifically binds TfR is connected or linked to the N-terminus of an Fc portion of the multi-specific protein and the second antigen-binding domain is linked to the C-terminus of the Fc portion of the multi -specific binding protein. In other aspects, the antigen-binding domain that specifically binds TfR is connected or linked to the C-terminus of an Fc portion of the multispecific binding protein and the second antigen-binding domain is linked to the N-terminus of the Fc portion of the multi-specific binding protein.

[0337] In some aspects, a multi-specific binding protein provided herein comprises two scFv molecules that comprise antigen-binding domains that specifically bind TfR and an antibody that binds to human pyroGlu3 Abeta (A0), wherein the antibody comprises two heavy chains and two light chains, and wherein one of the two scFv molecules is linked to the C-terminus of one of the antibody heavy chains, and wherein the other scFv molecule is linked to the C-terminus of the other antibody heavy chain. In some aspects, the scFvs are linked to the heavy chains of the antibody via a protein linker.

[0338] In some aspects, disclosed herein is a trivalent, bi-specific protein comprising (i) a single scFv, Fab or VHH antigen-binding domain that specifically binds to human TfR and (ii) an antibody that specifically binds to human pyroGlu3 Abeta (A0), wherein the antibody comprises two heavy chains and two light chains, and wherein the single scFv, Fab or VHH antigen-binding domain that specifically binds to human TfR is linked to the C-terminus or N-terminus of one of the two antibody heavy chains. The Fc may be a heterodimeric Fc, such as knob and hole Fc. An example of this 2+1 format with a knob-hole Fc is shown as (i) in Figure 3.

[0339] In some aspects, disclosed herein is a tetravalent, bi-specific protein comprising (i) two scFv, Fab or VHH antigen-binding domains that specifically bind to human TfR and (ii) anantibody that binds to human pyroGlu3 Abeta (A0), wherein the antibody comprises two heavy chains and two light chains, and wherein one of the scFv, Fab, or VHH antigen-binding domains that specifically binds to human TfR is linked to the C-terminus of one of the two antibody heavy chains, and the other scFv, Fab, or VHH antigen-binding domain that specifically binds to human TfR is linked to the C-terminus or N-terminus of the other of the two antibody heavy chains. In some aspects, the two scFv, Fab or VHH antigen-binding domains that bind to human TfR can comprise the same amino acid sequence. In some aspects, the Fc is a heterodimeric Fc, such as an Fc region with knob and hole mutations.

[0340] In some aspects, disclosed herein is a bivalent, bispecific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that specifically binds to human TfR linked to the N-terminus of an Fc domain of an Fc dimer, and (ii) a second antigen binding domain that binds to human pyroGlu3 Abeta (A0), wherein the second antigen binding domain is linked to the N-terminus of the second Fc domain of an Fc dimer. An example of this format is shown as (iii) in Figure 3.

[0341] In some aspects, disclosed herein is a bivalent, bi-specific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that specifically binds to human TfR linked to the N-terminus or C-terminus of an Fc domain of an Fc dimer and (ii) a second antigen binding domain, wherein the second antigen binding domain binds to human pyroGlu3 Abeta (A0) and comprises a single scFv, VHH or Fab linked to N-terminus of the Fc. In some aspects, disclosed herein is a bivalent, bi-specific protein comprising (i) a single scFv, VHH, or Fab antigenbinding domain that specifically binds to human TfR linked to the N-terminus or C-terminus of an Fc domain of an Fc dimer and (ii) a second antigen binding domain, wherein the second antigen binding domain binds to human pyroGlu3 Abeta (A0) and comprises a single scFv, VHH or Fab linked to the C-terminus of the second Fc domain of the Fc dimer. In some aspects, the Fc is a heterodimeric Fc, such as an Fc domain with knob and hole mutations. An example of this format is shown as (ii) in Figure 3, wherein a single scFv is linked to the N terminus of a first Fc domain of the Fc region and the second antigen binding domain is linked to the N terminus of the second Fc domain of the Fc region.

[0342] In some aspects, disclosed herein is a bivalent, bispecific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that specifically binds to human TfR and is linked to the N-terminus of an Fc domain of an Fc dimer, and (ii) a second antigen binding domain thatbinds to human pyroGlu3 Abeta (A0) and is linked to the N-terminus of the second Fc domain of an Fc dimer. An example of this format is shown as (iii) in Figure 3.

[0343] In some aspects, disclosed herein is a bivalent, bi-specific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that binds to human TfR and is linked to the N-terminus or C-terminus of an Fc domain of an Fc dimer and (ii) a second antigen binding domain, wherein the second antigen binding domain binds to human pyroGlu3 Abeta (A0) and comprises a single scFv, VHH or Fab linked to N-terminus of the Fc or C-terminus of a second Fc domain of the Fc dimer. In some aspects, the Fc is a heterodimeric Fc, such as an Fc domain with knob and hole mutations. An example of this 1+1 format is shown as (ii) in Figure 3, where a single scFv is linked to the N terminus of an Fc domain of an Fc region and a second antigen binding domain is linked to the N terminus of a second Fc domain of the Fc region.

[0344] As provided herein, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein can be multi-specific, e.g., bi-specific. Many different formats and uses of bi-specific binding molecules are known in the art (reviewed in, e.g., Kontermann; Drug Discov Today, 2015 July; 20(7):838-47; MAbs, 2012 March-April; 4(2): 182-97). A bispecific protein according to the present disclosure is not limited to any particular bispecific format or method of producing it. Accordingly, a bispecific protein of the present disclosure can include various configurations having a first antigen-binding domain that binds to human TfR and a second antigen-binding domain or antibody or antigen-binding fragment thereof that binds to human pyroGlu3 A0.

[0345] Bispecific molecules include, e.g., a kappa-lambda body, a dual-affinity re-targeting molecule (DART), a knob-in-hole antibody, a strand-exchange engineered domain body (SEEDbody), and a DuoBody. In some aspects, a bispecific antibody or antigen-binding fragment thereof, or multi-specific protein provided herein comprises a knob mutation and a hole mutation. In some aspects, the knob mutation comprises the amino acid substitution T366W according to EU numbering. In some aspects, the hole mutation comprises the amino acids substitutions T366S, L368A, and Y407V according to EU numbering. Examples of bispecific molecules that can be used in the present disclosure include (i) a single antibody that has two arms comprising different antigen-binding domains; (ii) a single chain antibody that has specificity to two different epitopes, e.g., via two scFvs linked in tandem by an extra peptide linker; (iii) a dual-variable-domain antibody (DVD-Ig), where each light chain and heavy chain contains two variable domains in tandem through a short peptide linkage (Wu et al., Generationand Characterization of a Dual Variable Domain Immunoglobulin (DVD-Ig. TM.) Molecule, In: Antibody Engineering, Springer Berlin Heidelberg (2010)); (iv) a chemically-linked bispecific (Fab’)2 fragment; (v) a Tandab, which is a fusion of two single chain diabodies resulting in a tetravalent bispecific antibody that has two binding sites for each of the target antigens; (vi) a flexibody, which is a combination of scFvs with a diabody resulting in a multivalent molecule; (vii) a so-called “dock and lock” molecule, based on the “dimerization and docking domain” in Protein Kinase A, which, when applied to Fabs, can yield a trivalent bispecific binding protein consisting of two identical Fab fragments linked to a different Fab fragment; (viii) a so-called Scorpion molecule, comprising, e.g., two scFvs fused to both termini of a human Fab-arm; and (ix) a diabody. Other examples of antibody structures are described in WO2019 / 246288, which is incorporated by reference.

[0346] In some aspects, a bispecific protein provided herein is selected from one of the following formats: CrossMab, DAF (two-in-one), DAF (four-in-one), DutaMab, DT-IgG, Charge pair, Fab-arm exchange, Triomab, LUZ-Y, Fcab, kappalambda-body, Orthogonal Fab, DVD-IgG, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L, H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv, scFv4-Ig, Zybody, DVI-IgG (four-in-one), Nanobody, Nanbody-HAS, BiTE, TandAb, scDiabody-CH3, Diabody-CH3, Triple Body, miniantibody, minibody, TriBi minibody, scFv-CH3 KIH, Fab-scFv, scFV-CH-CL-scFV, F(ab’)2, F(ab’)2-scFv2, scFV-KIH, Fab-scFv-Fc, Tetravalent HCAb, scDiabody-Fc, Diabody-Fc, Tandem scFv-Fc, Intrabody, ImmTAC, HSAbody, scDiabody-HAS, Tandem scFv-toxin, IgG-IgG, Cov-X-body, and scFvl-PEG-scFv2, as described in Spiess, C., et al., Alternative molecular formats and therapeutic applications for bispecific antibodies. Mol. Immunol. (2015), which is incorporated by reference herein.

[0347] In some aspects, a multi-specific binding protein provided herein is bivalent. In some aspects, a multi-specific binding protein provided herein is multivalent (e.g., bivalent). In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein is trivalent (e.g., in a 2+1 format). In some aspects, a trivalent format comprises a single TfR antigen-binding domain provided herein and two antigen-binding domains that bind to N3pE Abeta (AP). The two antigen-binding domains that bind to a CNS antigen or a brain antigen can comprise the same amino acid sequence or can comprise different amino acid sequences. In some aspects, the TfR antigen-binding domain is an scFv. In some aspects, the TfR antigen-binding domain is a VHH. In some aspects, the TfR antigen-binding domain is a Fab.

[0348] In some aspects, a multi-specific binding protein provided herein is tetravalent (e.g., in a 2+2 format). In some aspects, a tetravalent format comprises two TfR antigen-binding domains provided herein and two antigen-binding domains that bind to a CNS antigen or a brain antigen. The two TfR antigen-binding domains can comprise the same amino acid sequence or can comprise different amino acid sequences. In some aspects, the two TfR antigen-binding domains comprise the same amino acid sequence. In some aspects, one or both of the TfR antigen-binding domains is an scFv. The two antigen-binding domains that bind to a N3pE Abeta (AP) antigen can comprise the same amino acid sequence or can comprise different amino acid sequences

[0349] An antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein can comprise a linker. The linker can be e.g., a glycine linker, a glycine-rich linker, or a glycine-serine linker. The linker can comprise the amino acid sequence (GGGGS)x3 (SEQ ID NO: 207). The linker can comprise the amino acid sequence (GGSGG)x3 (SEQ ID NO: 208).

