Anti-CD371 antibodies and uses thereof

Abstract

The presently disclosed subject matter provides antibodies or antigen-binding fragments thereof that bind to CD371, and methods of using such antibodies or antigen-binding fragments thereof. In certain embodiments, the anti-CD371 antibodies or antigen-binding fragments thereof comprise a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:9, or SEQ ID NO:11.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 62 / 900,118, filed September 13, 2019, and U.S. Provisional Application No. 62 / 936,913, filed November 18, 2019, the contents of each of which are incorporated herein by reference in their entirety and to which priority is claimed. Sequence Listing

[0002] This application contains a Sequence Listing that has been submitted in ASCII format via EFS-Web and is incorporated herein by reference in its entirety. The ASCII copy was created on September 11, 2020, is designated 0727341146_ST25, and is 58,682 bytes in size.

[0003] 1. Field of the Invention The subject matter of this disclosure relates to antibodies that bind to CD371 and methods of using such antibodies. [Background technology]

[0004] 2. Background of the invention CD371 (CEC12A), also known as DCAL-2, MICL, or CLL-1, is a 30 kD C-type lectin transmembrane glycoprotein. It is expressed on monocytes, granulocytes, natural killer (NK) cells, and basophils. CD371 is an immune inhibitory receptor that recruits the Src homology phosphatases SHP-1 and SHP-2 to its phosphorylated cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM) (Sancho et al., Annu Rev. Immunol (2012); 30:491-529; Yan et al., Front Immunol (2015); 6:408; Lahoud et al., J Immunol (2011); 187:842). CD371 has been suggested to be a negative regulatory urate crystal (monosodium urate, MSU) receptor that controls autoimmune and inflammatory diseases (Neumann et al., Immunity (2014);40:389-99). CD371 is a negative regulator of granulocyte and monocyte function (Marshall et al., J Biol Chem (2004);279(15):14792-802; Pye et al., Eur J Immunol (2008);38(4):1157-63). Abnormal expression of CD371 has been reported in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) (Sadonik et al., Blood (2016);128:4234; Toft-Petersen et al., Br J Haematol (2016);175(3):393-41). Recent studies have shown that CD371 is expressed in 92% of acute myeloid leukemias (AML) and is absent from granulocyte-macrophage progenitor cells (GMPs) (Bakker et al., Cancer Res. (2004);64(22):8443-50).CD371 is also expressed in leukemia stem cells (LSCs), which have the ability to self-renew indefinitely and generate many daughter blast cells with a specific CD371 phenotype, and serves as one of the most important reasons for leukemia relapse (Siveen et al., Mol Cancer (2017); 16:13; Yoshida et al., Cancer Sci (2016); 107:5-11). Considering the important role of CD371 in disease, antibodies that bind to CD371 and methods using such agents are desirable. [Prior art documents] [Non-patent literature]

[0005] [Non-Patent Document 1] Sancho et al., Annu Rev. Immunol (2012); 30:491-529 [Non-patent document 2] Yan et al., Front Immunol (2015);6:408 [Non-patent document 3] Lahoud et al., J Immunol (2011);187:842 [Non-patent document 4] Marshall et al., J Biol Chem (2004);279(15):14792-802 [Non-Patent Document 5] Pye et al., Eur J Immunol (2008);38(4):1157-63 [Non-patent document 6] Sadonik et al., Blood (2016);128:4234 [Non-Patent Document 7] Toft-Petersen et al., Br J Haematol (2016);175(3):393-41 [Non-patent document 8] Bakker et al., Cancer Res. (2004);64(22):8443-50 [Non-Patent Document 9] Siveen et al., Mol Cancer (2017);16:13 [Non-Patent Document 10] Yoshida et al., Cancer Sci (2016);107:5-11 Summary of the Invention [Means for solving the problem]

[0006] 3. Summary of the Invention The presently disclosed subject matter provides antibodies or antigen-binding fragments thereof that specifically bind to CD371, and methods of using the antibodies or antigen-binding fragments thereof. In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11.

[0007] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, or SEQ ID NO:12.

[0008] In certain embodiments, an anti-CD371 antibody or antigen-binding fragment thereof comprises: (a) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11; and (b) a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, or SEQ ID NO:12.

[0009] In certain embodiments, the heavy and light chain variable regions of an anti-CD371 antibody or antigen-binding fragment thereof comprise: (a) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2; (b) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:3, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:4; (c) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:5, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:6; (d) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:7, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:8; (e) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:10; and (f) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:11, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:12; is selected from the group consisting of:

[0010] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, or SEQ ID NO: 11. In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, or SEQ ID NO: 12.

[0011] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11; and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, or SEQ ID NO:12.

[0012] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region and the light chain variable region are: (a) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 1, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 2; (b) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 3, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 4; (c) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 5, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 6; (d) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 8; (e) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 9, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 10; and (f) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 11, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 12 is selected from the group consisting of:

[0013] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising CDR1, CDR2, and CDR3 domains; and a light chain variable region comprising CDR1, CDR2, and CDR3 domains, wherein the heavy chain variable region CDR3 domain and the light chain variable region CDR3 domain are: (a) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33 and conservative modifications thereof; (b) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 39 and conservative modifications thereof; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45 and conservative modifications thereof; (d) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 51 and conservative modifications thereof; (e) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57 and conservative modifications thereof; and (f) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 and conservative modifications thereof. is selected from the group consisting of:

[0014] In certain embodiments, the heavy chain variable region CDR2 and the light chain variable region CDR2 of the antibody or antigen-binding portion thereof are: (a) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 29 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 32 and conservative modifications thereof; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38 and conservative modifications thereof; (c) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44 and conservative modifications thereof; (d) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 50 and conservative modifications thereof; (e) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56 and conservative modifications thereof; and (f) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62 and conservative modifications thereof. is selected from the group consisting of:

[0015] In certain embodiments, the anti-CD371 heavy chain variable region CDR1 domain and the anti-CD371 light chain variable region CDR1 domain of the antibody or antigen-binding portion thereof are (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31 and conservative modifications thereof; (b) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37 and conservative modifications thereof; (c) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43 and conservative modifications thereof; (d) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49 and conservative modifications thereof; (e) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55 and conservative modifications thereof; and (f) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61 and conservative modifications thereof. is selected from the group consisting of:

[0016] In certain embodiments, one or more of the CDR sequences have up to about 5 amino acid substitutions. In certain embodiments, one or more of the CDR sequences have up to about 3 amino acid substitutions.

[0017] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 29; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30; (b) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36; (c) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42; (d) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48; (e) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54; or (f) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60; Includes.

[0018] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof (a) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 32; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33; (b) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 39; (c) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 50; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 51; (e) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57; or (f) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 Includes.

[0019] In certain embodiments, the anti-CD371 antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 29; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 32; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33; (b) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 39; (c) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45; (d) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 50; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 51; (e) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57; or (f) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 Includes.

[0020] The presently disclosed subject matter provides antibodies or antigen-binding fragments thereof that cross-compete with any of the above antibodies or antigen-binding fragments thereof for binding to CD371.

[0021] The presently disclosed subject matter provides antibodies or antigen-binding fragments thereof that bind to the same epitope on CD371 as any of the above-described antibodies or antigen-binding fragments thereof.

[0022] In certain embodiments, the antibody sequence is in the light-heavy variable chain orientation (V L -V H In certain embodiments, the antibody or antigen-binding fragment thereof is present at about 1 x 10 -7 M to approximately 1 x 10 -8 Between m or approximately 1 x 10 -9 M to approximately 1 x 10 -8 The dissociation constant (K D ) and binds to human CD371.

[0023] In certain embodiments, the antibody or antigen-binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, or SEQ ID NO:21.

[0024] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a framework region of a human variable region. In certain embodiments, the antibody or antigen-binding fragment thereof is fully human or an antigen-binding fragment thereof. In certain embodiments, the antibody or antigen-binding fragment thereof is a chimeric antibody or antigen-binding fragment thereof. In certain embodiments, the antibody or antigen-binding fragment thereof is a humanized antibody or antigen-binding fragment thereof. In certain embodiments, the antigen-binding fragment of the antibody is a Fab, Fab', F(ab')2, variable fragment (Fv), or single-chain variable fragment (scFv).

[0025] The presently disclosed subject matter also provides immunoconjugates comprising an antibody or antigen-binding fragment thereof disclosed herein linked to a therapeutic agent. In certain embodiments, the therapeutic agent is a drug, a cytotoxin, or a radioisotope.

[0026] Additionally, the presently disclosed subject matter also provides bispecific molecules comprising an antibody or antigen-binding fragment thereof disclosed herein linked to a second functional moiety, in certain embodiments, the second functional moiety has a different binding specificity than the antibody or antigen-binding fragment thereof.

[0027] The presently disclosed subject matter also provides a composition comprising an antibody or antigen-binding fragment thereof disclosed herein, an immunoconjugate disclosed herein, or a bispecific antibody disclosed herein. In certain embodiments, the composition is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

[0028] Additionally, the presently disclosed subject matter provides nucleic acids encoding the antibodies or antigen-binding fragments thereof disclosed herein, expression vectors comprising such nucleic acid molecules, and host cells comprising such expression vectors.

[0029] The subject matter of the present disclosure is a whole cell or organizationIn certain embodiments, the method comprises contacting a cell or tissue with an antibody or antigen-binding fragment thereof disclosed herein, wherein the antibody or antigen-binding fragment thereof comprises a detectable label; and determining the amount of the labeled antibody or antigen-binding fragment thereof bound to the cell or tissue by measuring the amount of detectable label associated with the cell or tissue, wherein the amount of bound antibody or antigen-binding fragment thereof indicates the amount of CD371 in the cell or tissue.

[0030] Furthermore, the presently disclosed subject matter provides a method for treating tumor burden in a subject. In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof, immunoconjugate thereof, bispecific molecule thereof, or composition disclosed herein. In certain embodiments, the method reduces the number of tumor cells. In certain embodiments, the method reduces tumor size. In certain embodiments, the method eradicates the tumor in the subject. In certain embodiments, the subject is a human.

[0031] Furthermore, the presently disclosed subject matter provides a method for treating and / or preventing tumors or neoplasms in a subject. In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof, an immunoconjugate thereof, a bispecific molecule thereof, or a composition thereof disclosed herein. In certain embodiments, the method eradicates the tumor in the subject. In certain embodiments, the subject is a human.

[0032] In addition, the presently disclosed subject matter provides a method for increasing or extending the survival time of a subject with a tumor or neoplasm. In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof, an immunoconjugate thereof, a bispecific molecule thereof, or a composition thereof disclosed herein. In certain embodiments, the method eradicates the tumor in the subject. In certain embodiments, the subject is a human.

[0033] In addition, the presently disclosed subject matter provides a method for preventing and / or treating a tumor or neoplasm. In certain embodiments, the method comprises administering to a subject an antibody or antigen-binding fragment thereof, an immunoconjugate thereof, a bispecific molecule thereof, or a composition disclosed herein. In certain embodiments, the subject is a human.

[0034] In certain embodiments, the tumor or neoplasm is selected from the group consisting of acute myeloid leukemia (AML), multiple myeloma, non-Hodgkin's lymphoma, Hodgkin's lymphoma, chronic lymphocytic leukemia (CLL), glioblastoma, myelodysplastic syndrome (MDS), and chronic myeloid leukemia (CML). In certain embodiments, the tumor is AML.

[0035] Furthermore, the presently disclosed subject matter provides kits for treating tumor burden in a subject, for treating and / or preventing a tumor or neoplasm, and / or for increasing or prolonging survival of a subject having a tumor or neoplasm, comprising an antibody or antigen-binding fragment thereof, immunoconjugate thereof, bispecific molecule thereof, or composition thereof disclosed herein. In certain embodiments, the kit further comprises instructions for using the antibody or antigen-binding fragment thereof, immunoconjugate thereof, bispecific molecule thereof, or composition thereof disclosed herein for treating tumor burden in a subject, for treating and / or preventing a tumor or neoplasm, and / or for increasing or prolonging survival of a subject having a tumor or neoplasm. 4. Brief description of the drawings

[0036] The following detailed description, given by way of example and not intended to limit the invention to the specific embodiments described, may be understood in conjunction with the accompanying drawings, in which: [Brief explanation of the drawings]

[0037] [Figure 1]FIG. 1 shows the binding of anti-CD371 monoclonal phage preparations to HEK293H cells transfected with human CD371.

[0038] [Figure 2-1] Figures 2A and 2B show the binding of 1B10 and 1C3 formatted as human IgG1 to OCI cells. Figure 2A shows B10 (referred to as "1B10"). Figure 2B shows C3 (referred to as "1C3"). [Figure 2-2] Figures 2A and 2B show the binding of 1B10 and 1C3 formatted as human IgG1 to OCI cells. Figure 2A shows B10 (referred to as "1B10"). Figure 2B shows C3 (referred to as "1C3").

[0039] [Figure 3-1] FIG. 3 shows binding of scFv-Fc fusion proteins and scFv fragments to HEK293 cells expressing human CD371. [Figure 3-2] FIG. 3 shows binding of scFv-Fc fusion proteins and scFv fragments to HEK293 cells expressing human CD371. DETAILED DESCRIPTION OF THE INVENTION

[0040] 5. Detailed Description of the Embodiments of the Invention The presently disclosed subject matter provides anti-CD371 antibodies. Non-limiting embodiments of the present disclosure are illustrated herein and by example.