[0350] An antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein can comprise a constant region. In some aspects, a TfR antigen-binding domain provided herein is linked to the constant region, e.g., the C-terminus of the constant region. In some aspects, a constant domain is a human constant domain. In some aspects, a constant domain is a murine, rat, rabbit, or monkey (e.g., cynomolgus) constant domain. The constant region can be a heavy chain constant region. The constant region can be a human constant region. The constant region can be a human heavy chain constant region. The constant region can be an IgG constant region. The constant region can be an IgGl constant region. The constant region can be an IgG2 constant region. The constant region can be an IgG4 constant region. The constant region can be a human IgG constant region. The constant region can be a human IgGl constant region. The constant region can be a human IgG2 constant region. The constant region can be a human IgG4 constant region. The linker can comprise the amino acid sequence GGSGG (no repeats) (SEQ ID NO: 214). The linker may be 1 to 20 amino acids in length.

[0351] In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein comprises a heavy chain and a light chain. With respect to the heavy chain, in some aspects, the heavy chain of an antigen-binding protein described herein can be an alpha (a), delta (8), epsilon (a), gamma (y) or mu (p) heavy chain. In some aspects, the heavy chain can comprise a human alpha (a), delta (6), epsilon (a), gamma (y) or mu (p) heavy chain. In some aspects, the heavy chain comprises a human gamma (y) heavy chain constantregion. In some aspects, the heavy chain of comprises the amino acid sequence of an IgGl heavy chain constant region. In some aspects, the heavy chain comprises the amino acid sequence of an IgG2 (e.g., IgG2a or IgG2b) heavy chain constant region. In some aspects, the heavy chain comprises the amino acid sequence of an IgG4 heavy chain constant region. With respect to the light chain, in some aspects, the light chain is a kappa light chain. In some aspects, the light chain is a lambda light chain. In some aspects, the light chain is a human kappa light chain or a human lambda light chain.

[0352] In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein comprises constant regions comprising the amino acid sequences of the constant regions of an IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, or a human IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule. In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein comprises constant regions comprising the amino acid sequences of the constant regions of an IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, any class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2), or any subclass (e.g., IgG2a and IgG2b) of immunoglobulin molecule. In some aspects, the constant regions comprise the amino acid sequences of the constant regions of a human IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, any class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2), or any subclass (e.g., IgG2a and IgG2b) of immunoglobulin molecule.

[0353] Non-limiting examples of human constant region sequences have been described in e.g., U. S. Patent No. 5,693,780 and Kabat EA etal., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U. S. Department of Health and Human Services, NIH Publication No. 91-3242).

[0354] In some aspects, a constant region provided herein comprises a knob mutation. In some aspects, a constant region provided herein comprises a hole mutation. Accordingly, in some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein can comprise a constant region comprising a knob mutation and a constant region comprising a hole mutation. Description of knob and hole mutations are provided herein.Variations in Linker Length Between the C-terminus of an anti-pyroGlu3 Abeta (A0) antibody heavy chain Fc and an Antigen-Binding Domain that Binds to TfR

[0355] As described herein, certain anti-TfR antibodies can block binding of transferrin to its receptor (TfRl) on the surface of cells, interfering with iron transport. Disruption in iron transport into and uptake by erythroid progenitor cells (reticulocytes) can lead to a reduction in hematocrit, hemoglobin levels, and red blood cell count, resulting in anemia. Both antibody dependent cellular cytotoxicity (ADCC) and complement dependent cytotoxicity (CDC) have been described to also be involved in these adverse hemotologic processes due to Fc-mediated cytotoxic effects driven by Fc gamma receptor (FcyR) binding to the Fc region of the antibody. Eliminating or reducing the Fc effector function of an antibody (such as by mutating certain amino acid residues within the Fc region of the antibody) in order to reduce or eliminate ADCC and / or CDC is a potential mitigation strategy for the development of TfRl-based therapies designed to cross the blood-brain barrier in order to reduce such hematologic cytotoxic and adverse effects (Couch et al (2013), Sci Transl Med 5:183ra57).

[0356] However, changes in certain amino acid residues within the Fc region of an anti-A0 antibody in order to reduce ADCC and / or CDC can reduce or eliminate the effector function of the antibody when it is bound to its target. As such, Fc mutations resulting in reduced ADCC and / or CDC have the risk of reducing the full therapeutic efficacy of an anti-Abeta (A0) antibody. For instance, one mechanism of brain clearance of Abeta (A0) by therapeutic anti-Abeta (A0) antibodies is associated with the Fc effector function of the antibody, enabling recruitment of myeloid cells in the brain (e.g., microglial cells), which express Fc gamma receptors (FcyR), to aid in effective and efficient removal of Abeta (A0) plaques (See, e.g., Condello et al, 2015, Nat Commun 6:6176). Accordingly, in certain uses of anti-Abeta (A0) antibodies for treatment of various Abeta (A0)-associated brain disorders, such as Alzheimer’s disease, an antibody having Fc-mediated effector function is desirable in order to achieve the beneficial clinical effects.

[0357] Based on these observations, it is therefor desirable to have an effective anti-pyroGlu3 Abeta (A0) antibody comprising an antigen-binding domain that binds TfR to facilitate transport across the BBB that reduces hematologic adverse events mediated by the Fc effector function of the antibody, while still maintaining Fc effector function of the anti-pyroGlu3 Abeta (A0) antibody in the brain. Without wishing to be bound by this theory, reducing the length of the linker located between the C-terminal end of the anti-pyroGlu3 Abeta (A0) antibody heavy chain (e.g., antibody heavy chain constant domain or antibody heavy chain Fc) and the anti-TfR antigen binding domain (e.g., scFv in 2+1 format) is contemplated to position the anti-pyroGlu3Abeta (A0) antibody Fc region further into the TfR binding site upon anti-TfR scFv binding (or by reducing the flexibility / increasing the rigidity of the molecule), thereby preventing its ability to bind to FcyRs on immune cells when bound to, e.g., reticulocytes, thus reducing or eliminating the ADCC-mediated adverse hematologic events. In this way, the interaction between the anti-Abeta (A0) antibody Fc domain and FcyRs, which is necessary for the Fc-mediated ADCC response to occur, is prevented or reduced when the anti-Abeta (A0) / TfR composition is bound to reticulocytes. Additionally, it is contemplated that removal of the linker located between the C-terminusof the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain (e.g., scFv in 2+1 format) can create a more rigid, less flexible molecule that further helps reduce or eliminate ADCC-mediated adverse hematologic events associated with binding to FcyRs on immune cells.

[0358] Accordingly, in certain aspects, the present disclosure provides compositions comprising an anti-pyroGlu3 Abeta (A0) antibody having a fully active Fc region e.g., a human IgGl having a fully active Fc region) and a TfR antigen binding domain that can be transported across the blood brain barrier, wherein the TfR antigen binding domain is linked to the C-terminal region of the anti-pyroGlu3 Abeta (A0) antibody heavy chain (e.g., antibody heavy chain constant domain or antibody heavy chain Fc), wherein the composition reduces or eliminates ADCC-mediated adverse hematologic events associated with binding to FcyRs on immune cells when the composition is bound to reticulocytes. (See Example 29, Tables 23 and 24.)

[0359] In certain anti-pyroGlu3 Abeta (A0) antigen-binding domain / anti-TfR antigen binding domain compositions of the present disclosure, the linker located between the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv comprises the 5 amino acid linker GGSGG (SEQ ID NO: 214). In some aspects, the linker located between the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv comprises SEQ ID NO: 280. In some aspects, the linker located between the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv is shortened to 1-4 amino acids. In certain aspects, there is no linker locatedbetween the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv (e.g., SEQ ID NO: 281) (e.g., the C-terminus of the heavy chain Fc of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv are directly linked). In certain aspects, the N-terminal amino acid residues QV of the anti-TfR antigen binding domain scFv are removed, further reducing the length between the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv.

[0360] In certain aspects, the amino acid linker between the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody and the N-terminus of the anti-TfR antigen binding domain scFv may be shortened, for example, to 1-5 amino acids. To further increase the rigidity, the C-terminus of the anti-pyroGlu3 Abeta (A0) antibody heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) may be truncated and the N-terminus of the anti-TfR antigen binding domain may be truncated. For example, the amino acid sequence PGK may be removed from the C-terminus of the heavy chain (e.g., heavy chain constant domain or antibody heavy chain Fc) of the anti-pyroGlu3 Abeta (A0) antibody, and the amino acid sequence QV may be removed from the N-terminus of the anti-TfR antigen binding domain (e.g., SEQ ID NO:282).Multi-specific Binding Proteins that Comprise an Anti-pyroGlu3 Abeta (A0) Antigen-Binding Domain and an Antigen-Binding Domain that Binds to human CD98hc

[0361] Provided herein are multi-specific binding proteins that comprise an antigen-binding domain that specifically binds to human anti-pyroGlu3 Abeta (A0) and an antigen-binding domain that specifically binds to human CD98hc.

[0362] In some aspects, an antibody or antigen-binding fragment thereof provided herein comprises an antigen-binding domain that specifically binds to human CD98hc. In some aspects, an antibody or antigen-binding fragment thereof comprises an antigen-binding domain that specifically binds to human CD98hc linked to a second antigen-binding domain that specifically binds to human pyroGlu3 Abeta (A0). Also provided herein are antibodies or antigen-binding fragments thereof that bind to the same epitope of CD98hc as a CD98hc antigen-binding domainprovided herein. Also provided herein are antibodies or antigen-binding fragments thereof that competitively inhibit binding to CD98hc of a CD98hc antigen-binding domain provided herein.

[0363] In some aspects, a multi-specific binding protein provided herein comprises a first antigen-binding domain that specifically binds to human CD98hc linked to a second antigenbinding domain that specifically binds to human pyroGlu3 Abeta (A0). The first antigen-binding domain that specifically binds to human CD98hc can be any antigen-binding domain that binds to human CD98hc provided herein. The second antigen-binding domain can be any antigenbinding domain that specifically binds to human pyroGlu3 Abeta (A0) provided herein.

[0364] In some aspects, a multi-specific binding protein provided herein comprises an antigenbinding domain that specifically binds to human CD98hc that is linked to a second antibody or antigen-binding fragment thereof. The second antibody or antigen-binding fragment thereof specifically binds to human pyroGlu3 Abeta (A0). In some aspects, the second antibody or antigen-binding fragment does not bind to CD98hc.