[0041] For clarity of the disclosure, and not by way of limitation, the detailed description is divided into the following subsections: 5.1. Definition; 5.2.CD371; 5.3. Anti-CD371 antibody; 5.4. Nucleic acids encoding antibodies or antigen-binding fragments thereof; 5.5. Pharmaceutical compositions and methods of treatment; 5.6. Kits; and 5.7.Detection Method 5.1.Definition

[0042] In the description that follows, certain conventions will be followed regarding the use of terms. In general, the terms used herein are intended to be interpreted consistent with the meanings that those terms would have to those skilled in the art.

[0043] An "antigen-binding protein" is a protein or polypeptide that contains an antigen-binding region or portion, i.e., that has a strong affinity for another molecule to which it binds. Antigen-binding proteins include antibodies, chimeric antigen receptors (CARs), and fusion proteins.

[0044] "Antibody" and "antibodies," as these terms are known in the art, refer to antigen-binding proteins of the immune system. The term "antibody" referred to herein includes a full-length antibody having an antigen-binding region, as well as any fragment thereof, or a single chain thereof, e.g., a single-chain variable fragment (scFv), in which the "antigen-binding portion" or "antigen-binding region" is retained. A naturally occurring "antibody" is a glycoprotein comprising at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds. Each heavy chain comprises a heavy chain variable region (referred to herein as V H ) and heavy chain constant (C H The heavy chain constant region is composed of three domains: CH1, CH2, and CH3. Each light chain contains a light chain variable region (referred to herein as V L ) and light chain constant C L The light chain constant region consists of C L It consists of one domain: V H and V L The regions can be further subdivided into regions of hypervariability called complementarity determining regions (CDRs) interspersed with regions that are more conserved called framework regions (FRs). H and V Lis composed of three CDRs and four FRs, arranged from the amino terminus to the carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens. The constant region of the antibody can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system.

[0045] The term "human antibody," as used herein, is intended to include antibodies having variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains a constant region, the constant region also is derived from a human germline immunoglobulin sequence. The human antibodies of the presently disclosed subject matter may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo).

[0046] The term "monoclonal antibody," as used herein, refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and / or bind to the same epitope, except for possible variant antibodies (generally minor variants) that contain, for example, naturally occurring mutations or that arise during production of the monoclonal antibody preparation. In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. Thus, the modifier "monoclonal" indicates the character of the antibody as obtained from a substantially homogeneous population of antibodies and should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the subject matter of the present disclosure can be produced by a variety of techniques, including, but not limited to, hybridoma methods, recombinant DNA methods, phage display methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin loci; such methods, as well as other exemplary methods for producing monoclonal antibodies, are described herein.

[0047] The term "recombinant human antibody," as used herein, includes all human antibodies prepared, expressed, generated, or isolated by recombinant means, e.g., (a) antibodies isolated from animals (e.g., mice) that are transgenic or transchromosomal for human immunoglobulin genes or hybridomas prepared therefrom (described further below), (b) antibodies isolated from host cells transformed to express human antibodies, e.g., from transfectomas, (c) antibodies isolated from recombinant combinatorial human antibody libraries, and (d) antibodies prepared, expressed, generated, or isolated by any other means involving splicing human immunoglobulin gene sequences into other DNA sequences. Such recombinant human antibodies have variable regions in which the framework and CDR regions are derived from human germline immunoglobulin sequences. However, in certain embodiments, such recombinant human antibodies may be subjected to in vitro mutagenesis (or, if animals transgenic for human Ig sequences are used, in vivo mutagenesis) so that the amino acid sequences of the VH and VL regions of the recombinant antibodies are similar to those of human germline VH and VL regions. H and V L It is a sequence that is derived from and relates to a sequence, but which may not naturally occur within the human antibody germline repertoire in vivo.

[0048] The term "humanized antibody" is intended to refer to antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences. Additional framework region modifications may be made within the human framework sequences.

[0049] The term "chimeric antibody" is intended to refer to an antibody in which the variable region sequences are derived from one species and the constant region sequences are derived from another species, e.g., an antibody in which the variable region sequences are derived from a murine antibody and the constant region sequences are derived from a human antibody.

[0050] As used herein, an antibody that "specifically binds to CD371" is an antibody that specifically binds to CD371 and has a cell count of about 5 x 10-7 M or lower, approximately 1×10 -7 M or lower, approximately 5 x 10 -8 M or lower, approximately 1×10 -8 M or lower, approximately 5 x 10 -9 M or lower, approximately 1×10 -9 M or lower, approximately 5 x 10 -10 M or lower, approximately 1×10 -10 M or lower, approximately 5 x 10 -11 M or lower, or about 1 x 10 -11 M or lower dissociation constant (K d ) and is intended to refer to an antibody that binds to CD371 (e.g., human CD371).

[0051] An "antibody that competes for binding" or "cross-competes for binding" with a reference antibody for binding to an antigen, e.g., CD371, refers to an antibody that blocks the binding of the reference antibody to the antigen (e.g., CD371) by 50% or more in a competition assay, and conversely, the reference antibody blocks the binding of the antibody to the antigen (e.g., CD371) by 50% or more in a competition assay. Exemplary competition assays are described in "Antibodies," Harlow and Lane (Cold Spring Harbor Press, Cold Spring Harbor, NY).

[0052] As used herein, "isotype" refers to the antibody class (e.g., IgM or IgG1) that is encoded by heavy chain constant region genes.

[0053] The phrases "antibody that recognizes an antigen" and "antibody specific for an antigen" are used interchangeably herein with the term "antibody that specifically binds to an antigen (eg, a CD371 polypeptide)."

[0054] The term "antigen-binding portion" or "antigen-binding region" of an antibody, as used herein, refers to the region or portion of an antibody that binds to an antigen and confers antigen specificity to the antibody; antigen-binding proteins, e.g., antibodies, include one or more fragments of an antibody that retain the ability to specifically bind to an antigen (e.g., a CD391 polypeptide). It has been shown that the antigen-binding function of an antibody can also be performed by fragments of a full-length antibody. Examples of antigen-binding fragments encompassed by the term "antibody fragment" of an antibody include V L , V H , C L and CH1 domains; F(ab)2 fragment, a bivalent fragment containing two Fab fragments linked by a disulfide bridge at the hinge region; V H and the Fd fragment consisting of the CH1 domain; the V of a single arm of the antibody L and V H Fv fragment consisting of domains; V H These include dAb fragments consisting of domains (Ward et al., Nature 1989;341:544-546); as well as isolated complementarity determining regions (CDRs).

[0055] Furthermore, the two domains of the Fv fragment, V L and V H are encoded by separate genes, but recombinant methods can be used to combine them into a single protein chain (V L Area and V H These antibody fragments may be joined by a synthetic linker that allows for pairing of the Fv domains to form a monovalent molecule. These are known as single-chain Fvs (scFvs); see, e.g., Bird et al., Science (1988); 242:423-426; and Huston et al., Proc Natl Acad Sci (1998); 85:5879-5883. These antibody fragments are obtained using conventional techniques known to those of skill in the art, and the fragments are screened for utility in the same manner as are intact antibodies.

[0056] An "antibody" or "antigen binding protein" is one that has been identified and separated and / or recovered from a component of its natural environment. A "synthetic antibody" or "recombinant antibody" is generally produced using recombinant techniques or peptide synthesis techniques known to those of skill in the art.

[0057] As used herein, the term "single-chain variable fragment" or "scFv" refers to a V H heavy chains (V::VL) of immunoglobulins (e.g., murine or human) covalently linked to form a VL heterodimer. H ) and light chain (V L ) is a fusion protein of the variable region of the heavy chain (V H ) and light chain (V L ) are either directly bonded or V H N-terminus of V L and the C-terminus of V H The C-terminus of V L The extracellular antigen-binding domain is connected to the N-terminus of the heavy chain variable region and the light chain variable region by a peptide-encoding linker (e.g., 10, 15, 20, or 25 amino acids). The linker is usually rich in glycine for flexibility and serine or threonine for solubility. The linker can connect the heavy chain variable region and the light chain variable region of the extracellular antigen-binding domain.

[0058] Non-limiting examples of linkers are disclosed in Shen et al., Anal. Chem. (2008) 80(6):1910-1917 and WO 2014 / 087010, the contents of which are incorporated herein by reference in their entireties. In certain embodiments, the linker is a G4S linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13, which is provided below. GGGGSGGGGSGGGSGGGGS [SEQ ID NO: 13]

[0059] In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 14, provided below: GGGGSGGGGSGGGGS [SEQ ID NO: 14]

[0060] In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO:64, provided below: GGGGSGGGGSGGGGSGGGSGGGGS [SEQ ID NO: 64]

[0061] In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 65, provided below: GGGGSGGGGSGGGGSGGGGSGGGSGGGGS [SEQ ID NO: 65]

[0062] In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 66, provided below: GGGGS [SEQ ID NO: 66]

[0063] In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO:67, provided below: GGGGSGGGGS [SEQ ID NO: 67]

[0064] Despite the removal of the constant region and the introduction of the linker, the scFv protein retains the specificity of the original immunoglobulin. Single-chain Fv polypeptide antibodies are composed of V, V, and VF polypeptides, as described by Huston, et al. Proc. Nat. Acad. Sci. USA, (1988);85:5879-5883). H and V LIt can be expressed from a nucleic acid containing the coding sequence. See also U.S. Patent Nos. 5,091,513, 5,132,405, and 4,956,778; and U.S. Patent Application Publication Nos. 20050196754 and 20050196754. Antagonistic scFvs with inhibitory activity have been described (see, e.g., Zhao et al., Hyrbidoma (Larchmt) (2008);27(6):455-51; Peter et al., J Cachexia Sarcopenia Muscle (2012);August 12; Shieh et al., J Imunol (2009);183(4):2277-85; Giomarelli et al., Thromb Haemost (2007);97(6):955-63; Fife et al., J Clin Invst (2006);116(8):2252-61; Brocks et al., Immunotechnology 1997 3(3):173-84; Moosmayer et al., Ther Immunol 1995 2(10:31-40)). Agonistic scFvs with stimulatory activity have been described (see also Peter et al., J Biol Chern (2003);25278(38):36740-7; Xie et al., Nat Biotech 1997 15(8):768-71; Ledbetter et al., Crit Rev Immunol (1997);17(5-6):427-55; Ho et al., BioChim Biophys Acta (2003);1638(3):257-66).

[0065] As used herein, "F(ab)" refers to the fragment of an antibody structure that binds to an antigen but is monovalent and does not have the Fc portion; for example, digestion of an antibody with the enzyme papain produces two F(ab) fragments and one Fc fragment (e.g., heavy (H) chain constant region; the Fc region that does not bind to antigen).

[0066] As used herein, "F(ab')2" refers to an antibody fragment produced by pepsin digestion of a whole IgG antibody, which fragment contains two antigen-binding (ab') (bivalent) regions, each of which contains two separate amino acid chains, i.e., a portion of a heavy chain and a light (L) chain linked by an S-type disulfide bond for antigen binding, with the remainder of the heavy chains linked to each other. The "F(ab')2" fragment can be divided into two individual Fab' fragments.

[0067] As used herein, the term "vector" refers to any genetic element, e.g., a plasmid, phage, transposon, cosmid, chromosome, virus, virion, etc., which, when associated with the appropriate control elements, is capable of replication and of transferring genetic sequences into cells. Thus, the term includes cloning and expression vehicles, as well as viral and plasmid vectors.

[0068] "CDR" is defined as the complementarity-determining region amino acid sequence of an antibody, which is the hypervariable region of the immunoglobulin heavy and light chains. See, for example, Kabat et al., Sequences of Proteins of Immunological Interest, 4th US Department of Health and Human Services, National Institutes of Health (1987), or the IMGT numbering system (Lefranc, The Immunologist (1999); 7:132-136; Lefranc et al., Dev. Comp. Immunol. (2003); 27:55-77). The term "hypervariable region" or "HVR" as used herein refers to each of the regions of an antibody variable domain whose sequence is hypervariable ("complementarity-determining region" or "CDR") and / or forms a structurally defined loop ("hypervariable loop") and / or contains antigen-contacting residues ("antigen contact groups"). Generally, an antibody contains three heavy chain and three light chain CDRs or CDR regions in the variable region. CDRs provide the majority of contact residues for antibody binding to the antigen or epitope. In certain embodiments, CDRs are identified according to the IMGT system. In certain embodiments, CDRs are identified using the IMGT numbering system, which can be accessed at http: / / www.imgt.org / IMGT_vquest / input.

[0069] The term "isolated" refers to a degree of separation from the original source or environment.

[0070] An "isolated antibody" is one that has been separated from components of its natural environment. In certain embodiments, the antibody is purified to greater than 95% or 99% purity, as determined, for example, by electrophoresis (e.g., SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatography (e.g., ion exchange or reverse-phase HPLC). For a review of methods for assessing antibody purity, see, for example, Flatman et al., J. Chromatogr (2007); B 848:79-87.

[0071] An "isolated nucleic acid" refers to a nucleic acid molecule that has been separated from a component of its natural environment. Isolated nucleic acid includes a nucleic acid molecule that is contained within a cell that ordinarily contains the nucleic acid molecule, but where the nucleic acid molecule is present extrachromosomally or at a chromosomal location that is different from its natural chromosomal location.

[0072] An "isolated nucleic acid encoding an antibody" (including reference to a particular antibody, e.g., an anti-KLB antibody) refers to one or more nucleic acid molecules encoding the antibody heavy and light chains (or fragments thereof), including such nucleic acid molecule(s) in a single vector separate vectors, and such nucleic acid molecule(s) present in one or more locations within a host cell.

[0073] The term "vector," as used herein, refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is linked. The term includes vectors as autonomously replicating nucleic acid structures and vectors that integrate into the genome of a host cell into which they are introduced. Certain vectors are capable of directing the expression of nucleic acids to which they are operatively linked. Such vectors are referred to herein as "expression vectors."

[0074] An "immunoconjugate" is an antibody conjugated to one or more heterologous molecules, including, but not limited to, a cytotoxic agent.

[0075] An "effective amount" (i.e., a "therapeutically effective amount") is an amount sufficient to produce beneficial or desired clinical results upon treatment. An effective amount can be administered to a subject in one or more doses. In terms of treatment, an effective amount is an amount sufficient to palliate, relieve, stabilize, reverse, or slow the progression of a disease, or otherwise reduce the pathological consequences of a disease. An effective amount is generally determined by a physician on a case-by-case basis and is within the skill of one of ordinary skill in the art. When determining an appropriate dosage to achieve an effective amount, several factors are typically taken into consideration. These factors include the age, sex, and weight of the subject, the condition being treated, the severity of the condition, and the form and effective concentration of the cells to be administered.

[0076] As used herein, an "individual" or "subject" refers to a vertebrate, such as a human or non-human animal, e.g., a mammal. Mammals include, but are not limited to, humans, primates, farm animals, sport animals, rodents, and pet animals. Non-limiting examples of non-human animal subjects include rodents, such as mice, rats, hamsters, and guinea pigs, rabbits, dogs, cats, sheep, pigs, goats, cows, horses, and non-human primates, such as apes and monkeys.

[0077] As used herein, "treatment" (and grammatical variations thereof, such as "treat" or "treating") refers to a clinical intervention that attempts to alter the natural history of the treated individual and can be performed either prophylactically or during the course of clinical pathology. Desirable effects of treatment include, but are not limited to, preventing the occurrence or recurrence of disease, alleviating symptoms, attenuating any direct or indirect pathological consequences of the disease, preventing metastasis, slowing the rate of disease progression, ameliorating or alleviating the disease state, and achieving remission or improving prognosis. In certain embodiments, the antibodies of the presently disclosed subject matter are used to delay the onset of disease, slowing the progression of disease, e.g., tumors (e.g., acute myeloid leukemia (AML)).

[0078] "Neoplasia" means a disease characterized by the pathological proliferation of cells or tissues and their subsequent migration or invasion into other tissues or organs. Neoplastic growth is typically uncontrolled and progressive, occurring under conditions that do not induce or would cause the cessation of normal cell proliferation.

[0079] The terms "comprises" and "comprising" are intended to have the broad meaning ascribed to them in U.S. patent law and may mean "includes," "including," etc.

[0080] As used herein, the term "about" or "approximately" means within an acceptable error range for a particular value as determined by one of ordinary skill in the art, which depends in part on how the value is measured or determined, i.e., the limitations of the measurement system. For example, "about" can mean within 3 or more standard deviations, as practiced in the art. Alternatively, "about" can mean a range of up to 20%, preferably up to 10%, more preferably up to 5%, and more preferably still up to 1% of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, preferably within 5-fold, and more preferably within 2-fold of a value.

[0081] As described herein, any concentration range, percentage range, ratio range, or integer range should be understood to include any integer value within the stated range, and, where appropriate, fractions thereof (such as tenths and hundredths of integers), unless otherwise specified.

[0082] Other aspects of the presently disclosed subject matter are described in the disclosure that follows and are within the scope of the presently disclosed subject matter. 5.2.CD371

[0083] CD371 (CEC12A), also known as DCAL-2, MICL, or CLL-1, is a 30 kD C-type lectin transmembrane glycoprotein. It is expressed in monocytes, granulocytes, natural killer (NK) cells, and basophils. CD371 is an immune inhibitory receptor that recruits the Src homology phosphatases SHP-1 and SHP-2 to its phosphorylated cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM) (Sancho et al., Annu Rev. Immunol (2012); 30:491-529; Yan et al., Front Immunol (2015); 6:408; Lahoud et al., J Immunol (2011); 187:842). CD371 has been implicated as a negative regulatory urate crystal (monosodium urate, MSU) receptor that controls autoimmune and inflammatory diseases (Neumann et al., Immunity (2014);40:389-99). CD371 is a negative regulator of granulocyte and monocyte function (Marshall et al., J Biol Chem (2004);279(15):14792-802; Pyz et al., Eur J Immunol (2008);38(4):1157-63).

[0084] In certain embodiments, the CD371 is human CD371 comprising or consisting of the amino acid sequence having NCBI reference number: NP_612210.4 (SEQ ID NO: 15), or a fragment thereof. SEQ ID NO: 15 is provided below. [ka]

[0085] In certain embodiments, CD371 comprises or consists of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, at least about 100% identical to the amino acid sequence set forth in SEQ ID NO: 15, or a fragment thereof. 5.3. Anti-CD371 antibody

[0086] The antibodies of the presently disclosed subject matter are characterized by a particular functional feature or property of the antibody, for example, the antibody specifically binds to CD371 (e.g., binds to human CD371).

[0087] In certain embodiments, the antibodies or antigen-binding fragments of the disclosure are administered at a concentration of, for example, 1×10 -6 M or lower, e.g., about 1 x 10 -7 M or lower, approximately 1×10 -8 M or lower, approximately 1×10 -9 M or lower, approximately 1×10 -10 M or lower, or about 1 x 10 -11 M or lower dissociation constant (K d In certain embodiments, the antibody or antigen-binding fragment of the disclosure binds to CD371 (e.g., human CD371) with a binding affinity having a binding affinity of about 1 x 10 -8 M to approximately 1 x 10 -7 K between M d In certain embodiments, the antibody or antigen-binding fragment of the disclosure binds to CD371 (e.g., human CD371) at a concentration of about 1 x 10 -9 M to approximately 1 x 10 -8 K between M d In certain embodiments, the antibody or antigen-binding fragment of the disclosure binds to CD371 (e.g., human CD371) at a concentration of about 1.5 x 10 -8 M or lower K d In certain embodiments, the antibody or antigen-binding fragment of the disclosure binds to CD371 (e.g., human CD371) at a concentration of about 1 x 10 -8 M or lower K d In certain embodiments, the antibody or antigen-binding fragment of the disclosure binds to CD371 (e.g., human CD371) at a concentration of about 1 x 10 -8 K of M d In certain embodiments, the antibodies or antigen-binding fragments of the disclosure bind to CD371 (e.g., human CD371) at a concentration of about 9 x 10 -9 K of Md and binds to CD371 (e.g., human CD371).

[0088] The heavy and light chains of the antibodies or antigen-binding fragments of the disclosure may be full-length (e.g., the antibody may comprise at least one (e.g., one or two) complete heavy chain and at least one (e.g., one or two) complete light chain), or may comprise an antigen-binding portion (Fab, F(ab'), Fv, or single-chain Fv fragment ("scFv"). In certain embodiments, the antibody heavy chain constant region is chosen from, e.g., IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE, particularly, e.g., IgG1, IgG2, IgG3, and IgG4. In certain embodiments, the immunoglobulin isotype is IgG1 (e.g., human IgG1). In certain embodiments, the antibody light chain constant region is chosen from, e.g., kappa or lambda, particularly kappa. 5.3.1. Single-Chain Variable Fragments (scFv)

[0089] In certain embodiments, the subject matter of the present disclosure includes an antibody or antigen-binding fragment thereof having an scFv sequence fused to one or more constant domains to form an antibody with the Fc region of a human immunoglobulin, resulting in a bivalent protein and increasing the overall avidity and stability of the antibody. In addition, the Fc portion allows for the direct conjugation of other molecules to the antibody, including, but not limited to, fluorescent dyes, cytotoxins, radioisotopes, etc., for use in antigen quantitation studies, to immobilize antibodies for affinity measurements, for targeted delivery of therapeutic agents, to test for Fc-mediated cytotoxicity using immune effector cells, and many other uses.

[0090] The results presented herein highlight the specificity, sensitivity, and utility of the antibodies or antigen-binding fragments of the present disclosure in targeting CD371 polypeptides (e.g., human CD371).

[0091] The molecules of the present disclosure are based on the identification and selection, using phage display, of single-chain variable fragments (scFvs), the amino acid sequences of which confer the specificity of the molecule for a CD371 polypeptide of interest and form the basis of all antigen-binding proteins of the present disclosure. Thus, scFvs can be used to design a diverse series of "antibody" molecules, including, for example, full-length antibodies, fragments thereof, such as Fab and F(ab')2, minibodies, fusion proteins including scFv-Fc fusions, multivalent antibodies, i.e., antibodies with two or more specificities for the same or different antigens, e.g., bispecific antibodies, tribodies, etc. (See Cuesta et al., Multivalent antibodies: when design surpasses evolution. Trends in Biotechnology 28:355-362 2010).

[0092] In certain embodiments, the antigen-binding protein is a full-length antibody, and the heavy and light chains of the antibodies of the presently disclosed subject matter may be full-length (e.g., the antibody may comprise at least one or two complete heavy chains and at least one, preferably two complete light chains) or may comprise an antigen-binding fragment (Fab, F(ab')2, Fv, or scFv). In certain embodiments, the antibody heavy chain constant region is selected from IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, IgE, and the like. In certain embodiments, the immunoglobulin isotype is selected from IgG1, IgG2, IgG3, and IgG4. In certain embodiments, the immunoglobulin isotype is IgG1 (e.g., human IgG1). The choice of antibody isotype may depend on the immune effector function the antibody is designed to elicit.

[0093] In the construction of recombinant immunoglobulins, suitable amino acid sequences for the constant regions of various immunoglobulin isotypes and methods for the production of a wide range of antibodies are known to those skilled in the art.

[0094] In certain embodiments, the anti-CD371 scFv is a V selected from Table 1. H and V L In certain embodiments, the anti-CD371 scFv is a scFv-Fc fusion protein or a full-length human IgG having a V domain or CDR. H In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO:2. L In certain embodiments, the scFv is designed as "B031_P1_PH1B10" (also referred to as "B10").

[0095] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 1. H and V comprising the amino acid sequence set forth in SEQ ID NO:2 L In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO: 28, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 29 or a conservative modification thereof H CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 30 or a conservative modification thereof H SEQ ID NOs: 28 to 30 are provided in Table 1.

[0096] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 31 or a conservative modification thereof. L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 32 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 33 or a conservative modification thereof L SEQ ID NOs: 31 to 33 are provided in Table 1.

[0097] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 28 or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 29 or a conservative modification thereof HCDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 30 or a conservative modification thereof H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 31 or a conservative modification thereof L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 32 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 33 or a conservative modification thereof L Includes CDR3.

[0098] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 28. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 29 H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 30 H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 31 L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 32 L CDR2 and V comprising the amino acid sequence set forth in SEQ ID NO: 33 L Includes CDR3.

[0099] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 1. H and V comprising the amino acid sequence set forth in SEQ ID NO:2 L In certain embodiments, V H and V L are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0100] In certain embodiments, the variable region is a heavy chain variable region (V H In certain embodiments, the variable regions are linked together such that, from the N-terminus to the C-terminus, V H -V L In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 68, as provided in Table 1.

[0101] In certain embodiments, the light chain variable region (V L ) is located at the N-terminus. In certain embodiments, the variable region is L -V H In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 16. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 16 is set forth in SEQ ID NO: 22. SEQ ID NOs: 16 and 22 are provided in Table 1 below. Table 1 [Table 1-1] [Table 1-2]

[0102] In certain embodiments, the anti-CD371 scFv comprises a V chain comprising the amino acid sequence set forth in SEQ ID NO: 3, optionally with a linker sequence, e.g., a linker peptide, between the heavy chain variable region and the light chain variable region. H and V comprising the amino acid sequence set forth in SEQ ID NO:4 L In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13. SEQ ID NOs: 3 and 4 are provided in Table 2 below. In certain embodiments, the scFv is designated as "B031_P1_PH1C3" (also referred to as "C3").

[0103] In certain embodiments, the anti-CD371 scFv is a V scFv selected from Table 2. H and V L In certain embodiments, the anti-CD371 scFv is a scFv-Fc fusion protein or a full-length human IgG having a V domain or CDR. H In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO:4. L In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO:3. Hand V comprising the amino acid sequence set forth in SEQ ID NO:4 L Includes.

[0104] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 34, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 35 or a conservative modification thereof H CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 36 or a conservative modification thereof H SEQ ID NOs: 34-36 are provided in Table 2.

[0105] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 37 or a conservative modification thereof. L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 38 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 39 or a conservative modification thereof L SEQ ID NOs: 37-39 are provided in Table 2.

[0106] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 34, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 35 or a conservative modification thereof H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 36 or a conservative modification thereof H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 37 or conservative modifications thereof L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 38 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 39 or a conservative modification thereof L Includes CDR3.

[0107] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 34. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 35 HCDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 36 H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 37 L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 38 L CDR2 and V comprising the amino acid sequence set forth in SEQ ID NO: 39 L Includes CDR3.

[0108] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO:3. H and V comprising the amino acid sequence set forth in SEQ ID NO:4 L In certain embodiments, V H and V L are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0109] In certain embodiments, the heavy chain variable region (V H ) is located at the N-terminus. In certain embodiments, the variable region is H -V L In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 69, as provided in Table 2.

[0110] In certain embodiments, the light chain variable region (V L ) is located at the N-terminus. In certain embodiments, the variable region is L -V H In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 17. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 17 is set forth in SEQ ID NO: 23. SEQ ID NOs: 17 and 23 are provided in Table 2 below. Table 2 [Table 2-1] [Table 2-2]

[0111] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 5, optionally with a linker sequence, e.g., a linker peptide, between the heavy chain variable region and the light chain variable region. H and V comprising the amino acid sequence set forth in SEQ ID NO:6 H In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13. SEQ ID NOs: 5 and 6 are provided in Table 3 below. In certain embodiments, the anti-CD371 scFv is designated as "B031_P1_PH1D6" (also referred to as "D6").