[0365] In some aspects, a multi-specific binding protein provided herein comprises a 2+1 multi-specific protein format (as shown in Figure 1A and Figure IB) or a 2+2 multi-specific protein format (as shown in Figure 2).

[0366] In some aspects, a multi-specific binding protein provided herein comprises a CD98hc antigen-binding domain that is an scFv linked to an antibody that binds to Abeta (pyroGlu3 A0), wherein the antibody comprises two heavy chains and two light chains. In some aspects, the scFv is linked to the C-terminus of one of the two antibody heavy chains, e.g., via a protein linker.

[0367] In some aspects, the multi-specific binding protein comprises 1) an antigen-binding domain that specifically binds CD98hc, 2) a second antigen-binding domain that specifically binds to human pyroGlu3 Abeta (A0), and 3) an Fc region, wherein the CD98hc antigen-binding domain and the second antigen-binding domain are connected or linked to the Fc region of the multi-specific protein. In other aspects, the multi-specific protein comprises 1) an antigenbinding domain that comprises a heavy chain variable region and specifically binds CD98hc, 2) a second antigen binding domain that comprises a heavy-chain variable region and binds to human pyroGlu3 Abeta (A0), and 3) an Fc region, wherein the CD98hc antigen-binding domain and the second antigen -binding domain are connected or linked to the Fc region of the multi-specific protein.

[0368] In some aspects, the multi-specific binding protein comprises 1) an antigen-binding domain that specifically binds CD98hc, 2) a second antigen-binding domain that binds to humanpyroGlu3 Abeta (A0), and 3) an Fc region, wherein the antigen-binding domain that specifically binds CD98hc and the second antigen-binding domain are both connected or linked to the N-terminus of the Fc portion of the multi-specific protein. In other aspects, the antigen -binding domain that specifically binds CD98hc is connected or linked to the N-terminus of an Fc portion of the multi-specific protein and the second antigen-binding domain is linked to the C-terminus of the Fc portion of the multi-specific protein. In other aspects, the antigen-binding domain that specifically binds CD98hc is connected or linked to the C-terminus of an Fc portion of the multispecific protein and the second antigen-binding domain is linked to the N-terminus of the Fc portion of the multi-specific protein.

[0369] In some aspects, a multi-specific binding protein provided herein comprises two scFv molecules that comprise antigen-binding domains that specifically bind CD98hc and an antibody that binds to human pyroGlu3 Abeta (A0), wherein the antibody comprises two heavy chains and two light chains, and wherein one of the two scFv molecules is linked to the C-terminus of one of the antibody heavy chains, and wherein the other scFv molecule is linked to the C-terminus of the other antibody heavy chain. In some aspects, the scFvs are linked to the heavy chains of the antibody via a protein linker.

[0370] In some aspects, disclosed herein is a trivalent, bi-specific protein comprising (i) a single scFv, Fab or VHH antigen-binding domain that specifically binds to human CD98hc and (ii) an antibody that specifically binds to human pyroGlu3 Abeta (A0), wherein the antibody comprises two heavy chains and two light chains, and wherein the single scFv, Fab or VHH antigen-binding domain that specifically binds to human CD98hc is linked to the C-terminus or N-terminus of one of the two antibody heavy chains. The Fc may be a heterodimeric Fc, such as knob and hole Fc. An example of this 2+1 format with a knob-hole Fc is shown as (i) in Figure 3.

[0371] In some aspects, disclosed herein is a tetravalent, bi-specific protein comprising (i) two scFv, Fab or VHH antigen-binding domains that specifically bind to human CD98hc and (ii) an antibody that binds to human pyroGlu3 Abeta (A0), wherein the antibody comprises two heavy chains and two light chains, and wherein one of the scFv, Fab, or VHH antigen-binding domains that specifically binds to human CD98hc is linked to the C-terminus of one of the two antibody heavy chains, and the other scFv, Fab, or VHH antigen-binding domain that specifically binds to human CD98hc is linked to the C-terminus or N-terminus of the other of the two antibody heavy chains. In some aspects, the two scFv, Fab or VHH antigen-binding domains that bind to humanCD98hc can comprise the same amino acid sequence. In some aspects, the Fc is a heterodimeric Fc, such as an Fc region with knob and hole mutations.

[0372] In some aspects, disclosed herein is a bivalent, bispecific protein comprising (i) a single scFv, VHH, or Fab antigen -binding domain that specifically binds to human CD98hc linked to the N-terminus of an Fc domain of an Fc dimer, and (ii) a second antigen binding domain that binds to human pyroGlu3 Abeta (A0), wherein the second antigen binding domain is linked to the N-terminus of the second Fc domain of an Fc dimer. An example of this 1+1 format is shown as (iii) in Figure 3.

[0373] In some aspects, disclosed herein is a bivalent, bi-specific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that specifically binds to human CD98hc linked to the N-terminus or C-terminus of an Fc domain of an Fc dimer and (ii) a second antigen binding domain, wherein the second antigen binding domain binds to human pyroGlu3 Abeta (A0) and comprises a single scFv, VHH or Fab linked to N-terminus of the Fc. In some aspects, disclosed herein is a bivalent, bi-specific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that specifically binds to human CD98hc linked to the N-terminus or C-terminus of an Fc domain of an Fc dimer and (ii) a second antigen binding domain, wherein the second antigen binding domain binds to human pyroGlu3 Abeta (A0) and comprises a single scFv, VHH or Fab linked to the C-terminus of the second Fc domain of the Fc dimer. In some aspects, the Fc is a heterodimeric Fc, such as an Fc region with knob and hole mutations. An example of this 1+1 format is shown as (ii) in Fig. 3, wherein a single scFv is linked to the N terminus of a first Fc domain of the Fc region and the second antigen binding domain is linked to the N terminus of the second Fc domain of the Fc region.

[0374] In some aspects, disclosed herein is a bivalent, bispecific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that specifically binds to human CD98hc and is linked to the N-terminus of an Fc domain of an Fc dimer, and (ii) a second antigen binding domain that binds to human pyroGlu3 Abeta (A0) and is linked to the N-terminus of the second Fc domain of an Fc dimer. An example of this 1+1 format is shown as (iii) in Figure 3.

[0375] In some aspects, disclosed herein is a bivalent, bi-specific protein comprising (i) a single scFv, VHH, or Fab antigen-binding domain that binds to human CD98hc and is linked to the N-terminus or C-terminus of an Fc domain of an Fc dimer and (ii) a second antigen binding domain, wherein the second antigen binding domain binds to human pyroGlu3 Abeta (A0) and comprises a single scFv, VHH or Fab linked to N-terminus of the Fc or C-terminus of a secondFc domain of the Fc dimer. In some aspects, the Fc is a heterodimeric Fc, such as an Fc domain with knob and hole mutations. An example of this 1+1 format is shown as (ii) in Figure 3, where a single scFv is linked to the N terminus of an Fc domain of an Fc region and a second antigen binding domain is linked to the N terminus of a second Fc domain of the Fc region.

[0376] As provided herein, an antibody or antigen-binding fragment thereof, or multi-specific protein provided herein can be multi-specific, e.g., bi-specific. As described above, a bispecific protein of the present disclosure can include various configurations having a first antigen-binding domain that binds to human CD98hc and a second antigen-binding domain that binds to human pyroGlu3 Ab eta (A0).

[0377] In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein provided herein comprises a CD98hc binding domain and a knob mutation and a hole mutation. Knob and hole mutations are described in detail herein and similar apply to agents comprising CD98hc binding domains. In some aspects, the knob mutation comprises the amino acid substitution T366W according to EU numbering. In some aspects, the hole mutation comprises the amino acids substitutions T366S, L368A, and Y407V according to EU numbering. Examples of bispecific molecules that can be used are described herein.

[0378] In some aspects, bispecific protein formats are described above. In some aspects, an antibody or antigen-binding fragment thereof or multi-specific binding protein comprising a CD98hc binding domain and provided herein is bivalent or tetravalent. Formats comprising two CD98hc antigen-binding domains can comprise the same amino acid sequence or can comprise different amino acid sequences. In some aspects, the two CD98hc antigen-binding domains comprise the same amino acid sequence. In some aspects, one or both of the CD98hc antigenbinding domains is an scFv. The two antigen-binding domains that bind to a N3pE Abeta (AP) antigen can comprise the same amino acid sequence or can comprise different amino acid sequences.

[0379] An antibody or antigen-binding fragment thereof, or multi-specific binding protein comprising a CD98hc binding domain and provided herein can comprise a linker. The linker can be e.g., a glycine linker, a glycine-rich linker, or a glycine-serine linker. The linker can comprise the amino acid sequence (GGGGS)x3 (SEQ ID NO: 207). The linker can comprise the amino acid sequence (GGSGG)x3 (SEQ ID NO: 208).

[0380] An antibody or antigen-binding fragment thereof, or multi-specific binding protein comprising a CD98hc binding domain and provided herein can comprise a constant region, as described herein.

[0381] In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific binding protein comprising a CD98hc binding domain as provided herein comprises a heavy chain and a light chain, as described herein.

[0382] In some aspects, an antibody or antigen-binding fragment thereof, or multi-specific protein comprising a CD98hc binding domain as provided herein comprises constant regions comprising the amino acid sequences of the constant regions of an IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, or a human IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, or any class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2), or any subclass (e.g., IgG2a and IgG2b) of immunoglobulin molecule. Non-limiting examples of human constant regions and human constant region sequences are described herein.Multi-specific Binding Proteins that Comprise an Antigen-Binding Domain that Binds to PyroGlu3 Abeta (A0) and an Antigen-Binding Domain that Binds to Blood-Brain Barrier Receptors or Proteins

[0383] Provided herein are compositions that specifically bind to human receptors or proteins of the blood-brain barrier.

[0384] In some aspects, the composition comprises a multispecific protein comprising (i) an antigen-binding domain that specifically binds to human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc) and (ii) an isolated anti-pyroGlu3 Abeta (A0) antibody or antigen binding fragment thereof.

[0385] In some aspects, provided herein are multispecific proteins comprising (i) an antigenbinding domain that specifically binds to a blood brain barrier (BBB) target selected from human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc) and (ii) an antigen-binding domain that specifically binds to pyroGlu3 Abeta (A0).

[0386] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is an antibody. In some aspects, the antibody is a full-length antibody.