[0112] In certain embodiments, the anti-CD371 scFv is a V scFv selected from Table 3. H and V L In certain embodiments, the anti-CD371 scFv is a scFv-Fc fusion protein or a full-length human IgG having a V domain or CDR. H In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO:6. L Includes.

[0113] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO:5. H and V comprising the amino acid sequence set forth in SEQ ID NO:6 L Includes.

[0114] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 40, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 41 or a conservative modification thereof H CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 42 or a conservative modification thereof H SEQ ID NOs: 40-42 are provided in Table 3.

[0115] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 43, or a conservative modification thereof. L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 44 or conservative modifications thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 45 or a conservative modification thereof L SEQ ID NOs: 43-45 are provided in Table 3.

[0116] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 40, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 41 or a conservative modification thereof H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 42 or a conservative modification thereof H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 43 or conservative modifications thereof L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 44 or conservative modifications thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 45 or a conservative modification thereof L Includes CDR3.

[0117] In certain embodiments, the anti-CD371 scFv comprises a V scFv comprising the amino acid sequence set forth in SEQ ID NO: 40. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 41 H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 42 H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 43 L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 44 L CDR2 and V comprising the amino acid sequence set forth in SEQ ID NO: 45 L Includes CDR3.

[0118] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO:5. H and V comprising the amino acid sequence set forth in SEQ ID NO:6 L In certain embodiments, VH and V L are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0119] In certain embodiments, the heavy chain variable region (V H ) is located at the N-terminus. In certain embodiments, the variable region is H -V L In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 70, as provided in Table 3.

[0120] In certain embodiments, the light chain variable region (V L ) is located at the N-terminus. In certain embodiments, the variable region is L -V H In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 18. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 18 is set forth in SEQ ID NO: 24. SEQ ID NOs: 18 and 24 are provided in Table 3 below. Table 3 [Table 3-1] [Table 3-2]

[0121] In certain embodiments, the anti-CD371 scFv comprises a V chain comprising the amino acid sequence set forth in SEQ ID NO: 7, optionally with a linker sequence, e.g., a linker peptide, between the heavy chain variable region and the light chain variable region. H and V comprising the amino acid sequence set forth in SEQ ID NO:8 L In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13. SEQ ID NOs: 7 and 8 are provided in Table 4 below. In certain embodiments, the anti-CD371 scFv is designated as "B031_P1_PH2A11" (also referred to as "A11").

[0122] In certain embodiments, the anti-CD371 scFv is a V scFv selected from Table 4. H and V L In certain embodiments, the anti-CD371 scFv is a scFv-Fc fusion protein or a full-length human IgG having a V domain or CDR. H In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO: 14. L In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO:7. H and V comprising the amino acid sequence set forth in SEQ ID NO:8 L SEQ ID NOs: 7 and 8 are provided in Table 4.

[0123] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 46, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 47 or conservative modifications thereof H CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 48 or a conservative modification thereof H SEQ ID NOs: 46-48 are provided in Table 4.

[0124] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 49 or a conservative modification thereof. L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 50 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 51 or a conservative modification thereof L SEQ ID NOs: 49-51 are provided in Table 4.

[0125] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 46, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 47 or conservative modifications thereof HCDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 48 or a conservative modification thereof H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 49 or a conservative modification thereof L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 50 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 51 or a conservative modification thereof L Includes CDR3.

[0126] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 46. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 47 H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 48 H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 49 L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 50 L CDR2 and V comprising the amino acid sequence set forth in SEQ ID NO: 51 L Includes CDR3.

[0127] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO:7. H and V comprising the amino acid sequence set forth in SEQ ID NO:8 L In certain embodiments, V H and V L are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0128] In certain embodiments, the heavy chain variable region (V H ) is located at the N-terminus. In certain embodiments, the variable region is H -V L In certain embodiments, the CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 71, as provided in Table 4.

[0129] In certain embodiments, the light chain variable region (V L) is located at the N-terminus. In certain embodiments, the variable region is L -V H In certain embodiments, the scFv comprises the amino acid sequence set forth in SEQ ID NO: 19. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 19 is set forth in SEQ ID NO: 25. SEQ ID NOs: 19 and 25 are provided in Table 4 below. Table 4 [Table 4-1] [Table 4-2]

[0130] In certain embodiments, the anti-CD371 scFv comprises a V chain comprising the amino acid sequence set forth in SEQ ID NO: 9, optionally with a linker sequence, e.g., a linker peptide, between the heavy chain variable region and the light chain variable region. H and V comprising the amino acid sequence set forth in SEQ ID NO: 10 L In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13. SEQ ID NOs: 9 and 10 are provided in Table 5. In certain embodiments, the anti-CD371 scFv is designated as "B031_P1_PH2E4" (also referred to as "E4").

[0131] In certain embodiments, the anti-CD371 scFv is a V scFv selected from Table 5. H and V L In certain embodiments, the anti-CD371 scFv is a VFc fusion protein or a full-length human IgG having a VFc region or CDR. H In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO: 10. L In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO:9. H and V comprising the amino acid sequence set forth in SEQ ID NO: 10 L Includes.

[0132] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 52, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 53 or a conservative modification thereof H CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 54 or a conservative modification thereof H SEQ ID NOs: 52-54 are provided in Table 5.

[0133] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 55 or a conservative modification thereof. L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 56 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 57 or a conservative modification thereof L SEQ ID NOs: 55-57 are provided in Table 5.

[0134] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 52, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 53 or a conservative modification thereof H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 54 or conservative modifications thereof H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 55 or conservative modifications thereof L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 56 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 57 or a conservative modification thereof L Includes CDR3.

[0135] In certain embodiments, the anti-CD371 scFv comprises a V scFv comprising the amino acid sequence set forth in SEQ ID NO: 52. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 53 H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 54 HCDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 55 L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 56 L CDR2 and V comprising the amino acid sequence set forth in SEQ ID NO: 57 L Includes CDR3.

[0136] In certain embodiments, the anti-CD371 scFv comprises a V scFv comprising the amino acid sequence set forth in SEQ ID NO:9. H and V comprising the amino acid sequence set forth in SEQ ID NO: 10 L In certain embodiments, V H and V L are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0137] In certain embodiments, the light chain variable region (V L ) is located at the N-terminus. In certain embodiments, the variable region is L -V H In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 20. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 20 is set forth in SEQ ID NO: 26. SEQ ID NOs: 20 and 26 are provided in Table 5 below. Table 5 [Table 5]

[0138] In certain embodiments, the anti-CD371 scFv comprises a V chain comprising the amino acid sequence set forth in SEQ ID NO: 11, optionally with a linker sequence, e.g., a linker peptide, between the heavy chain variable region and the light chain variable region. H and V comprising the amino acid sequence set forth in SEQ ID NO: 12 LIn certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13. SEQ ID NOs: 11 and 12 are provided in Table 6 below. In certain embodiments, the anti-CD371 scFv is designated as "B031_P1_PH2E8" (also referred to as "E8").

[0139] In certain embodiments, the anti-CD371 scFv is a V scFv selected from Table 6. H and V L In certain embodiments, the anti-CD371 scFv is a scFv-Fc fusion protein or a full-length human IgG having a V domain or CDR. H In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO: 12. L In certain embodiments, the anti-CD371 scFv comprises a V comprising the amino acid sequence set forth in SEQ ID NO: 11. H and V comprising the amino acid sequence set forth in SEQ ID NO: 12 L Includes.

[0140] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 58, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 59 or a conservative modification thereof H CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 60 or a conservative modification thereof H SEQ ID NOs: 58-60 are provided in Table 6.

[0141] In certain embodiments, the anti-CD371 scFv comprises a V nucleotide comprising the amino acid sequence set forth in SEQ ID NO: 61, or a conservative modification thereof. L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 62 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 63 or a conservative modification thereof L SEQ ID NOs: 61-63 are provided in Table 6.

[0142] In certain embodiments, the anti-CD371 scFv comprises a VFv comprising the amino acid sequence set forth in SEQ ID NO: 58, or a conservative modification thereof. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 59 or a conservative modification thereof H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 60 or a conservative modification thereof H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 61 or a conservative modification thereof L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 62 or a conservative modification thereof L CDR2 and a V comprising the amino acid sequence set forth in SEQ ID NO: 63 or a conservative modification thereof L Includes CDR3.

[0143] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 58. H CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 59 H CDR2, V comprising the amino acid sequence set forth in SEQ ID NO: 60 H CDR3, V comprising the amino acid sequence set forth in SEQ ID NO: 61 L CDR1, V comprising the amino acid sequence set forth in SEQ ID NO: 62 L CDR2 and V comprising the amino acid sequence set forth in SEQ ID NO: 63 L Includes CDR3.

[0144] In certain embodiments, the anti-CD371 scFv comprises a V domain comprising the amino acid sequence set forth in SEQ ID NO: 11. H and V comprising the amino acid sequence set forth in SEQ ID NO: 12 L In certain embodiments, V H and V L are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0145] In certain embodiments, the heavy chain variable region (V H ) is located at the N-terminus. In certain embodiments, the variable region is H -V LIn certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 73, as provided in Table 6.

[0146] In certain embodiments, the light chain variable region (V L ) is located at the N-terminus. In certain embodiments, the variable region is L -V H In certain embodiments, the anti-CD371 scFv comprises the amino acid sequence set forth in SEQ ID NO: 21. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 21 is set forth in SEQ ID NO: 27. SEQ ID NOs: 21 and 27 are provided in Table 6 below. Table 6 [Table 6-1] [Table 6-2] Monoclonal Antibodies

[0147] The presently disclosed subject matter provides antibodies (e.g., human antibodies, e.g., human monoclonal antibodies) that specifically bind to CD371 (e.g., human CD371). The V of anti-CD371 antibodies B031_P1_PH1B10 (also referred to as "B10"), B031_P1_PH1C3 (also referred to as "C3"), B031_P1_PH1D6 (also referred to as "D6"), B031_P1_PH2A11 (also referred to as "A11"), B031_P1_PH2E4 (also referred to as "E4"), and B031_P1_PH2E8 (also referred to as "E8"). H The amino acid sequences are shown in SEQ ID NOs: 1, 3, 5, 7, 9 and 11, respectively. L The amino acid sequences are shown in SEQ ID NOs: 2, 4, 6, 8, 10 and 12, respectively.

[0148] Considering that each of the B031_P1_PH1B10 (B10), B031_P1_PH1C3 (C3), B031_P1_PH1D6 (D6), B031_P1_PH2A11 (A11), B031_P1_PH2E4 (E4), and B031_P1_PH2E8 (E8) antibodies can bind to CD371, V H and V sequences can be "mixed and matched" to create other anti-CD371 binding molecules. The binding of such "mixed and matched" antibodies to CD371 can be tested using binding assays known in the art, including, for example, ELISAs, Western blots, RIAs, and Biacore analysis. Preferably, V H and V L When chains are mixed and matched, a particular V H / V L V derived from involution H The sequence is structurally similar to V H Similarly, a particular V H / V L V derived from involution L The sequence is structurally similar to V L It is replaced by an array.

[0149] In certain embodiments, the presently disclosed subject matter comprises: (a) a heavy chain variable region (V) comprising an amino acid sequence selected from SEQ ID NOs: 1, 3, 5, 7, 9, and 11; H and (b) a light chain variable region (V) comprising an amino acid sequence selected from SEQ ID NOs: 2, 4, 6, 8, 10, and 12. L ), wherein the antibody or antigen-binding fragment specifically binds to CD371, e.g., human CD371. H and V L teeth, (a) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 1, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 2; or (b) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 3, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 4; (c) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 5, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 6; (d) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 8; (e) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 9, and a light chain variable region comprising amino acids having the sequence set forth in SEQ ID NO: 10; and (f) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 11, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 12. is selected from.

[0150] In certain embodiments, the presently disclosed subject matter provides antibodies or antigen-binding fragments thereof comprising the heavy and light chain CDR1, CDR2 and CDR3 of B10, C3, D6, A11, E4 and E8.

[0151] Vs at B10, C3, D6, A11, E4 and E8 H The amino acid sequences of CDR1 are shown in SEQ ID NOs: 28, 34, 40, 46, 52, and 58, respectively. H The amino acid sequences of CDR2 are shown in SEQ ID NOs: 29, 35, 41, 47, 53 and 59, respectively. H The amino acid sequences of CDR3 are shown in SEQ ID NOs: 30, 36, 42, 48, 54 and 60, respectively.

[0152] Vs at B10, C3, D6, A11, E4 and E8 L The amino acid sequences of CDR1 are shown in SEQ ID NOs: 31, 37, 43, 49, 55, and 61, respectively. L The amino acid sequences of CDR2 are shown in SEQ ID NOs: 32, 38, 44, 50, 56 and 62, respectively. LThe amino acid sequences of CDR3 are set forth in SEQ ID NOs: 33, 39, 45, 51, 57, and 63, respectively. CDR regions are delineated using the IMGT system. In certain embodiments, CDR regions are delineated using the IMGT numbering system, accessible at http: / / www.imgt.org / IMGT_vquest / input.

[0153] Considering that each of these antibodies or antigen-binding fragments thereof binds to CD371 and that antigen-binding specificity is primarily provided by the CDR1, CDR2, and CDR3 regions, V H CDR1, CDR2 and CDR3 sequences and V L CDR1, CDR2, and CDR3 sequences may be "mixed and matched" (i.e., CDRs from different antibodies may be mixed and matched, but each antibody may have a V H CDR1, CDR2 and CDR3 and V L Other anti-CD371 binding molecules can be created that contain CDR1, CDR2, and CDR3. The binding of such "mixed and matched" antibodies to CD371 can be tested using the binding assays described above. H When CDR sequences are mixed and matched, a particular V H The CDR1, CDR2 and / or CDR3 sequences derived from the V sequence are replaced with structurally similar CDR sequence(s). L When CDR sequences are mixed and matched, a particular V L The CDR1, CDR2 and / or CDR3 sequences derived from the sequence are replaced with structurally similar CDR sequence(s). H and V L The array contains one or more V H and / or V L It will be readily apparent to one skilled in the art that the CDR regions can be generated by replacing the sequences with structurally similar sequences derived from the CDR sequences of antibodies or antigen-binding fragments thereof B10, C3, D6, A11, E4 and E8 disclosed herein.

[0154] In certain embodiments, the subject matter of the present disclosure comprises: (a) a heavy chain variable region CDR1 comprising an amino acid sequence selected from SEQ ID NOs: 28, 34, 40, 46, 52, and 58; (b) a heavy chain variable region CDR2 comprising an amino acid sequence selected from SEQ ID NOs: 29, 35, 41, 47, 53, and 59; (c) a heavy chain variable region CDR3 comprising an amino acid sequence selected from SEQ ID NOs: 30, 36, 42, 48, 54, and 60; (d) a light chain variable region CDR1 comprising an amino acid sequence selected from SEQ ID NOs: 31, 37, 43, 49, 55, and 61; (e) a light chain variable region CDR2 comprising an amino acid sequence selected from SEQ ID NOs: 32, 38, 44, 50, 56, and 62; and (f) a light chain variable region CDR3 comprising an amino acid sequence selected from SEQ ID NOs: 33, 39, 45, 51, 57, and 63; The present invention provides an antibody or antigen-binding fragment comprising:

[0155] In certain embodiments, the antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 30 or SEQ ID NO: 29; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31; (e) a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 33, SEQ ID NO: 33, or SEQ ID NO: 32; (f) a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33 Includes.

[0156] In certain embodiments, the antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37; (e) a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38; and (f) A light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 39.

[0157] In certain embodiments, the antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43; (e) a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44; and (f) a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45 Includes.

[0158] In certain embodiments, the antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49; (e) a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 50; and (f) a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 51 Includes.

[0159] In certain embodiments, the antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55; (e) a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56; and (f) a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57 Includes.

[0160] In certain embodiments, the antibody or antigen-binding fragment thereof (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61; (e) a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62; and (f) a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 Includes.

[0161] The constant / framework regions of the anti-CD371 antibodies disclosed herein can be altered, for example, by amino acid substitution, to modify the properties of the antibody (e.g., binding affinity to the antigen, F C The antibody may be modified to increase or decrease one or more of: receptor binding, carbohydrate of the antibody, e.g., glycosylation, fucosylation, etc., number of cysteine ​​residues, effector cell function, complement function, or introduction of conjugation sites).

[0162] In certain embodiments, the anti-CD371 antibody of the present disclosure is a fully human antibody, such as any one of B10, C3, D6, A11, E4, and E8. Fully human mAbs cause serious side effects when administered to humans, including anaphylactic and hypersensitivity reactions.

[0163] The use of phage display libraries has made it possible to select large antibody repertoires for unique, rare Abs directed against highly defined epitopes (see McCafferty et al., Phage antibodies: filamentous phage displaying antibody variable domains. Nature, 348: 552-554 for further details regarding phage display). This has enabled the rapid identification of highly specific human Fab or single-chain Fv (scFv) fragments directed against tumor antigen-derived peptide-MHC complex molecules. Additionally, by using Fab fragments to engineer full-length monoclonal antibodies (mAbs), it is possible to directly generate therapeutic human mAbs, avoiding the several months of time-consuming research typically required for the development of therapeutic mAbs. The subject matter of the present disclosure concerns developing fully human mAbs that recognize a human CD371 polypeptide (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 23), for example, cancer therapy, e.g., for treating AML. 5.3.3. Homologous Antibodies

[0164] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises heavy and light chain variable regions comprising amino acid sequences that are homologous or identical to the amino acid sequences of an antibody described herein (e.g., B10, C3, D6, A11, E4, and E8 antibodies), and said antibody or antigen-binding fragment thereof retains the desired functional properties of the anti-CD371 antibody or antigen-binding fragment thereof of the presently disclosed subject matter.

[0165] For example, the presently disclosed subject matter provides an antibody or antigen-binding fragment or portion thereof comprising a heavy chain variable region and a light chain variable region, (a) the heavy chain variable region comprises an amino acid sequence that is at least about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11; (b) the light chain variable region comprises an amino acid sequence that is at least about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, or SEQ ID NO:12; The antibody or antigen-binding fragment thereof is administered at a concentration of 1×10 -7 M or lower K d or 1×10 -8 M or lower K d It specifically binds to human CD371.

[0166] In certain embodiments, V H and / or V L The amino acid sequence can be at least about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% homologous or identical to the sequences set forth above. H and V L V with high (i.e., 80% or higher) homology or identity to the region H and V L Antibodies having regions can be obtained by mutagenesis (e.g., site-directed or PCR-mediated mutagenesis) followed by testing the encoded altered antibodies for retained function (i.e., binding affinity) using the binding assays described herein.

[0167] As used herein, the percent homology between two amino acid sequences is equal to the percent identity between the two sequences. The percent identity or homology between two sequences is a function of the number of identical positions shared by the sequences (i.e., % homology = number of identical positions / total number of positions x 100), taking into account the number of gaps that need to be introduced for optimal alignment of the two sequences and the length of each gap. The comparison of sequences and determination of percent identity between two sequences can be achieved using a mathematical algorithm, as described in the non-limiting examples below.

[0168] The percent homology or identity between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., (1988);14:11-17) incorporated into the ALIGN program (version 2.0) using a PAM120 weighted residue table, a gap length penalty of 12, and a gap penalty of 4. Additionally, the percent homology between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol. (1970) 48:444-453) algorithm incorporated into the GAP program in the GCG software package (available at www.gcg.com) using either a Blossum62 matrix or a PAM250 matrix, and gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6.

[0169] Additionally or alternatively, the protein sequences of the presently disclosed subject matter can be further used as a "query sequence" to perform a search against public databases, for example, to identify related sequences. Such searches can be performed using the XBLAST program (version 2.0) of Altschul et al., J Mol Biol (1990); 215:403-10. BLAST protein searches can be performed using the XBLAST program, score=50, word length=3, to obtain amino acid sequences homologous to the antibody molecules of the present invention. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul et al., Nucleic Acids Res (1997); 25(17):3389-3402. When utilizing BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used. (). 5.3.4. Antibodies with Conservative Modifications

[0170] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a heavy chain variable region comprising CDR1, CDR2, and CDR3 sequences and a light chain variable region comprising CDR1, CDR2, and CDR3 sequences, wherein one or more of these CDR sequences comprise a specific amino acid sequence based on a preferred antibody described herein (e.g., B10, C3, D6, A11, E4, and E8 antibodies), or a conservative modification thereof, and wherein the antibody retains the desired functional properties of an anti-CD371 antibody or antigen-binding fragment thereof of the presently disclosed subject matter. The presently disclosed subject matter provides an antibody or antigen-binding fragment or portion thereof comprising a heavy chain variable region comprising CDR1, CDR2, and CDR3 sequences and a light chain variable region comprising CDR1, CDR2, and CDR3 sequences, (a) the heavy chain variable region CDR3 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 30, 36, 42, 48, 54, and 60, and conservative modifications thereof; (b) the light chain variable region CDR3 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 33, 39, 45, 51, 57, and 63, and conservative modifications thereof; and The antibody or antigen-binding fragment thereof is administered at a concentration of 1×10 -7 M or lower K d or 1×10 -8 M or lower K d and binds to human CD371.

[0171] In certain embodiments, the heavy chain variable region CDR3 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 30, 36, 42, 48, 54, and 60, and conservative modifications thereof, and the light chain variable region CDR3 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 33, 39, 45, 51, 57, and 63, and conservative modifications thereof.

[0172] In certain embodiments, the heavy chain variable region CDR2 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 29, 35, 41, 47, 53, and 59, and conservative modifications thereof, and the light chain variable region CDR2 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 32, 38, 44, 50, 56, and 62, and conservative modifications thereof.

[0173] In certain embodiments, the heavy chain variable region CDR1 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 28, 34, 40, 46, 52, and 58, and conservative modifications thereof, and the light chain variable region CDR1 sequence comprises an amino acid sequence selected from the amino acid sequences of SEQ ID NOs: 31, 37, 43, 49, 55, and 61, and conservative modifications thereof.

[0174] As used herein, the term "conservative sequence modifications" refers to amino acid modifications that do not significantly affect or alter the binding characteristics of the antibody containing the amino acid sequence. Such conservative modifications include amino acid substitutions, additions, and deletions. Modifications can be introduced into the antibodies of the present invention by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis.

[0175] A conservative amino acid substitution is one in which an amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues with similar side chains have been defined in the art. Exemplary conservative amino acid substitutions are shown in Table 7. Amino acid substitutions can be introduced into an antibody of interest, and the product screened for a desired activity, such as retention / improvement of antigen binding, reduced immunogenicity, or improved ADCC or CDC. In certain embodiments, the sequences disclosed herein, e.g., CDR sequences, V H Array or V L The sequences may have up to about 1, up to about 2, up to about 3, up to about 4, up to about 5, up to about 6, up to about 7, up to about 8, up to about 9 or up to about 10 modified and / or substituted amino acid residues. Table 7 [Table 7]

[0176] Amino acids can be grouped according to common side chain properties: - Hydrophobic: Norleucine, Met, Ala, Val, Leu, Ile; - Neutral, hydrophilic: Cys, Ser, Thr, Asn, Gln; - Acidic: Asp, Glu; - Basic: His, Lys, Arg; - Residues that influence chain orientation: Gly, Pro; - Aromatics: Trp, Tyr, Phe.

[0177] Non-conservative substitutions involve exchanging a member of one of these classes for another class. 5.3.5. Anti-CD371 Antibodies that Cross-Compete for Binding to CD371 with Anti-CD371 Antibodies of the Invention

[0178] The presently disclosed subject matter provides antibodies or antigen-binding fragments thereof that cross-compete with any of the disclosed anti-CD371 antibodies for binding to CD371 (e.g., human CD371). For example, without limitation, a cross-competing antibody may bind to the same epitope region, e.g., the same epitope, an adjacent epitope, or an overlapping epitope, as any of the anti-CD371 antibodies or antigen-binding fragments thereof of the presently disclosed subject matter. In certain embodiments, the reference antibody or reference antigen-binding fragment thereof for cross-competition studies may be any one of the anti-CD371 antibodies or antigen-binding fragments thereof disclosed herein, e.g., the B10, C3, D6, A11, E4, and E8 antibodies.

[0179] Such cross-competing antibodies can be identified based on their ability to cross-compete with any one of the anti-CD371 antibodies or antigen-binding fragments thereof of the present disclosure in a standard CD371 binding assay. For example, Biacore analysis, ELISA assays, or flow cytometry can be used to demonstrate cross-competition with the antibodies of the presently disclosed subject matter. The ability of the test antibody to inhibit the binding of, for example, any one of the anti-CD371 antibodies (e.g., B10, C3, D6, A11, E4, and E8 antibodies) of the present disclosure to CD371 (e.g., human CD371) demonstrates that the test antibody can compete with any one of the anti-CD371 antibodies or antigen-binding fragments thereof of the present disclosure for binding to CD371 (e.g., human CD371), and thus binds to the same epitope region on CD371 (e.g., human CD371) as any one of the anti-CD371 antibodies or antigen-binding fragments thereof of the present disclosure. In certain embodiments, the cross-competing antibody or antigen-binding fragment thereof binds to the same epitope on CD371 (e.g., human CD371) as any one of the anti-CD371 antibodies or antigen-binding fragments thereof of the present disclosure. 5.3.6. Characterization of Antibody Binding to Antigen

[0180] The subject antibodies or antigen-binding fragments of the present disclosure can be tested for binding to CD371, for example, by standard ELISA. To determine whether a selected anti-CD371 antibody binds to a unique epitope, each antibody can be biotinylated using commercially available reagents (Pierce, Rockford, IL). Competition studies using unlabeled and biotinylated monoclonal antibodies can be performed using ELISA plates coated with CD371 as described above. Binding of biotinylated mAbs can be detected using a streptavidin-alkaline phosphatase probe.

[0181] To determine the isotype of purified antibodies, isotype ELISA can be performed using reagents specific for antibodies of a particular isotype. Anti-CD371 human IgG can be further tested for reactivity with the CD371 antigen by Western blotting.

[0182] In certain embodiments, K d is measured by a radiolabeled antigen binding assay (RIA). In certain embodiments, the RIA is performed using a Fab version of the antibody of interest and its antigen. For example, the Fab is bound to a minimal concentration of ( 125 I) The solution binding affinity of a Fab to an antigen is measured by equilibrating with labeled antigen and then capturing the bound antigen on an anti-Fab antibody-coated plate (see, e.g., Chen et al., J Mol Biol (1999);293:865-881). In certain embodiments, K dis measured using a BIACORE® surface plasmon resonance assay, e.g., an assay using a BIACORE®-2000 or BIACORE®-3000 (BIAcore, Inc., Piscataway, NJ). 5.3.7. Immunoconjugates

[0183] The presently disclosed subject matter provides an anti-CD371 antibody or antigen-binding fragment thereof conjugated to a therapeutic moiety, such as a cytotoxin, a drug (e.g., an immunosuppressant), or a radiotoxin. Such conjugates are referred to herein as "immunoconjugates." Immunoconjugates that include one or more cytotoxins are referred to as "immunotoxins." A cytotoxin or cytotoxic agent includes any agent that is detrimental to (e.g., kills) cells. Non-limiting examples of cytotoxins include taxol (such as ricin, diphtheria, and gelonin), cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxy anthracin dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, and puromycin, and analogs or homologs thereof. Therapeutic agents include, for example, calicheamicin, auristatins, antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracil), and the like. Also included are dacarbazine), alkylating agents (e.g., mechlorethamine, thioepa, chlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU), cyclothosphamide, busulfan, dibromomannitol, streptozotocin, mitomycin C, and cis-dichlorodiamineplatinum(II) (DDP) cisplatin), anthracyclines (e.g., daunorubicin (formerly daunomycin) and doxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin), bleomycin, mithramycin, and anthramycin (AMC)), hypomethylating agents (azacytidine and decitabine), and antimitotic agents (e.g., vincristine and vinblastine).