[0387] In some aspects, the antibody is an IgG antibody. In some aspects, the IgG antibody is an IgGl antibody or an IgG4 antibody.Ill

[0388] In some aspects, the anti-pyroGlu3 Abeta (A0) antibody comprises a heavy chain with a Fc hole mutation.

[0389] In some aspects, the anti-pyroGlu3 Abeta (A0) antibody comprises a heavy chain with a Fc knob mutation.

[0390] In some aspects, the anti-BBB antigen-binding domain is a scFv.

[0391] In some aspects, the anti-BBB scFv is linked to the C-terminus of the anti-pyroGlu3 A0 antibody heavy chain with a Fc hole mutation.

[0392] In some aspects, the anti-BBB scFv is linked to the C-terminus of the anti-pyroGlu3 A0 antibody heavy chain with a Fc knob mutation.

[0393] In some aspects, the multi specific protein comprises two antigen-binding domains that specifically bind to a BBB target selected from human TfR and human CD98hc. In some aspects, both of the anti-BBB antigen-binding domains are scFvs.

[0394] In some aspects, the first anti-BBB scFv is linked to the C-terminus of the anti-pyroGlu3 Abeta (A0) antibody heavy chain with a Fc hole mutation. In some aspects, the second anti-BBB scFv is linked to the C-terminus of the anti-pyroGlu3 Abeta (A0) antibody heavy chain with a Fc knob mutation.

[0395] In some aspects, the anti-BBB antigen-binding domain is an antibody. In some aspects, the antibody is a full-length antibody.

[0396] In some aspects, the antibody is an IgG antibody. In some aspects, the IgG antibody is an IgGl antibody or an IgG4 antibody.

[0397] In some aspects, the anti-BBB antibody comprises a heavy chain with a Fc hole mutation.

[0398] In some aspects, the anti-BBB antibody comprises a heavy chain with a Fc knob mutation.

[0399] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a scFv.

[0400] In some aspects, the anti-pyroGlu3 Abeta (A0) scFv is linked to the C-terminus of the anti-BBB antibody heavy chain with a Fc hole mutation.

[0401] In some aspects, the anti-pyroGlu3 Abeta (A0) scFv is linked to the C-terminus of the anti-BBB antibody heavy chain with a Fc knob mutation.

[0402] In some aspects, the multi specific protein comprises two antigen-binding domains that specifically bind to a pyroGlu3 Abeta (A0), wherein both of the anti-pyroGlu3 Abeta (A0) antigen-binding domains are scFvs.

[0403] In some aspects, the first anti-pyroGlu3 Abeta (A0) scFv is linked to the C-terminus of the anti-BBB antibody heavy chain with a Fc hole mutation. In some aspects, the second anti-pyroGlu3 Abeta (A0) scFv is linked to the C-terminus of the anti-BBB antibody heavy chain with a Fc knob mutation.

[0404] In some aspects, the antibody is a monoclonal antibody.

[0405] In some aspects, the antibody is a humanized antibody.

[0406] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is selected from the group consisting of a VHH, a Fab, a Fab’, a Fab’-SH, a F(ab’)2, a Fv, or a scFv.

[0407] In some aspects, the anti-BBB antigen-binding domain is a VHH, a Fab, a Fab’, a Fab’-SH, a F(ab’)2, a Fv, or a scFv.

[0408] In some aspects, the multispecific protein further comprises iii) an Fc region.

[0409] In some aspects, the Fc region is fully active. In some aspects, a multispecific protein of the present disclosure has a fully active Fc region, and thus, enables recruitment of myeloid cells in the brain (e.g., microglial cells) to aid in the effective and efficient removal of Abeta (A0) plaques.

[0410] In some aspects, the Fc region is partially active.

[0411] In some aspects, the Fc region is not silent.

[0412] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a VHH.

[0413] In some aspects, the anti-BBB antigen-binding domain is a VHH.

[0414] In some aspects, the multispecific protein comprises an IgGFc region, the anti-BBB antigen-binding domain is a VHH that is N-terminal to one protein fragment of the Fc region, and the anti pyroGlu3 Abeta (A0) antigen-binding domain is a VHH that is N-terminal to the other protein fragment of the Fc region.

[0415] In some aspects, the anti pyroGlu3 Abeta (A0) antigen-binding domain is a Fab.

[0416] In some aspects, the anti-BBB antigen-binding domain is a Fab.

[0417] In some aspects, the multispecific protein comprises an IgGFc region, the anti-BBB antigen-binding domain is a Fab that is N-terminal to one protein fragment of the Fc region, and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a Fab that is N-terminal to the other protein fragment of the Fc region.

[0418] In some aspects, the anti-BBB antigen-binding domain is a scFv.

[0419] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a scFv.

[0420] In some aspects, the multispecific protein comprises an IgGFc region, the anti-BBB antigen-binding domain is a scFv that is N-terminal to one protein fragment of the Fc region, and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a scFv that is N-terminal to the other protein fragment of the Fc region.

[0421] In some aspects, the multispecific protein comprises an IgGFc region, the anti-BBB antigen-binding domain is a scFv that is N-terminal to one protein fragment of the Fc region, and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a Fab that is N-terminal to the other protein fragment of the Fc region.

[0422] In some aspects, the multispecific protein comprises an IgGFc region, the anti-BBB antigen-binding domain is a Fab that is N-terminal to one protein fragment of the Fc region, and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is a Fab that is N-terminal to the other protein fragment of the Fc region

[0423] In some aspects, the IgG Fc region is an IgGl Fc region or an IgG4 Fc region.

[0424] In some aspects, the IgG Fc region comprises knob and hole mutations.

[0425] In some aspects, the multispecific protein is bivalent.

[0426] In some aspects, the multispecific protein is trivalent.

[0427] In some aspects, the trivalent protein comprises one antigen-binding domain that binds to the BBB target and two antigen-binding domains that bind to pyroGlu3 Abeta (A0).

[0428] In some aspects, the trivalent protein comprises two antigen-binding domains that bind to the BBB target and one antigen-binding domain that binds to pyroGlu3 Abeta (A0).

[0429] In some aspects, the multispecific protein is tetravalent, optionally wherein the tetravalent protein comprises two of antigen-binding domains that bind to the BBB target and two antigen-binding domains that bind to pyroGlu3 Abeta (A0).

[0430] In some aspects, the multispecific protein is a bispecific T-cell engager (BiTE), dualaffinity re-targeting protein (DARTs), or Tandem diabody (TandAb).

[0431] In some aspects, the anti-BBB antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen-binding domain are contained within a single polypeptide chain. In some aspects, the single polypeptide chain further comprises an Fc domain.

[0432] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is N-terminal to the anti-BBB antigen-binding domain.

[0433] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain thereof is C-terminal to the anti-BBB antigen-binding domain.

[0434] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain and the anti-BBB antigen-binding domain are directly connected via a peptide bond.

[0435] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain and the anti-BBB antigen-binding domain are connected via a linker.

[0436] In some aspects, the linker is a peptide linker.

[0437] In some aspects, the anti-BBB antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen -binding domain are N-terminal to the Fc domain.

[0438] In some aspects, the anti-BBB antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen-binding domain are C-terminal to the Fc domain.

[0439] In some aspects, the anti-BBB antigen-binding domain is N-terminal to the Fc domain and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is C-terminal to the Fc domain.

[0440] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is N-terminal to the Fc domain and the anti-BBB antigen-binding domain is C-terminal to the Fc domain.

[0441] In some aspects, the multispecific protein comprises two copies of the anti-pyroGlu3 Abeta (A0) antigen-binding domain and / or two copies of the anti-BBB antigen-binding domain.

[0442] In some aspects, an anti-pyroGlu3 Abeta (A0) antigen-binding domain of the present disclosure is an antagonist to pyroGlu3 Abeta (A0) activity or pathology.

[0443] In some aspects, an anti-pyroGlu3 Abeta (A0) antigen-binding domain of the present disclosure reduces pyroGlu3 Abeta (A0) levels in the brain.

[0444] In some aspects, an anti-pyroGlu3 Abeta (A0) antigen-binding domain of the present disclosure competes with one or more antibodies selected from anti-pyroGlu3 Abeta (A0) antibody AB-1, AB-2, AB-3, AB-4, AB-5, AB-6, AB-7, AB-8, AB-9, AB-10, and any combination thereof for binding to pyroGlu3 Abeta (A0).

[0445] In some aspects, the anti- pyroGlu3 Abeta (A0) antigen-binding domain binds human pyroGlu3 Abeta (A0), binds mouse Abeta (A0), binds cynomolgus Abeta (A0), binds both human and mouse Abeta (A0), or binds human, mouse, and cynomolgus Abeta (A0).

[0446] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain binds human pyroGlu3 Abeta (A0) with an affinity of about 1 nM to about 100 nM, of about 5 nM to about 100 nMfrom about 7 nM to about 88 nM. In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain binds human pyroGlu3 Abeta (A0) with an affinity of about 10 nM to about 50 nM.

[0447] In some aspects, the antigen-binding domain is an anti-TfR antigen-binding domain.

[0448] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain in the multispecific protein is N-terminal to the anti-TfR antigen-binding domain.

[0449] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is C-terminal to the anti-TfR antigen-binding domain.

[0450] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain and the anti-TfR antigen-binding domain are directly connected via a peptide bond.

[0451] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain and the anti-TfR antigen-binding domain are connected via a linker.

[0452] In some aspects, the linker is a peptide linker.

[0453] In some aspects, the multispecific protein further comprises iii) an Fc region.

[0454] In some aspects, the Fc region is fully active. In some aspects, a multispecific protein of the present disclosure has a fully active Fc region, and thus, enables recruitment of myeloid cells in the brain (e.g., microglial cells) to aid in the effective and efficient removal of Abeta (A0) plaques.

[0455] In some aspects, the Fc region is partially active.

[0456] In some aspects, the Fc region is not silent.

[0457] In some aspects, the anti-TfR antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen -binding domain are linked to the N-terminus of the Fc portion of the multi-specific binding protein.

[0458] In some aspects, the anti-TfR antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen-binding domain are both linked to the C-terminus of the Fc portion of the multispecific binding protein.

[0459] In some aspects, the anti-TfR antigen-binding domain is linked to the N-terminus of the Fc portion and the anti- pyroGlu3 Abeta (A0) antigen-binding domain is linked to the C-terminus of the Fc portion of the multispecific protein.