[0184] Other examples of therapeutic cytotoxins that can be conjugated to the anti-CD371 antibodies disclosed herein include duocarmycin, calicheamicin, maytansine, and auristatin, as well as derivatives thereof. Cytotoxins can be conjugated to the anti-CD371 antibodies or antigen-binding fragments thereof disclosed herein using linker technology available in the art. Examples of linker types that have been used to conjugate cytotoxins to antibodies include, but are not limited to, hydrazone, thioether, ester, disulfide, and peptide-containing linkers. For example, linkers can be selected that are susceptible to cleavage by the low pH in lysosomal compartments or by proteases preferentially expressed in tumor tissues, such as cathepsins (e.g., cathepsins B, C, and D). For further discussion of types of cytotoxins, linkers, and methods for conjugating therapeutic agents to antibodies, see also Saito, G. et al. (2003) Adv. Drug Deliv. Rev. 55:199-215; Trail, PA et al. (2003) Cancer Immunol. Immunother. 52:328-337; Payne, G. (2003) Cancer Cell 3:207-212; Allen, TM (2002) Nat. Rev. Cancer 2:750-763; Pastan, I. and Kreitman, RJ (2002) Curr. Opin. Investig. Drugs 3:1089-1091; Senter, PD and Springer, CJ (2001) Adv. Drug Deliv. Rev. 53:247-264.

[0185] The anti-CD371 antibodies or antigen-binding fragments thereof of the presently disclosed subject matter can also be conjugated to radioisotopes to produce cytotoxic radiopharmaceuticals, also referred to as radioimmunoconjugates. Non-limiting examples of radioisotopes that can be conjugated to antibodies for diagnostic or therapeutic use include: 90 Y, 131 I, 225 Ac, 213 Bi, 223 Ra and 227 The method for preparing radioimmunoconjugate is established in the art. Examples of radioimmunoconjugate include Zevalin (trademark) (IDEC Pharmaceuticals) and Bexxar (trademark) (Corixa Pharmaceuticals) are commercially available, and the antibody of the present invention can be used to prepare radioimmunoconjugate by using similar methods.

[0186] The antibody conjugates of the presently disclosed subject matter can be used to modify a given biological response, and the drug moiety should not be construed as limited to classical chemical therapeutic agents. For example, the drug moiety may be a protein or polypeptide possessing a desired biological activity. Such proteins may include, for example, enzymatically active toxins, such as abrin, ricin A, pseudomonas exotoxin, or diphtheria toxin, or active fragments thereof; proteins such as tumor necrosis factor (TNF) or interferon-γ; or biological response modifiers, such as lymphokines, interleukin-1 (IL-1), interleukin-2 (IL-2), interleukin-6 (IL-6), granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte-colony-stimulating factor (G-CSF), or other growth factors.

[0187] Techniques for conjugating such therapeutic moieties to antibodies are well known and are described, for example, in Arnon et al., "Monoclonal Antibodies For Immunotargeting Of Drugs In Cancer Therapy," in Monoclonal Antibodies And Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss, Inc. 1985); Hellstrom et al., "Antibodies For Drug Delivery," in Controlled Drug Delivery (2nd Ed.), Robinson et al. (eds.), pp. 623-53 (Marcel Dekker, Inc. 1987); Thorpe, "Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review," in Monoclonal Antibodies '84: Biological And Clinical Applications, Pinchera et al. (eds.), pp. 475-506 (1985); "Analysis, Results, And Future Prospective See, "Of The Therapeutic Use Of Radiolabeled Antibodies In Cancer Therapy," in Monoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al. (eds.), pp. 303-16 (Academic Press 1985), and Thorpe et al., "The Preparation And Cytotoxic Properties Of Antibody-Toxin Conjugates," Immunol. Rev., 62:119-58 (1982). 5.3.8. Bispecific molecules

[0188] The presently disclosed subject matter provides bispecific molecules comprising the anti-CD371 antibodies, or fragments thereof, disclosed herein. The antibodies, or antigen-binding fragments thereof, of the present disclosure can be derivatized or linked to another functional molecule, e.g., another peptide or protein (e.g., a ligand for another antibody or receptor), to generate bispecific molecules that bind to at least two different binding sites or target molecules. Indeed, the antibodies, or antigen-binding fragments thereof, of the present disclosure can be derivatized or linked to two or more other functional molecules to generate multispecific molecules that bind to three or more different binding sites and / or target molecules; such multispecific molecules are also intended to be encompassed by the term "bispecific molecule" as used herein. To generate bispecific molecules, the anti-CD371 antibodies, or antigen-binding fragments thereof, of the present disclosure can be functionally linked (e.g., by chemical coupling, genetic fusion, noncovalent association, or otherwise) to one or more other binding molecules, such as another antibody, antibody fragment, peptide, or binding mimetic, resulting in a bispecific molecule.

[0189] The presently disclosed subject matter provides bispecific molecules comprising at least a first binding specificity for CD371 and a second binding specificity for a second target epitope. The second target epitope may be a CD371 epitope or a non-CD371 epitope, e.g., a different antigen. In certain embodiments, the bispecific molecule is multispecific, and the molecule may further comprise a third binding specificity. For example, if the first portion of the bispecific antibody binds to an antigen on a tumor cell and the second portion of the bispecific antibody recognizes an antigen on the surface of a human immune effector cell, the antibody can specifically bind to the effector antigen on the human immune effector cell and thereby activate the effector cell. Thus, in certain embodiments, the bispecific antibody can form a link between an effector cell, e.g., a T cell, and a tumor cell, thereby enhancing effector function. In certain embodiments, the bispecific antibody of the present disclosure comprises at least a first binding specificity for CD371 and at least a second binding specificity for an immune cell.

[0190] The bispecific molecules of the presently disclosed subject matter can be prepared by conjugating the constituent binding specificities using methods known in the art. For example, each binding specificity of the bispecific molecule can be generated separately and then conjugated to each other. When the binding specificities are proteins or peptides, various coupling or cross-linking agents can be used for covalent conjugation. Non-limiting examples of cross-linking agents include protein A, carbodiimide, N-succinimidyl-S-acetyl-thioacetate (SATA), 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB), o-phenylenedimaleimide (oPDM), N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP), and sulfosuccinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate (sulfo-SMCC) (see, e.g., Karpovsky et al. (1984) J. Exp. Med. 160:1686; Liu, MA et al. (1985) Proc. Natl. Acad. Sci. USA 82:8648). Other methods include those described in Paulus (1985) Behring Ins. Mitt. No. 78, 118-132; Brennan et al. (1985) Science 229:81-83), and Glennie et al. (1987) J. Immunol. 139: 2367-2375. Conjugating agents can also be SATA and sulfo-SMCC, both of which are available from Pierce Chemical Co. (Rockford, IL).

[0191] When the binding specificity is an antibody, they can be conjugated via sulfhydryl bonds at the C-terminal hinge regions of the two heavy chains. In certain embodiments, the hinge region is modified to contain an odd number of sulfhydryl residues, preferably one, prior to conjugation.

[0192] Alternatively, both binding specificities can be encoded in the same vector and expressed and assembled in the same host cell, an approach that is particularly useful when the bispecific molecule is a mAb x mAb, mAb x Fab, Fab x F(ab')2, or ligand x Fab fusion protein.

[0193] The binding of bispecific molecules to their specific targets can be confirmed, for example, by enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), FACS analysis, bioassay (e.g., growth inhibition), or Western blot assay. Each of these assays generally detects the presence of a protein-antibody complex of particular interest by using a labeled reagent (e.g., an antibody) specific to the complex of interest. Alternatively, the complex can be detected using any of a variety of other immunoassays. For example, the antibody can be radiolabeled and used in a radioimmunoassay (RIA) (see, e.g., Weintraub, B., Principles of Radioimmunoassays, Seventh Training Course on Radioligand Assay Techniques, The Endocrine Society, March 1986, incorporated herein by reference). Radioisotopes can be detected by means such as the use of a gamma counter or scintillation counter, or by autoradiography. 5.3.9. SELECTION OF HIGH AFFINITY ScFv AGAINST CD371 POLYPEPTIDES

[0194] The next step is to select phage that bind with high affinity to the target antigen of interest (e.g., CD371) from non-binding or low-affinity binding phage in a phage display library (e.g., a human phage display library). This can be achieved by repeatedly binding phage to an antigen bound to a solid support, such as beads or mammalian cells, followed by removal of unbound phage and elution of specifically bound phage. In certain embodiments, the antigen (e.g., CD371) is immobilized on a surface (e.g., a polystyrene surface). The phage library is incubated with cells, beads, or other solid support, and unbound phage is removed by washing. Binding clones are selected and tested.

[0195] Once selected, positive scFv clones are tested for their binding to cell surface CD371 (human CD371) by flow cytometry. Briefly, phage clones are incubated with HEK293H cells overexpressing CD371. The cells are washed and then incubated with M13 coat protein mAb. The cells are washed again and labeled with PE-labeled anti-mouse Fab2 before flow cytometry.

[0196] In other embodiments, the anti-CD371 antibodies may contain amino acid substitutions in one or more framework regions designed to improve protein stability, antibody binding, expression levels, or to introduce sites for conjugation of therapeutic agents. These scFvs are then used to produce recombinant human monoclonal Igs according to methods known to those of skill in the art. 5.3.10. Engineering Full-Length mAbs Using Selected ScFv Fragments

[0197] Phage display technology allows for the rapid selection and production of antigen-specific scFv and Fab fragments that are useful in themselves or can be further developed to yield complete antibodies, antigen-binding proteins, or antigen-binding fragments thereof. Intact mAbs with Fc domains have several advantages over scFv and Fab antibodies. First, only full-length Abs exert immunological functions, such as Fc domain-mediated CDC and ADCC. Second, bivalent mAbs provide stronger antigen-binding affinity than monomeric Fab Abs. Third, Fabs and bivalent mAbs have different plasma half-lives and renal clearance rates. The specific characteristics and advantages of each can be tailored to the intended effector strategy. Fourth, bivalent mAbs may be internalized at different rates than scFvs and Fabs, thereby altering their immune or carrier functions. For example, alpha emitters do not need to be internalized to kill their targets, whereas many drugs and toxins benefit from the internalization of immune complexes. Thus, in one particular embodiment, once a scFv clone specific for CD371 was obtained from a phage display library, the scFv fragment was used to produce a full-length IgG mAb.

[0198] To produce recombinant human monoclonal IgG in Chinese hamster ovary (CHO) cells, full-length IgG mAbs can be engineered based on methods known to those skilled in the art (Tomomatsu et al., Production of human monoclonal antibodies against FceRIa by a method combining in vitro immunization with phage display. Biosci Biotechnol Biochem 73(7): 1465-1469 2009). Briefly, antibody variable regions can be subcloned into mammalian expression vectors, matching lambda or kappa light chain constant sequences and IgG1 subclass Fc (for example) (Lidija P, et al. An integrated vector system for the eukaryotic expression of antibodies or their fragments after selection from phage display libraries. Gene 1997; 187(1): 9-18; Lisa JH, et al. Crystallographic structure of an intact IgG1 monoclonal antibody. Journal of Molecular Biology 1998; 275(5): 861-872). Kinetic binding analysis (Yasmina NA, et al. Probing the binding mechanism and affinity of tanezumab, a recombinant humanized anti-NGF monoclonal antibody, using a repertoire of biosensors. Protein Science 2008; 17(8): 1326-1335) demonstrated K values ​​in the nanomolar range. d This can be used to confirm the specific binding of full-length IgG to CD371 at RT. 5.4. Nucleic Acids Encoding Antibodies or Antigen-Binding Fragments

[0199] The presently disclosed subject matter provides nucleic acids encoding the anti-CD371 antibodies or antigen-binding fragments thereof disclosed herein. In certain embodiments, the nucleic acids comprise or consist of the nucleotide sequence set forth in SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27.

[0200] Furthermore, a vector comprising the nucleic acid of the present disclosure is provided. In certain embodiments, the vector is an expression vector. The presently disclosed subject matter further provides a host cell comprising the expression vector disclosed herein. 5.5. Pharmaceutical Compositions and Methods of Treatment

[0201] The presently disclosed subject matter provides compositions comprising an anti-CD371 antibody or antigen-binding fragment thereof, an immunoconjugate, or a bispecific antibody of the present disclosure. In certain embodiments, the composition is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

[0202] The presently disclosed subject matter provides various methods using the anti-CD371 antibodies or antigen-binding fragments thereof, immunoconjugates, bispecific antibodies, and compositions disclosed herein. For example, the presently disclosed subject matter provides a method for reducing tumor burden in a subject. In certain embodiments, the method comprises administering to a subject one or more of the anti-CD371 antibodies or antigen-binding fragments thereof, immunoconjugates, bispecific antibodies, or compositions disclosed herein. The anti-CD371 antibodies or antigen-binding fragments thereof of the present disclosure reduce the number of tumor cells, reduce tumor size, and / or eradicate tumors in the subject.