[0460] In some aspects, the anti-pyroGlu3 Abeta (A0) antigen-binding domain is linked to the N-terminus of the Fc portion of the multi-specific binding protein and the anti-TfR antigenbinding domain is linked to the C-terminus of the Fc portion.

[0461] In some aspects, the anti-TfR antigen-binding domain is a single scFv, or a VHH, or a Fab.

[0462] In some aspects, the anti-TfR antigen-binding domain binds to a TfR domain with a binding affinity of about 1 nM to about 100 nM, about 1 nM to about 1000 nM, about 1 nM to about 2000 nM, or about 40 nM to about 2200 nM.

[0463] In some aspects, the multi-specific binding protein comprises two copies of the anti-pyroGlu3 Abeta (A0) antigen-binding domain.

[0464] In some aspects, the multi-specific binding protein comprises an Fc domain or Fc region.

[0465] In some aspects, the single scFv, Fab or VHH is linked to the C-terminus of the Fc domain or Fc region, wherein the two copies of the anti-pyroGlu3 Abeta (A0) antigen-binding domain are linked to the N-terminus of the Fc domain or Fc region.

[0466] In some aspects, the single scFv, Fab or VHH is linked to the N-terminus of the Fc domain or Fc region, wherein the two copies of the anti-pyroGlu3 Abeta (A0) antigen-binding domain are linked to the C-terminus of the Fc domain or Fc region.

[0467] In some aspects, the anti-TfR antigen binding domain is an anti-TfR antibody, and wherein the antibody comprises two heavy chains and two light chains.

[0468] In some aspects, two copies of an anti-pyroGlu3 Abeta (A0) antigen-binding domain are linked to the C-terminus of the two anti-TfR antibody heavy chains.

[0469] In some aspects, the anti-TfR antigen binding domain comprises two scFv, Fab, or VHH domains linked to the C-terminus of the heavy chain.

[0470] In some aspects, two copies of the anti-pyroGlu3 Abeta (A0) antigen-binding domain are linked to the N-terminus of the Fc domain or Fc region.

[0471] In some aspects, the anti-TfR antigen binding domain comprises two scFv, Fab, or VHH antigen-binding domains that specifically bind to human TfR linked to the N-terminus of the heavy chain.

[0472] In some aspects, two copies of the anti-pyroGlu3 Abeta (A0) antigen-binding domain are linked to the C-terminus of the Fc domain or Fc region.

[0473] In some aspects, the anti-TfR antigen binding domain comprises a single scFv, VHH, or Fab antigen-binding domain.

[0474] In some aspects, the multi-specific binding protein further comprises (ii) an Fc domain or Fc region, and (iii) a single copy of the anti-pyroGlu3 Abeta (A0) antigen-binding domain.

[0475] In some aspects, the single scFv, VHH, or Fab antigen-binding domain is linked to the C-terminus of the Fc domain or Fc region and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is linked to N-terminus of the Fc domain or Fc region.

[0476] In some aspects, the single scFv, VHH, or Fab antigen-binding domain is linked to the N-terminus of the Fc domain or Fc region and the anti-pyroGlu3 Abeta (A0) antigen-binding domain is linked to C-terminus of the Fc domain or Fc region.

[0477] In some aspects, the single scFv, VHH, or Fab antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen-binding domain are both linked to the N-terminus of the Fc domain or Fc region.

[0478] In some aspects, the single scFv, VHH, or Fab antigen-binding domain and the anti-pyroGlu3 Abeta (A0) antigen-binding domain are both linked to the C-terminus of the Fc domain or Fc region.

[0479] In some aspects, the Fc is a single chain, engineered monovalent Fc domain.

[0480] In some aspects, the multi-specific binding protein is a bispecific protein.

[0481] In some aspects, provided herein is a composition comprising any of the multi-specific binding proteins disclosed herein.Polynucleotides and Methods of Making the Same

[0482] In some aspects, provided herein is a nucleotide sequence encoding an antigen-binding domain that specifically binds to a human blood brain barrier protein or receptor and an antigenbinding domain that specifically binds to a human pyroGlu3 Abeta (A0) peptide. In some aspects, provided herein are polynucleotides encoding an antigen-binding domain that specifically binds to a human blood brain barrier protein or receptor and encoding an antigenbinding thereof domain that specifically binds to a human pyroGlu3 Abeta (A0) peptide. In some aspects, the human blood brain barrier protein or receptor is TfR. In some aspects, the human blood brain barrier protein or receptor is CD98hc.

[0483] Provided herein are polynucleotides comprising a nucleotide sequence encoding the compositions described herein, or a domain thereof described herein, that are optimized, e.g., by codon / RNA optimization, replacement with heterologous signal sequences, and / or elimination of mRNA instability elements. Methods to generate optimized nucleic acids for recombinant expression by introducing codon changes (e.g., a codon change that encodes the same amino acid due to the degeneracy of the genetic code) and / or eliminating inhibitory regions in the mRNAcan be carried out by adapting the optimization methods described in, e.g., U. S. Patent Nos. 5,965,726; 6,174,666; 6,291,664; 6,414,132; and 6,794,498, accordingly.

[0484] A polynucleotide comprising a nucleotide sequence encoding the compositions described herein, or a domain thereof described herein, can be generated from nucleic acid from a suitable source (e.g., a hybridoma) using methods well known in the art (e.g., PCR and other molecular cloning methods). For example, PCR amplification using synthetic primers hybridizable to the 3’ and 5’ ends of a known sequence can be performed using genomic DNA obtained from hybridoma cells producing the antibody of interest. Such PCR amplification methods can be used to obtain nucleic acids comprising, e.g., the sequence encoding the light chain and / or heavy chain of an antigen-binding domain, antibody, or antigen-binding fragment thereof. The amplified nucleic acids can be cloned into vectors for expression in host cells and for further cloning, for example, to generate an antigen-binding domain that specifically bind to human TfR, fusion protein, complex, antibody, antigen-binding fragment thereof, or multispecific protein described herein, or a domain thereof described herein.

[0485] Polynucleotides provided herein can be in the form of RNA or in the form of DNA. DNA includes cDNA, genomic DNA, and synthetic DNA, and DNA can be double-stranded or single-stranded. If single stranded, DNA can be the coding strand or non-coding (anti-sense) strand. In some aspects, the polynucleotide is a cDNA or a DNA lacking one more endogenous introns. In some aspects, a polynucleotide is a non-naturally occurring polynucleotide. In some aspects, a polynucleotide is recombinantly produced. In some aspects, the polynucleotides are isolated. In some aspects, the polynucleotides are substantially pure.

[0486] In some aspects, polynucleotides provided herein are in the form of RNA. In some aspects, polynucleotides provided herein are in the form of RNA encoding a fusion protein or complex provided herein. In some aspects, a polynucleotide provided herein is a synthetic messenger RNA (mRNA). In some aspects, the synthetic mRNA has at least one nucleoside modification. In some aspects, the at least one nucleoside modification is selected from the group consisting of pyridin-4-one ribonucleoside, 5 -aza-uridine, 2-thio-5-aza-uridine, 2-thiouridine, 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxyuridine, 3 -methyluridine, 5-carboxymethyl-uridine, 1-carboxymethyl-pseudouridine, 5-propynyl-uridine, 1-propynyl-pseudouridine, 5-taurinomethyluridine, 1-taurinomethyl-pseudouridine, 5-taurinomethyl-2-thio-uridine, 1-taurinomethyl-4-thio-uridine, 5-methyl-uridine, 1-methyl-pseudouridine, 4-thio-l -methylpseudouridine, 2-thio-l-methyl-pseudouridine, 1 -methyl- 1-deaza-pseudouri dine, 2-thio-l-methyl- 1-deaza-pseudouri dine, dihydrouridine, dihydropseudouridine, 2-thio-dihydrouridine, 2-thio-dihydropseudouridine, 2-methoxyuridine, 2-methoxy-4-thio-uridine, 4-methoxy-pseudouridine, 4-methoxy-2-thio-pseudouridine, 5-aza-cytidine, pseudoisocytidine, 3-methyl-cytidine, N4-acetylcytidine, 5-formylcytidine, N4-methylcytidine, 5-hydroxymethylcytidine, 1-methyl-pseudoisocytidine, pyrrolo-cytidine, pyrrolo-pseudoisocytidine, 2-thio-cytidine, 2-thio-5-methyl-cytidine, 4-thio-pseudoisocytidine, 4-thio-l-methy...

Claims

WHAT IS CLAIMED IS:

1. An isolated antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) wherein the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and a light chain variable region (VL) CDR1, CDR2, and CDR3 comprising the sequences of:(a) SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively;(b) SEQ ID NOs: 4, 5, 6, 21, 22, and 23, respectively;(c) SEQ ID NOs: 7, 8, 9, 24, 25, and 26, respectively;(d) SEQ ID NOs: 10, 11, 12, 27, 28, and 29, respectively;(e) SEQ ID NOs: 16, 17, 18, 32, 33, and 34, respectively;(f) SEQ ID NOs: 4, 5, 6, 21, 22, and 23, respectively;(g) SEQ ID NOs: 7, 8, 9, 24, 25, and 26, respectively;(h) SEQ ID NOs: 10, 19, 12, 27, 35, and 29, respectively;(i) SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; or(j) SEQ ID NOs: 16, 20, 18, 36, 33, and 34, respectively.

2. The antibody or antigen-binding fragment of claim 1, wherein the antibody or antigenbinding fragment comprises a VH that is at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or 100% identical to an amino acid sequence selected from any one of SEQ ID NOs: 43, 37, 39, 41, 45, 47, 49, 51, and 54.

3. The antibody or antigen-binding fragment of claim 1 or 2, wherein the antibody or antigen-binding fragment comprises a VL that is at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or 100% identical to an amino acid sequence selected from any one of SEQ ID NOs: 53, 38, 40, 42, 44, 46, 48, 50, 52, and 55.

4. The antibody or antigen-binding fragment of any one of claims 1-3, wherein the antibody or antigen -binding fragment comprises a VH and a VL, wherein the VH and VL comprise amino acid sequences at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to the amino acid sequences of:(a) SEQ ID NOs: 43 and 53, respectively;(b) SEQ ID NOs: 47 and 48, respectively;(c) SEQ ID NOs: 41 and 42, respectively;(d) SEQ ID NOs: 43 and 44, respectively;(e) SEQ ID NOs: 45 and 46, respectively;(f) SEQ ID NOs: 39 and 40, respectively;(g) SEQ ID NOs: 49 and 50, respectively;(h) SEQ ID NOs: 51 and 52, respectively;(i) SEQ ID NOs: 37 and 38, respectively; or(j) SEQ ID NOs: 54 and 55, respectively.

5. The antibody or antigen-binding fragment of any one of claims 1-4, wherein the antibody or antigen-binding fragment comprises a VH and / or a VL comprising the amino acid sequences of:(a) SEQ ID NOs: 43 and 53, respectively;(b) SEQ ID NOs: 47 and 48, respectively;(c) SEQ ID NOs: 41 and 42, respectively;(d) SEQ ID NOs: 43 and 44, respectively;(e) SEQ ID NOs: 45 and 46, respectively;(f) SEQ ID NOs: 39 and 40, respectively;(g) SEQ ID NOs: 49 and 50, respectively;(h) SEQ ID NOs: 51 and 52, respectively;(i) SEQ ID NOs: 37 and 38, respectively; or(j) SEQ ID NOs: 54 and 55, respectively.

6. An antibody or antigen-binding fragment thereof that specifically binds to human N- terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) selected from the group consisting of:(a) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 53;(b) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 47 and a VL comprising the amino acid sequence of SEQ ID NO: 48;(c) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 37 and a VL comprising the amino acid sequence of SEQ ID NO: 38;(d) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 39 and a VL comprising the amino acid sequence of SEQ ID NO: 40;(e) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 41 and a VL comprising the amino acid sequence of SEQ ID NO: 42;(f) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 44;(g) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 45 and a VL comprising the amino acid sequence of SEQ ID NO: 46;(h) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 49 and a VL comprising the amino acid sequence of SEQ ID NO: 50;(i) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 51 and a VL comprising the amino acid sequence of SEQ ID NO: 52;(j) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 54 and a VL comprising the amino acid sequence of SEQ ID NO: 55; and(k) an antibody that binds to the same human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) epitope as the antibody or antigen-binding fragment thereof of any one of (a)-(j).

7. An antibody or antigen-binding fragment thereof that specifically binds to human N- terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) selected from the group consisting of:(a) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 53;(b) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 47 and a VL comprising the amino acid sequence of SEQ ID NO: 48;(c) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 37 and a VL comprising the amino acid sequence of SEQ ID NO: 38;(d) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 39 and a VL comprising the amino acid sequence of SEQ ID NO: 40;(e) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 41 and a VL comprising the amino acid sequence of SEQ ID NO: 42;(f) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 43 and a VL comprising the amino acid sequence of SEQ ID NO: 44;(g) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 45 and a VL comprising the amino acid sequence of SEQ ID NO: 46;(h)an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 49 and a VL comprising the amino acid sequence of SEQ ID NO: 50;(i) an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 51 and a VL comprising the amino acid sequence of SEQ ID NO: 52;(j)an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 54 and a VL comprising the amino acid sequence of SEQ ID NO: 55; and(k) an antibody that competitively inhibits the binding of any one of (a)-(j) to human N- terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta).

8. The antibody or antigen-binding fragment of any one of claims 1-7, wherein the antibody or antigen -binding fragment comprises a heavy chain constant region and a light chain constant region.

9. The antibody or antigen-binding fragment of claim 8, wherein the heavy chain constant region is an isotype selected from the group consisting of human IgGi, IgG2, IgGs, and IgG4 isotypes.

10. The antibody or antigen-binding fragment of any one of claims 1-9, wherein the antibody or antigen -binding fragment comprises a heavy chain constant region and a light chain constant region, wherein the heavy chain constant region is a human IgGi heavy chainconstant region, and wherein the light chain constant region is a human IgGK light chain constant region.

11. The antibody or antigen-binding fragment of any one of claims 1-10, wherein the antibody or antigen-binding fragment is a monoclonal antibody.

12. The antibody or antigen-binding fragment of any one of claims 1-11, wherein the antibody or antigen-binding fragment thereof is a murine, chimeric, humanized, or human antibody or antigen-binding fragment thereof.

13. The antibody or antigen binding fragment of any one of claims 1-12, which is a full length antibody.

14. The antibody or antigen binding fragment of any one of claims 1-12, which is an antigen binding fragment.

15. The antigen binding fragment of claim 14, wherein the antigen binding fragment is a Fab, Fab’, F(ab’)2, single chain Fv (scFv), disulfide linked Fv, V-NAR domain, IgNar, intrabody, IgGACH2, minibody, F(ab’)s, tetrabody, triabody, diabody, single-domain antibody, DVD-Ig, Fcab, mAb2, (scFv)2, or scFv-Fc.

16. The antibody or antigen-binding fragment of any one of claims 1-15, wherein the antibody or antigen-binding fragment thereof binds at least 2-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human non-modified E3-24 Abeta polypeptide.

17. The antibody or antigen-binding fragment of any one of claims 1-16, wherein the antibody or antigen-binding fragment thereof binds about 2-fold to about 5-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human non-modified E3-24 Abeta polypeptide.

18. The antibody or antigen-binding fragment of any one of claims 1-17, wherein the antibody or antigen-binding fragment thereof binds at least 40-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human non-modified E3-40 Abeta polypeptide or wherein the antibody or antigen-binding fragment thereof binds about 40- fold to about 65-fold more strongly to human pyro-Glu3-24 Abeta polypeptide than to human non-modified E3-40 A-beta polypeptide.

19. The antibody or antigen-binding fragment of any one of claims 1-18, wherein the antibody or antigen-binding fragment promotes phagocytosis of Abeta plaques, optionally wherein 0.1 pg / ml increases phagocytosis of Abeta plaques by about 50% as compared to a control.

20. The antibody or antigen-binding fragment of any one of claims 1-19, wherein the antibody or antigen-binding fragment promotes uptake of pyroGlu3 Abeta by microglia in the CNS.

21. The antibody or antigen-binding fragment of any one of claims 1-20, wherein the antibody or antigen-binding fragment thereof is monospecific.

22. The antibody or antigen-binding fragment of any one of claims 1-20, wherein the antibody or antigen-binding fragment thereof is bispecific or multispecific.

23. The antibody or antigen-binding fragment of any one of claims 1-20, further comprising an antigen-binding domain that binds to blood brain barrier (BBB) target.

24. A complex comprising the antibody or antigen-binding fragment of any one of claims 1- 20 and an antigen-binding domain that binds to a BBB target.

25. The antibody or antigen-binding fragment or complex of any one of claims 1-24, further comprising a detectable label.

26. The antibody or antigen-binding fragment or complex of any one of claims 23-25, wherein the BBB target is human transferrin receptor (TfR) or human CD98 heavy chain (CD98hc).

27. The antibody or antigen-binding fragment or complex of claim 26, wherein(a) the BBB target is human TfR and wherein the antibody or antigen-binding fragment or complex comprises a human TfR antigen-binding domain comprising heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and light chain variable region (VL) CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of:(i) SEQ ID NOs: 56, 57, 59, 69, 67, and 68, respectively;(ii) SEQ ID NOs: 56, 57, 58, 66, 67, and 68, respectively;(iii) SEQ ID NOs: 56, 57, 58, 69, 67, and 68, respectively;(iv) SEQ ID NOs: 56, 57, 60, 69, 67, and 68, respectively;(v) SEQ ID NOs: 56, 57, 59, 70, 67, and 68, respectively;(vi) SEQ ID NOs: 56, 57, 59, 71, 67, and 68, respectively;(vii) SEQ ID NOs: 56, 57, 59, 72, 67, and 68, respectively;(viii) SEQ ID NOs: 56, 57, 59, 73, 67, and 68, respectively;(ix) SEQ ID NOs: 56, 57, 61, 69, 67, and 68, respectively;(x) SEQ ID NOs: 62, 63, 64, 74, 75, and 76, respectively;(xi) SEQ ID NOs: 62, 65, 64, 77, 75, and 76, respectively;(xii) SEQ ID NOs: 62, 63, 64, 78, 75, and 76, respectively;(xiii) SEQ ID NOs: 62, 63, 64, 79, 75, and 76, respectively;(xiv) SEQ ID NOs: 62, 65, 64, 80, 75, and 76, respectively;(xv) SEQ ID NOs: 56, 57, 59, 66, 67, and 68, respectively;(xvi) SEQ ID NOs: 62, 275, 64, 276, 75, and 277, respectively; or (xvii) SEQ ID NOs: 62, 275, 64, 77, 75, and 76, respectively; or (b) the BBB target is human CD98hc and wherein the antibody or antigen-binding fragment or complex comprises a human CD98hc antigen-binding domain comprising heavy chain variable region (VH) complementarity determining region (CDR) 1, VH CDR2, VH CDR3 and light chain variable region (VL) CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of:(i) SEQ ID NOs: 98, 99, 100, 122, 123, and 124, respectively; (ii) SEQ ID NOs: 98, 101, 100, 122, 123, and 124, respectively; (iii) SEQ ID NOs: 98, 102, 100, 122, 123, and 124, respectively; (iv) SEQ ID NOs: 98, 103, 100, 122, 123, and 124, respectively; (v) SEQ ID NOs: 98, 104, 100, 122, 123, and 124, respectively; (vi) SEQ ID NOs: 98, 105, 100, 122, 123, and 124, respectively; (vii) SEQ ID NOs: 98, 106, 100, 122, 123, and 124, respectively; (viii) SEQ ID NOs: 98, 107, 100, 122, 123, and 124, respectively; (ix) SEQ ID NOs: 98, 108, 100, 122, 123, and 124, respectively; (x) SEQ ID NOs: 98, 109, 100, 122, 123, and 124, respectively; (xi) SEQ ID NOs: 98, 110, 100, 122, 123, and 124, respectively; (xii) SEQ ID NOs: 98, 99, 100, 125, 123, and 124, respectively; (xiii) SEQ ID NOs: 98, 99, 100, 126, 123, and 124, respectively; (xiv) SEQ ID NOs: 98, 99, 100, 127, 123, and 124, respectively; (xv) SEQ ID NOs: 98, 99, 100, 128, 123, and 124, respectively;(xvi) SEQ ID NOs: 98, 99, 100, 129, 123, and 124, respectively;(xvii) SEQ ID NOs: 98, 99, 100, 130, 123, and 124, respectively;(xviii) SEQ ID NOs: 98, 99, 100, 131, 123, and 124, respectively;(xix) SEQ ID NOs: 98, 106, 100, 127, 123, and 124, respectively;(xx) SEQ ID NOs: 98, 111, 100, 122, 123, and 124, respectively;(xxi) SEQ ID NOs: 98, 99, 112, 122, 123, and 124, respectively;(xxii) SEQ ID NOs: 98, 99, 113, 122, 123, and 124, respectively;(xxiii) SEQ ID NOs: 98, 99, 114, 122, 123, and 124, respectively;(xxiv) SEQ ID NOs: 98, 99, 115, 122, 123, and 124, respectively;(xxv) SEQ ID NOs: 98, 99, 116, 122, 123, and 124, respectively;(xxvi) SEQ ID NOs: 98, 99, 117, 122, 123, and 124, respectively;(xxvii) SEQ ID NOs: 98, 99, 118, 122, 123, and 124, respectively;(xxviii)SEQ ID NOs: 98, 99, 119, 122, 123, and 124, respectively;(xxix) SEQ ID NOs: 98, 99, 120, 122, 123, and 124, respectively;(xxx) SEQ ID NOs: 98, 99, 100, 132, 123, and 124, respectively;(xxxi) SEQ ID NOs: 98, 99, 100, 133, 123, and 124, respectively;(xxxii) SEQ ID NOs: 98, 99, 100, 134, 123, and 124, respectively;(xxxiii)SEQ ID NOs: 98, 99, 100, 135, 123, and 124, respectively;(xxxiv)SEQ ID NOs: 98, 99, 100, 136, 123, and 124, respectively;(xxxv) SEQ ID NOs: 98, 99, 100, 137, 123, and 124, respectively;(xxxvi)SEQ ID NOs: 98, 99, 100, 122, 123, and 138, respectively;(xxxvii) SEQ ID NOs: 98, 99, 100, 122, 123, and 139, respectively; (xxxviii) SEQ ID NOs: 98, 99, 100, 122, 123, and 140, respectively; (xxxix)SEQ ID NOs: 98, 99, 100, 122, 123, and 141, respectively;(xl) SEQ ID NOs: 98, 99, 100, 122, 123, and 142, respectively;(xli) SEQ ID NOs: 98, 99, 100, 122, 123, and 143, respectively;(xlii) SEQ ID NOs: 98, 99, 100, 122, 123, and 144, respectively;(xliii) SEQ ID NOs: 98, 99, 100, 122, 123, and 145, respectively;(xliv) SEQ ID NOs: 98, 99, 100, 122, 123, and 146, respectively;(xlv) SEQ ID NOs: 98, 99, 100, 122, 123, and 147, respectively;(xlvi) SEQ ID NOs: 121, 99, 100, 122, 123, and 124, respectively; or (xlvii) SEQ ID NOs: 121, 99, 100, 122, 123, and 147, respectively.