[0203] The subject matter of the present disclosure provides a method for increasing or extending the survival time of a subject with a tumor or neoplasm.In certain embodiments, the method comprises administering to the subject one or more of the anti-CD371 antibodies or their antigen-binding fragments, immunoconjugates, bispecific antibodies, or compositions disclosed herein.The method can reduce or eradicate the tumor burden in the subject.

[0204] The presently disclosed subject matter further provides methods for treating and / or preventing a tumor or neoplasm in a subject. In certain embodiments, the method comprises administering to the subject one or more of the anti-CD371 antibodies or antigen-binding fragments thereof, immunoconjugates, bispecific antibodies, or compositions disclosed herein.

[0205] Such methods include administering an effective amount of an anti-CD371 antibody or antigen-binding fragment thereof of the present disclosure, a composition (e.g., a pharmaceutical composition) of the present disclosure to achieve the desired effect (alleviation of an existing condition or prevention of recurrence). For treatment, the amount administered is an amount effective to produce the desired effect. An effective amount can be given in one or a series of administrations. An effective amount can be given as a bolus or by continuous perfusion.

[0206] Non-limiting examples of neoplasms or tumors include acute myeloid leukemia (AML), multiple myeloma, chronic lymphocytic leukemia (CLL), lymphoma (Hodgkin's lymphoma, non-Hodgkin's lymphoma), glioblastoma, myelodysplastic syndrome (MDS), and chronic myeloid leukemia (CML), bone cancer, intestinal cancer, liver cancer, skin cancer, head and neck cancer, melanoma (cutaneous or intraocular malignant melanoma), kidney cancer (e.g., clear cell carcinoma), throat cancer, prostate cancer (e.g., hormonal refractory prostate adenocarcinoma), blood cancers (e.g., leukemia, lymphoma, and myeloma), cancer of the uterus, rectum, anal region, bladder, brain, stomach, testicle, fallopian tube, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, leukemia (e.g., acute leukemia, acute lymphocytic leukemia, acute myelocytic leukemia, acute myeloblastic leukemia, acute promyelocytic leukemia, acute monocytic leukemia, acute erythroleukemia, chronic leukemia, chronic myelocytic leukemia, polycythemia vera, cancer of the small intestine, cancer of the endocrine system, cancer of the thyroid gland, cancer of the parathyroid gland, cancer of the adrenal gland, sarcoma of soft tissue, cancer of the urethra, cancer of the penis, solid tumors of childhood, lymphocytic lymphoma, cancer of the bladder, cancer of the kidney or ureter, cancer of the renal pelvis, neoplasms of the central nervous system (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumors, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, including those induced by asbestos Environmentally induced cancers include Waldenstrom's hypergammaglobulinemia, heavy chain disease, and sarcomas and carcinomas (e.g., fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endothelial tumor, lymphangiosarcoma, lymphangioendothelial tumor, synovium, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, hepatoma, cholangiocarcinoma (nile carcinoma) solid tumors such as duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, cervical cancer, salivary gland cancer, uterine cancer, testicular cancer, bladder cancer, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, schwannoma, meningioma, melanoma, neuroblastoma, and retinoblastoma.

[0207] Non-limiting examples of suitable tumors or neoplasms include acute myeloid leukemia (AML), multiple myeloma, non-Hodgkin's lymphoma, Hodgkin's lymphoma, chronic lymphocytic leukemia (CLL), glioblastoma, myelodysplastic syndrome (MDS), and chronic myelogenous leukemia (CML). In certain embodiments, the tumor or neoplasm is AML.

[0208] The anti-CD371 antibody of the present disclosure can be administered using any suitable method or route, and an anti-neoplastic agent can be co-administered if necessary. Routes of administration include, but are not limited to, oral administration, intravenous administration, intraperitoneal administration, subcutaneous administration, intramuscular administration, intranodal administration, intratumoral administration, intraosseous administration, intrathecal administration, pleural administration, intrapleural administration, and direct administration. However, it should be emphasized that the subject matter of the present disclosure is not limited to any particular method or route of administration.

[0209] The anti-CD371 antibodies or antigen-binding fragments thereof of the present disclosure can be administered as conjugates that specifically bind to a receptor and deliver a toxic lethal payload after internalization of the ligand-toxin.

[0210] The anti-CD371 antibody or antigen-binding fragment thereof of the presently disclosed subject matter may be administered in the form of a composition further comprising a pharmaceutically acceptable carrier. Suitable pharmaceutically acceptable carriers include, for example, one or more of water, saline, phosphate-buffered saline, dextrose, glycerol, ethanol, etc., and combinations thereof. The pharmaceutically acceptable carrier may further contain minor amounts of auxiliary substances, such as wetting or emulsifying agents, preservatives, or buffers, which enhance the shelf life or effectiveness of the binding protein. Compositions for injection can be formulated to provide rapid, sustained, or delayed release of the active ingredient after administration to a mammal, as is well known in the art.

[0211] The presently disclosed subject matter also provides the use of antibodies and the nucleic acids encoding them for the treatment of tumors or neoplasms (e.g., AML), for diagnostic and prognostic applications, and as research tools for detecting CD371 in cells and tissues. Pharmaceutical compositions comprising the antibodies and nucleic acids of the present disclosure are encompassed by the presently disclosed subject matter. Vectors comprising the nucleic acids of the presently disclosed subject matter for antibody-based treatments via vectored immunotherapy are also contemplated by the presently disclosed subject matter. Vectors include expression vectors that enable the expression and secretion of antibodies, as well as vectors that direct the cell surface expression of antigen-binding proteins, such as chimeric antigen receptors.

[0212] Cells containing nucleic acids, for example cells transfected with a vector of the invention, are also encompassed by the subject matter of the invention. 5.6.Kit

[0213] The subject matter of the present disclosure provides a kit for the treatment and / or prevention of tumors or neoplasms (e.g., AML) to reduce tumor burden and / or increase or prolong the survival time of subjects with tumors or neoplasms (e.g., AML). In certain embodiments, the kit includes a composition comprising an anti-CD371 antibody or antigen-binding fragment thereof, immunoconjugate, bispecific antibody, or composition disclosed herein in unit dosage form. In certain embodiments, the kit includes a sterile container containing a therapeutic or prophylactic vaccine; such a container may be in the form of a box, an ampoule, a bottle, a vial, a tube, a bag, a sachet, a blister pack, or any other suitable container known in the art. Such a container may be made of plastic, glass, laminated paper, metal foil, or any other material suitable for holding a medicament.

[0214] In certain embodiments, the kit further comprises instructions for administering to a subject an anti-CD371 antibody or antigen-binding fragment thereof, immunoconjugate, bispecific antibody, or composition disclosed herein. The instructions may generally include information about the use of the anti-CD371 antibody or antigen-binding fragment thereof, immunoconjugate, bispecific antibody, and composition disclosed herein to treat and / or prevent a tumor or neoplasm (e.g., AML), reduce tumor burden, and / or increase or prolong survival of a subject with a tumor or neoplasm (e.g., AML). In certain embodiments, the instructions include at least one of the following: a description of the therapeutic agent; dosing schedules and administration for the treatment and / or prevention of a tumor or neoplasm (e.g., AML) or symptoms thereof; precautions; warnings; indications; contraindications; overdose information; adverse reactions; animal pharmacology; clinical studies; and / or references. The instructions may be printed directly on the container (if present), or may be printed as a label affixed to the container, or as a separate sheet, pamphlet, card, or folder supplied in or with the container. 5.7.Detection Method

[0215] The subject matter of the present disclosure is a whole cell or organization In certain embodiments, the method comprises: a) contacting a cell or tissue with an anti-CD371 antibody or antigen-binding fragment disclosed herein, wherein the antibody or antigen-binding fragment comprises a detectable label; b) determining the amount of labeled antibody or antigen-binding fragment thereof bound to the cells or tissue; and Includes.

[0216] In certain embodiments, b) comprises measuring the amount of detectable label associated with the cell or tissue, wherein the amount of bound antibody or antigen-binding fragment thereof indicates the amount of CD371 in the cell or tissue.

[0217] The cell or tissue may be any cell or tissue, including any normal, healthy, or cancerous cell or tissue. [Example]

[0218] 6. Working Example The following examples are presented so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the antibodies, bispecific antibodies, compositions comprising same, screening, and therapeutic methods of the presently disclosed subject matter, and are not intended to limit the scope of what the inventors regard as the presently disclosed subject matter. It will be understood that various other embodiments can be practiced given the general description provided above. Example 1 Generation of anti-CD371 antibodies and scFvs

[0219] The extracellular domain and the portion of CD371 corresponding to amino acids His 65-Ala 265 (UniProt accession number Q5QGZ9) were recombinantly produced as a soluble protein with a polyhistidine tag for purification. The extracellular domain of mouse CD371 (Thr 67-Arg 267) was also produced with a polyhistidine tag, and antibodies were screened for cross-species reactivity.

[0220] A proprietary naive, semi-synthetic scFv phage display library was screened for antibodies binding to the CD371 protein using standard solid-phase phage display panning techniques. Briefly, recombinant CD371 was immobilized on a polystyrene surface, which was then blocked with approximately 5% milk and incubated with the phage library. Subsequent washes, elution, and phage amplification steps were performed to complete each round of biopanning. Three rounds of panning were completed using the amplified CD371 binder-enriched phage pool from the previous round of panning as input for the next round. To identify clones that showed high specificity for CD371, single clones from the third round of panning were analyzed for binding to human CD371, mouse CD371, and BSA (as a nonspecific control) by enzyme-linked immunosorbent assay (ELISA) using anti-M13 phage antibodies. Only those monoclonal phage supernatants that showed specific binding to CD371 were selected for antibody sequencing, and six antibodies with unique sequences were identified: B10 (also referred to as "1B10"), C3 (also referred to as "1C3"), D6 (also referred to as "1D6"), A11 (also referred to as "2A11"), E4 (also referred to as "2E4"), and E8 (also referred to as "2E8"). None of the screened antibodies showed binding to either the human or mouse homolog of CD371.

[0221] To test whether the antibodies recovered from the phage panning campaign were able to bind to cell surface CD371 in its native conformation, monoclonal phage preparations were also screened by flow cytometry on CD371-transfected and wild-type HEK293H cells. Figure 1 shows the binding profiles of the 1B10, 1C3, 1D6, 2A11, 2E4, and 2E8 antibodies. Example 2 Antibody binding to cell lines expressing CD371

[0222] Based on preliminary in vitro functional characterization, two mAbs (1B10 and 1C3) were reformatted against human IgG1 and transfected into OCI cells (CD371 + AML cell lines). Both mAbs demonstrated dose-dependent binding, as shown in Figures 2A and 2B. B10 was administered in both orientations (V H -V L or V L -V H These scFv-Fc fusion constructs were further engineered into several formats, including scFv-Fc fusions with the variable domains of CD371. + Specific binding to cells was demonstrated (see Figure 3). Similarly, binding to cells was measured by V L -V H However, the V H -V L No binding of the oriented B10 scFv was observed by flow cytometry, suggesting lower affinity and thus the need for bivalent binding. Example 3 Antibody binding to recombinant CD371 in solution

[0223] Affinity measurements of B10 variants determined by biolayer interferometry by capturing IgG and Fc fusions with anti-Fc antibodies and using soluble CD371 as the analyte. For affinity measurements of scFv, biotinylated CD371 was captured with streptavidin and soluble scFv was used as the analyte. Table 8 shows the dissociation constants (K) for various antibody formats. D ), binding rate constant (k on ) and dissociation rate constant (k off ) are shown. Consistent with the flow cytometry results, full IgG and scFv-Fc fusions bound more strongly than scFv (Table 8), likely due to their bivalent interactions, resulting in an avidity effect. H -V LWeak binding of the oriented 1B10 scFv was observed, but the dissociation constant could not be calculated by any curve fit method. Table 8. Binding affinity of various formats of antibody B10 to soluble CD371 [Table 8] Embodiments of the subject matter of the present disclosure

[0224] From the foregoing description, it will be apparent that variations and modifications may be made to the subject matter of the present disclosure to adapt it for various uses and conditions. Such embodiments also fall within the scope of the following claims.

[0225] The recitation of a list of elements in any definition of a variable herein includes definitions of that variable as any single element or combination (or subcombination) of the listed elements. The recitation of an embodiment herein includes that embodiment as any single embodiment or in combination with any other embodiment or portion thereof.