28. The antibody or antigen-binding fragment or complex of claim 26 or 27, wherein(a) the BBB target is human TfR and wherein the human TfR antigen-binding domain comprises a VH and a VL, wherein the VH and VL comprise amino acid sequences at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to the amino acid sequences of:(i) SEQ ID NOs: 84 and 83, respectively;(ii) SEQ ID NOs: 81 and 82, respectively;(iii) SEQ ID NOs: 81 and 83, respectively;(iv) SEQ ID NOs: 85 and 83, respectively;(v) SEQ ID NOs: 84 and 86, respectively;(vi) SEQ ID NOs: 84 and 87, respectively;(vii) SEQ ID NOs: 84 and 88, respectively;(viii) SEQ ID NOs: 84 and 89, respectively;(ix) SEQ ID NOs: 90 and 83, respectively;(x) SEQ ID NOs: 91 and 92, respectively;(xi) SEQ ID NOs: 93 and 94, respectively;(xii) SEQ ID NOs: 91 and 95, respectively;(xiii) SEQ ID NOs: 91 and 96, respectively;(xiv) SEQ ID NOs: 93 and 97, respectively;(xv) SEQ ID NOs: 269 and 270, respectively;(xvi) SEQ ID NOs: 271 and 272, respectively; or(xvii) SEQ ID NOs: 273 and 274, respectively; or(b) the BBB target is human CD98hc and wherein the human CD98hc antigen-binding domain comprises a VH and a VL, wherein the VH and VL comprise amino acid sequences at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to the amino acid sequences of:(i) SEQ ID NOs: 148 and 149, respectively;(ii) SEQ ID NOs: 150 and 149, respectively;(iii) SEQ ID NOs: 151 and 149, respectively;(iv) SEQ ID NOs: 152 and 149, respectively;(v) SEQ ID NOs: 153 and 149, respectively;(vi) SEQ ID NOs: 154 and 149, respectively;(vii) SEQ ID NOs: 155 and 149, respectively;(viii) SEQ ID NOs: 156 and 149, respectively;(ix) SEQ ID NOs: 157 and 149, respectively;(x) SEQ ID NOs: 158 and 149, respectively;(xi) SEQ ID NOs: 159 and 149, respectively;(xii) SEQ ID NOs: 148 and 160, respectively;(xiii) SEQ ID NOs: 148 and 161, respectively;(xiv) SEQ ID NOs: 148 and 162, respectively;(xv) SEQ ID NOs: 148 and 163, respectively;(xvi) SEQ ID NOs: 148 and 164, respectively;(xvii) SEQ ID NOs: 148 and 165, respectively;(xviii) SEQ ID NOs: 148 and 166, respectively;(xix) SEQ ID NOs: 155 and 162, respectively;(xx) SEQ ID NOs: 167 and 149, respectively;(xxi) SEQ ID NOs: 168 and 149, respectively;(xxii) SEQ ID NOs: 169 and 149, respectively;(xxiii) SEQ ID NOs: 170 and 149, respectively;(xxiv) SEQ ID NOs: 171 and 149, respectively;(xxv) SEQ ID NOs: 172 and 149, respectively;(xxvi) SEQ ID NOs: 173 and 149, respectively;(xxvii) SEQ ID NOs: 174 and 149, respectively;(xxviii)SEQ ID NOs: 175 and 149, respectively;(xxix) SEQ ID NOs: 176 and 149, respectively;(xxx) SEQ ID NOs: 148 and 177, respectively;(xxxi) SEQ ID NOs: 148 and 178, respectively;(xxxii) SEQ ID NOs: 148 and 179, respectively;(xxxiii)SEQ ID NOs: 148 and 180, respectively;(xxxiv)SEQ ID NOs: 148 and 181, respectively;(xxxv) SEQ ID NOs: 148 and 182, respectively;(xxxvi)SEQ ID NOs: 148 and 183, respectively;(xxxvii) SEQ ID NOs: 148 and 184, respectively; (xxxviii) SEQ ID NOs: 148 and 185, respectively; (xxxix)SEQ ID NOs: 148 and 186, respectively;(xl) SEQ ID NOs: 148 and 187, respectively;(xli) SEQ ID NOs: 148 and 188, respectively;(xlii) SEQ ID NOs: 148 and 189, respectively;(xliii) SEQ ID NOs: 148 and 190, respectively;(xliv) SEQ ID NOs: 148 and 191, respectively;(xlv) SEQ ID NOs: 148 and 192, respectively;(xlvi) SEQ ID NOs: 193 and 194, respectively; or(xlvii) SEQ ID NOs: 193 and 192, respectively.

29. The antibody or antigen-binding fragment or complex of any one of claims 23-28, wherein the human TfR antigen-binding domain comprises a disulfide staple.

30. The antibody or antigen-binding fragment or complex of any one of claims 23-29, wherein the human TfR antigen-binding domain comprises the amino acid sequence of SEQ ID NO:202 or SEQ ID NO:203.

31. The antibody or antigen-binding fragment or complex of claim 27, wherein the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises the VH CDR 1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; and the human TfR antigen-binding domain comprises VH CDR1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs: 56, 57, 59, 69, 67, and 68, respectively.

32. The antibody or antigen-binding fragment or complex of claim 27, wherein the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises the VH CDR 1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 13, 14, 15, 24, 30, and 31, respectively; and the human TfR antigen-binding domain comprises VH CDR1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs: 56, 57, 59, 72, 67, and 68, respectively.

33. The antibody or antigen-binding fragment or complex of claim 27, wherein the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises the VH CDR 1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences of: SEQ ID NOs: 13, 14,15, 24, 30, and 31, respectively; and the human TfR antigen-binding domain comprises VH CDR1, VH CDR2, VH CDR3 and VL CDR1, CDR2, and CDR3 sequences comprising the amino acid sequences of SEQ ID NOs: 56, 57, 59, 73, 67, and 68, respectively.

34. The antibody or antigen-binding fragment or complex of claim 28, wherein the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises a VH and a VL comprising the amino acid sequences of SEQ ID NO:43 and 53 respectively; and the human TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs: 84 and 83, respectively.

35. The antibody or antigen-binding fragment or complex of claim 28, wherein the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises a VH and a VL comprising the amino acid sequences of SEQ ID NO:43 and 53 respectively; and the human TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs: 84 and 88, respectively.

36. The antibody or antigen-binding fragment or complex of claim 28, wherein the antibody or antigen-binding fragment thereof that specifically binds to human N-terminal truncated pyroglutamate amyloid beta peptide (pyroGlu3 Abeta) comprises a VH and a VL comprising the amino acid sequences of SEQ ID NO:47 and 48 respectively; and the human TfR antigen-binding domain comprises a VH and a VL comprising the amino acid sequences of SEQ ID NOs: 84 and 89, respectively.

37. The antibody or antigen-binding fragment or complex of any one of claims 23-36, wherein (i) the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta is a full-length antibody, a single-chain fragment variable (scFv), a VHH, or a Fab, and (ii) the anti-BBB antigen-binding domain is a single-chain fragment variable (scFv).

38. The antibody or antigen-binding fragment or complex of any one of claims 23-36, wherein (i) the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta is a full-length antibody, a single-chain fragment variable (scFv), a VHH, or a Fab and (ii) the anti-BBB antigen-binding domain is a VHH.

39. The antibody or antigen-binding fragment or complex of any one of claims 23-36, wherein (i) the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta is a full-length antibody, a single-chain fragment variable (scFv), a VHH, or a Fab and (ii) the anti-BBB antigen-binding domain is a Fab.

40. The antibody or antigen-binding fragment or complex of any one of claims 23-39, wherein the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the anti-BBB antigen-binding domain are connected by a linker.

41. The antibody or antigen-binding fragment or complex of claim 40, wherein the linker connects the C-terminus of the heavy chain Fc of the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the N-terminus of the anti-BBB antigenbinding domain.

42. The antibody or antigen-binding fragment or complex of claim 40 or 41, wherein the linker comprises GGSGG (SEQ ID NO: 214).

43. The antibody or antigen-binding fragment or complex of any one of claims 23-39, wherein the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the anti-BBB antigen-binding domain are directly connected.

44. The antibody or antigen-binding fragment or complex of any one of claims 23-43, wherein the antibody or antigen-binding fragment that binds to pyroGlu3 Abeta and the anti-BBB antigen-binding domain comprises the amino acid sequence of SEQ ID NO:280, SEQ ID NO:281, or SEQ ID NO:282.

45. The antibody or antigen-binding fragment or complex of any one of claims 23-44, wherein the antibody or antigen-binding fragment or complex is bivalent.

46. The antibody or antigen-binding fragment or complex of any one of claims 23-44, wherein the antibody or antigen-binding fragment or complex is trivalent, optionally wherein the antibody or antigen-binding fragment or complex comprises (i) one antigenbinding domain that binds to the BBB target and two antigen-binding domains that bind to pyroGlu3 Abeta or (ii) two antigen-binding domains that bind to the BBB target and one antigen-binding domain that binds to pyroGlu3 Abeta.

47. The antibody or antigen-binding fragment or complex of any one of claims 23-44, wherein the antibody or antigen-binding fragment or complex is tetravalent, optionallywherein the antibody or antigen-binding fragment or complex comprises two antigenbinding domains that bind to the BBB target and two antigen-binding domains that bind to pyroGlu3 Abeta.

48. The antibody or antigen-binding fragment or complex of any one of claims 23-28, comprising the amino acid sequences of (a) SEQ ID NOs: 252, 230, and 231 (b) SEQ ID NOs: 223, 224, and 225, (c) SEQ ID NOs: 229, 230, and 231; (d) SEQ ID NOs: 241, 224, and 225; (e) SEQ ID NOs: 242, 224, and 225; (f) SEQ ID NOs: 243, 224, and 225; (g) SEQ ID NOs: 244, 224, and 225; (h) SEQ ID NOs 245, 224, and 225; (i) SEQ ID NOs: 246, 224, and 225; (j) SEQ ID NOs: 247, 224, and 225; (k) SEQ ID NOs: 248, 224, and 225; (1) SEQ ID NOs: 249, 224, and 225; (m) SEQ ID NOs: 250, 224, and 225; (n) SEQ ID NOs: 251, 230, and 231; (o) SEQ ID NOs: 226, 227, and 228(p) SEQ ID NOs: 253, 230, and 231; (q) SEQ ID NOs: 254, 230, and 231; (r) SEQ ID NOs: 255, 230, and 231; (s) SEQ ID NOs: 256, 230, and 231; (t) SEQ ID NOs: 257, 230, and 231; (u) SEQ ID NOs: 258, 230, and 231; (v) SEQ ID NOs: 259, 230, and 231; (w) SEQ ID NOs: 260, 230, and 231; (x) SEQ ID NOs: 261, 230, and 231; (y) SEQ ID NOs: 262, 230, and 231; (z) SEQ ID NOs: 263, 230, and 231; (aa) SEQ ID NOs: 264, 230, and 231; (bb) SEQ ID NOs: 265, 230, and 231; SEQ ID NOs: 266, 230, and 231; (cc) SEQ ID NOs: 278, 224, and 225; or (dd) SEQ ID NOs: 279, 230, and 231.

49. The antibody or antigen-binding fragment or complex of any one of claims 23-48, wherein the antibody or antigen-binding fragment or complex enhances brain uptake of the antibody or antigen-binding fragment thereof that specifically binds to human N- terminal truncated pyroGlu3 Abeta in one or more of the frontal cortex, hippocampus, and thalamus of an animal.

50. The antibody or antigen-binding fragment or complex of any one of claims 23-48, wherein the antibody or antigen-binding fragment or complex increases the concentration of the antibody or antigen-binding fragment thereof that specifically binds to human N- terminal truncated pyroGlu3 Abeta in the CSF of an animal.

51. An isolated polynucleotide comprising a nucleic acid molecule encoding the heavy chain variable region or heavy chain of the antibody or antigen-binding fragment thereof of any one of claims 1-23.

52. The isolated polynucleotide of claim 51, wherein the nucleic acid molecule encodes the VH of any one of SEQ ID NOs: 43, 37, 39, 41, 45, 47, 49, 51, and 54.

53. An isolated polynucleotide comprising a nucleic acid molecule encoding the light chain variable region or light chain of the antibody or antigen-binding fragment thereof of any one of claims 1-23.

54. The isolated polynucleotide of claim 53, wherein the nucleic acid molecule encodes the VL of any one of SEQ ID NOs: 53, 38, 40, 42, 44, 46, 48, 50, 52, and 55.

55. An isolated polynucleotide comprising a nucleic acid molecule encoding the heavy chain variable region or heavy chain of the antibody or antigen-binding fragment thereof of any one of claims 1-23 and the light chain variable region or light chain of the antibody or antigen-binding fragment thereof of any one of claims 1-23.

56. An isolated vector comprising the polynucleotide of any one of claims 51-55.

57. A host cell comprising (a) the polynucleotide of any one of claims 51-55 (b) the vector of claim 54, or (c) a first vector comprising the polynucleotide of claim 51 or 52 and a second vector comprising the polynucleotide of claim 53 or 54.

58. The host cell of claim 57, which is selected from the group consisting of E. coli, Pseudomonas, Bacillus, Streptomyces, yeast, CHO, YB / 20, NSO, PER-C6, HEK-293T, NIH-3T3, HeLa, BHK, Hep G2, SP2 / 0, Rl.l, B-W, L-M, COS 1, COS 7, BSC1, BSC40, BMT10 cell, plant cell, insect cell, and human cell in tissue culture, optionally wherein the host cell is a HEK-293 cell.

59. A method of producing an antibody or antigen-binding fragment thereof that binds to human pyroGlu3 Abeta comprising culturing the host cell of claim 57 or 58 so that the nucleic acid molecule is expressed and the antibody or antigen-binding fragment thereof is produced, optionally wherein the method further comprises isolating the antibody or antigen-binding fragment thereof from the culture.

60. An isolated antibody or antigen-binding fragment thereof that specifically binds to human pyroGlu3 Abeta and is encoded by the polynucleotide of any one of claims 51-55, encoded by the vector of claim 56, or produced by the method of claim 59.

61. A pharmaceutical composition comprising the antibody or antigen-binding fragment thereof or complex of any one of claims 1-50 and 60 and a pharmaceutically acceptable excipient.

62. A method of promoting phagocytosis of Abeta plaques comprising contacting a composition comprising Abeta plaques and macrophages and / or microglia with the antibody or antigen binding fragment thereof or complex of any one of claims 1-50 and 60 or the pharmaceutical composition of claim 61.

63. The method of claim 62, wherein the contacting is in vitro.

64. The method of claim 62, wherein the contact is in a subject.

65. A method of slowing the progression of neurodegeneration in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex of any one of claims 1-50 and 60 or the pharmaceutical composition of claim 61.

66. A method of decreasing cognitive decline in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex of any one of 1-50 and 60 or the pharmaceutical composition of claim 61.

67. The method of any one of claims 64-66, wherein the subject has Alzheimer’s disease.

68. A method of treating a neurodegenerative disease in a subject, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex of any one of 1-50 and 60 or the pharmaceutical composition of claim 61.

69. A method of preventing a neurodegenerative disease in a subject, the method comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex of any one of 1-50 and 60 or the pharmaceutical composition of claim 61.

70. The method of claim 68 or 69, wherein the neurodegenerative disease is Alzheimer’s disease.

71. The method of claim 70, wherein the Alzheimer’s disease is familial Alzheimer’s disease.n72. The method of claim 68 or 69, wherein the neurodegenerative disease is early Alzheimer’s disease or mild cognitive impairment (MCI).

73. The method of any one of claims 65-72, comprising administering to the subject a therapeutically effective amount of the antibody or antigen binding fragment thereof or complex of any one of claims 23-50 and 60 or the pharmaceutical composition of claim 61, wherein the BBB target is TfR.

74. The method of claim 73, wherein the administration reduces the likelihood of anemia.

75. The method of claim 73 or 75, wherein the administration results in minimal hematological adverse events.

76. The method of any one of claims 73-75, wherein the administration results in no adverse or long-term reduction in reticulocyte count in the subject.

77. The method of any one of claims 73-76, wherein the administration results in no adverse or long-term reduction in red blood cell count in the subject.

78. The method of any one of claims 73-77, wherein the administration results in no adverse or long-term reduction in hematocrit in the subject.

79. The method of any one of claims 73-78, wherein the administration results in no adverse or long-term reduction in hemoglobin level in the subject.

80. The method of any one of claims 73-79, wherein the administration results in rapid clearance or reduction of Abeta in the brain of the subject.

81. The method of any one of claims 73-80, wherein the administration requires less frequent monitoring of Abeta levels or no monitoring of Abeta levels in the brain of the subject as compared to the monitoring required of other anti-Abeta antibody therapies, optionally wherein the monitoring is MRI monitoring.

82. The method of any one of claims 73-81, wherein the subject has not been APOE genotyped prior to the administration.

83. The method of any one of claims 73-82, wherein the administration is associated with less ARIA as compared to the ARIA associated with other anti-Abeta antibody therapies.

84. The method of any one of claims 65-83, wherein the administration is intravenous or subcutaneous.

85. The method of any one of claims 65-84, further comprising administering an additional active agent, optinally wherein the additional active agent is one or more of a tau- targeting agent, an ApoER2 -targeting agent, an ApoE4-targeting agent, an ApoE2- targeting agent, a NLRP3 -targeting agent, a GSK3B targeting agent, and a GLP-1 targeting agent.

86. The antibody or antigen-binding fragment thereof or complex of any one of claims 1-50 and 60 or the pharmaceutical composition of claim 61 for use in the method of claim 62 or 63 or for use in the manufacture of a medicament for use in the method of any one of claims 64-85.

87. Use of the antibody or antigen-binding fragment thereof or complex of any one of claims 1-50 and 60 or the pharmaceutical composition of claim 61 in the method of any one of claims 62-85.

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