[0226] All patents and publications mentioned in this specification are herein incorporated by reference to the same extent as if each individual patent or publication was specifically and individually indicated to be incorporated by reference. The present invention provides, for example, the following items. (Item 1) An anti-CD371 antibody or antigen-binding fragment thereof comprising a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11. (Item 2) An anti-CD371 antibody or antigen-binding fragment thereof comprising a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, or SEQ ID NO:12. (Item 3) (a) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11; and (b) a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, or SEQ ID NO:12. An anti-CD371 antibody or an antigen-binding fragment thereof comprising: (Item 4) An anti-CD371 antibody or antigen-binding fragment thereof comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region and the light chain variable region are: (a) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:1, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:2; (b) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:3, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:4; (c) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:5, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:6; (d) at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:7 a heavy chain variable region comprising an amino acid sequence, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:8; (e) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:10; and (f) a heavy chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:11, and a light chain variable region comprising an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO:12; An anti-CD371 antibody or antigen-binding fragment thereof selected from the group consisting of: (Item 5) An anti-CD371 antibody or antigen-binding fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, or SEQ ID NO: 11. (Item 6) An anti-CD371 antibody or antigen-binding fragment thereof comprising a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, or SEQ ID NO: 12. (Item 7) (a) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, or SEQ ID NO:11; and (b) a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, or SEQ ID NO: 12; An anti-CD371 antibody or an antigen-binding fragment thereof comprising: (Item 8) (a) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 1, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 2; (b) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 3, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 4; (c) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 5, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 6; (d) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 8; (e) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 9, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 10; and (f) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 11, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 12 8. The antibody or antigen-binding fragment thereof according to any one of items 1 to 7, comprising: (Item 9) an anti-CD371 antibody or an anti-CD371 antibody comprising a heavy chain variable region comprising CDR1, CDR2 and CDR3 domains; and a light chain variable region comprising CDR1, CDR2 and CDR3 domains; An antigen-binding fragment, comprising a heavy chain variable region CDR3 domain and a light chain variable region CDR3 domain: (a) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33 and conservative modifications thereof; (b) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 39 and conservative modifications thereof; (c) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45 and conservative modifications thereof; (d) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 51 and conservative modifications thereof; (e) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57 and conservative modifications thereof; and (f) a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60 and conservative modifications thereof; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 and conservative modifications thereof. an anti-CD371 antibody or antigen-binding fragment thereof selected from: (Item 10) a heavy chain variable region CDR2 domain and a light chain variable region CDR2 domain, (a) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 29 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 32 and conservative modifications thereof; (b) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38 and conservative modifications thereof; (c) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44 and conservative modifications thereof; (d) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 50 and conservative modifications thereof; (e) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56 and conservative modifications thereof; and (f) a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59 and conservative modifications thereof; and a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62 and conservative modifications thereof. 10. The antibody or antigen-binding fragment thereof according to item 9, selected from: (Item 11) a heavy chain variable region CDR1 domain and a light chain variable region CDR1 domain, (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31 and conservative modifications thereof; (b) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37 and conservative modifications thereof; (c) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43 and conservative modifications thereof; (d) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49 and conservative modifications thereof; (e) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55 and conservative modifications thereof; and (f) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58 and conservative modifications thereof; and a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61 and conservative modifications thereof. 11. The antibody or antigen-binding fragment thereof according to item 9 or 10, selected from: (Item 12) 12. The antibody or antigen-binding fragment thereof according to any one of items 9 to 11, wherein one or more of the CDR sequences have up to about five amino acid substitutions. (Item 13) 12. The antibody or antigen-binding fragment thereof according to any one of items 9 to 11, wherein one or more of the CDR sequences have up to about three amino acid substitutions. (Item 14) (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 29; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30; (b) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36; (c) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42; (d) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48; (e) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54; or (f) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59; and a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60; An anti-CD371 antibody or an antigen-binding fragment thereof comprising: (Item 15) (a) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 32; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33; (b) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 38; (c) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45; (d) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 51; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 52; (e) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57; or (f) a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 An anti-CD371 antibody or an antigen-binding fragment thereof comprising: (Item 16) (a) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 28; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 29; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 30; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 31; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 32; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 33; (b) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 34; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 35; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 36; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 37; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 38; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 39; (c) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 40; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 41; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 42; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 43; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 44; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 45; (d) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 46; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 47; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 48; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 49; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 50; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 51; (e) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 52; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 53; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 54; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 55; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 56; a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 57; or (f) a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 58; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 59; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 60; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 61; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 62; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 63 An anti-CD371 antibody or an antigen-binding fragment thereof comprising: (Item 17) 17. The antibody or antigen-binding fragment thereof according to any one of items 1 to 16, which binds to CD371 and comprises the amino acid sequence set forth in SEQ ID NO: 15 or a fragment thereof. (Item 18) 18. An antibody or antigen-binding fragment thereof that cross-competes for binding to CD371 with the antibody or antigen-binding fragment thereof according to any one of items 1 to 17. (Item 19) 18. An antibody or antigen-binding fragment thereof that binds to the same epitope on CD371 as the antibody or antigen-binding fragment thereof according to any one of items 1 to 17. (Item 20) The antibody sequence is in the light-heavy variable chain orientation (V L -V H 20. The antibody or antigen-binding fragment thereof according to any one of items 1 to 19, wherein the antibody or antigen-binding fragment thereof is present in a region corresponding to the region of interest. (Item 21) Approximately 1×10 -7 M to approximately 1 x 10 -8 Between m or approximately 1 x 10 -9 M to approximately 1 x 10 -8 The dissociation constant (K D 21. The antibody or antigen-binding fragment thereof according to any one of items 1 to 20, which binds to human CD371 at the .beta. (Item 22) An antibody or antigen-binding fragment thereof comprising the amino acid sequence set forth in SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20 or SEQ ID NO: 21. (Item 23) 23. The antibody or antigen-binding fragment thereof according to any one of items 1 to 22, wherein the antibody comprises a framework region of a human variable region. (Item 24) 24. The antibody or antigen-binding fragment thereof according to any one of items 1 to 23, which is fully human or an antigen-binding fragment thereof. (Item 25) 24. The antibody or antigen-binding fragment thereof according to any one of items 1 to 23, which is a chimeric antibody or antigen-binding fragment thereof. (Item 26) 24. The antibody or antigen-binding fragment thereof according to any one of items 1 to 23, which is a humanized antibody or antigen-binding fragment thereof. (Item 27) 27. The antibody or antigen-binding fragment thereof according to any one of items 1 to 26, wherein the antigen-binding fragment of the antibody is Fab, Fab', F(ab')2, a variable fragment (Fv), or a single-chain variable region (scFv). (Item 28) A composition comprising the antibody or antigen-binding fragment thereof according to any one of items 1 to 27. (Item 29) 29. The composition according to item 28, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier. (Item 30) 28. An immunoconjugate comprising the antibody or antigen-binding fragment thereof according to any one of items 1 to 27, linked to a therapeutic agent. (Item 31) 31. The immunoconjugate of item 30, wherein the therapeutic agent is a drug, a cytotoxin, or a radioisotope. (Item 32) 32. A composition comprising the immunoconjugate of item 30 or 31. (Item 33) 33. The composition according to item 32, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier. (Item 34) 28. A bispecific molecule comprising the antibody or antigen-binding fragment thereof according to any one of items 1 to 27, linked to a second functional moiety. (Item 35) 35. The bispecific molecule of item 34, wherein the second functional moiety has a different binding specificity than the antibody or antigen-binding fragment thereof. (Item 36) 36. A composition comprising the bispecific molecule of item 34 or 35. (Item 37) 37. The composition according to item 36, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier. (Item 38) A nucleic acid encoding the antibody or antigen-binding fragment thereof according to any one of Items 1 to 27. (Item 39) 39. An expression vector comprising the nucleic acid molecule of item 38. (Item 40) A host cell comprising the expression vector of item 39. (Item 41) Whole cells or organization A method for detecting CD371 in contacting a cell or tissue with the antibody or antigen-binding fragment thereof according to any one of items 1 to 27, wherein the antibody or antigen-binding fragment thereof comprises a detectable label; determining the amount of labeled antibody or antigen-binding fragment thereof bound to said cells or tissue by measuring the amount of detectable label associated with said cells or tissue, wherein the amount of bound antibody or antigen-binding fragment thereof indicates the amount of CD371 in said cells or tissue; A method comprising: (Item 42) 43. A method for reducing tumor burden in a subject, comprising administering to the subject the antibody or antigen-binding fragment thereof described in any one of Items 1 to 27, the immunoconjugate described in Item 30 or 31, the bispecific molecule described in Item 34 or 35, or the composition described in any one of Items 28, 29, 32, 33, 36, and 37. 43. The method of claim 42, wherein the number of tumor cells is reduced, the tumor size is reduced, and / or the tumor is eradicated in the subject. (Item 44) 36. A method for treating and / or preventing a tumor or neoplasm in a subject, comprising administering to the subject the antibody or antigen-binding fragment thereof according to any one of items 1 to 27, the immunoconjugate according to item 30 or 31, the bispecific molecule according to item 34 or 35, or the composition according to any one of items 28, 29, 32, 33, 36, and 37. (Item 45) 36. A method for increasing or prolonging the survival of a subject having a tumor or neoplasm, comprising administering to the subject the antibody or antigen-binding fragment thereof of any one of items 1 to 27, the immunoconjugate of item 30 or 31, the bispecific molecule of item 34 or 35, or the composition of any one of items 28, 29, 32, 33, 36, and 37. (Item 46) 46. ​​The method of claim 45, which can reduce or eradicate tumor burden in the subject. (Item 47) 47. The method of any one of items 42 to 46, wherein the tumor or neoplasm is selected from acute myeloid leukemia (AML), multiple myeloma, non-Hodgkin's lymphoma, Hodgkin's lymphoma, chronic lymphocytic leukemia (CLL), glioblastoma, myelodysplastic syndrome (MDS), and chronic myeloid leukemia (CML). (Item 48) 48. The method of any one of items 42 to 47, wherein the tumor or neoplasm is AML. (Item 49) 49. The method according to any one of items 42 to 48, wherein the subject is a human. (Item 50) 28. A kit for reducing tumor burden in a subject, treating and / or preventing a tumor or neoplasm in a subject, and / or increasing or prolonging survival of a subject having a tumor or neoplasm, comprising the antibody or antigen-binding fragment thereof according to any one of items 1 to 27. (Item 51) 51. The kit of item 50, further comprising written instructions for using the antibody or antigen-binding fragment thereof to treat and / or prevent a tumor or neoplasm in a subject and / or to increase or prolong survival of a subject having a tumor or neoplasm.

Claims

1. a heavy chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO:28; a heavy chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO:29; a heavy chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO:30; a light chain variable region CDR1 comprising the amino acid sequence set forth in SEQ ID NO:31; a light chain variable region CDR2 comprising the amino acid sequence set forth in SEQ ID NO:32; and a light chain variable region CDR3 comprising the amino acid sequence set forth in SEQ ID NO:33 An anti-CD371 antibody or antigen-binding fragment thereof comprising:

2. The antibody or antigen-binding fragment thereof according to claim 1, comprising a heavy chain variable region comprising an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% identical to the amino acid sequence set forth in SEQ ID NO:

1.

3. The antibody or antigen-binding fragment thereof according to claim 1, comprising a light chain variable region comprising an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% identical to the amino acid sequence set forth in SEQ ID NO:

2.

4. The antibody or antigen-binding fragment thereof according to claim 1, comprising a heavy chain variable region comprising an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% identical to the amino acid sequence set forth in SEQ ID NO: 1, and a light chain variable region comprising an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% identical to the amino acid sequence set forth in SEQ ID NO:

2.

5. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 4, comprising a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 1 and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

2.

6. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, which binds to CD371 and comprises the amino acid sequence set forth in SEQ ID NO: 15 or a fragment thereof.

7. 1 x 10 －７ M to 1 x 10 －８ Between M or 1 x 10 －９ M to 1 x 10 －８ The dissociation constant (K Ｄ The antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, which binds to human CD371 at the 5'-terminal end of the antibody or antigen-binding fragment thereof.

8. The antibody or antigen-binding fragment thereof according to claim 1, comprising the amino acid sequence set forth in SEQ ID NO:

68.

9. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, wherein the antibody comprises a framework region of a human variable region.

10. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 9, which is a chimeric antibody or antigen-binding fragment thereof.

11. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 9, which is a humanized antibody or antigen-binding fragment thereof.

12. The antigen-binding fragment of the antibody may be Fab, Fab', F(ab') ２ 12. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 11, which is a variable fragment (Fv), or a single-chain variable region (scFv).

13. A composition comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 12.

14. 14. The composition of claim 13, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

15. An immunoconjugate comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 12 linked to a therapeutic agent.

16. The immunoconjugate of claim 15 , wherein the therapeutic agent is a drug, a cytotoxin, or a radioisotope.

17. A composition comprising the immunoconjugate of claim 15 or 16.

18. 18. The composition of claim 17, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

19. A bispecific molecule comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 12 linked to a second functional moiety.

20. 20. The bispecific molecule of claim 19, wherein the second functional moiety has a different binding specificity than the antibody or antigen-binding fragment thereof.

21. 21. A composition comprising the bispecific molecule of claim 19 or 20.

22. 22. The composition of claim 21, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

23. A nucleic acid encoding the antibody or antigen-binding fragment thereof according to any one of claims 1 to 12.

24. 24. An expression vector comprising the nucleic acid molecule of claim 23.

25. A host cell comprising the expression vector of claim 24.

26. 13. A composition for use in a method for detecting CD371 in whole cells or tissues, comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 12, wherein the antibody or antigen-binding fragment thereof comprises a detectable label; The method comprises: contacting a cell or tissue with the composition; determining the amount of labeled antibody or antigen-binding fragment thereof bound to said cells or tissue by measuring the amount of detectable label associated with said cells or tissue, wherein the amount of bound antibody or antigen-binding fragment thereof indicates the amount of CD371 in said cells or tissue; A composition comprising:

27. 23. The composition of any one of claims 13, 14, 17, 18, 21, and 22, for use in a method of reducing tumor burden in a subject, the method comprising administering the composition to the subject.

28. 28. The composition of claim 27, wherein the method reduces the number of tumor cells, reduces tumor size, and / or eradicates a tumor in the subject.

29. 23. The composition of any one of claims 13, 14, 17, 18, 21, and 22, for use in a method for treating and / or preventing a tumor or neoplasm in a subject, the method comprising administering the composition to the subject.

30. 23. The composition of any one of claims 13, 14, 17, 18, 21, and 22, for use in a method for increasing or extending the survival time of a subject having a tumor or neoplasm, the method comprising administering the composition to the subject.

31. 31. The composition of claim 30, which is capable of reducing or eradicating tumor burden in the subject.

32. 32. The composition of any one of claims 27 to 31, wherein the tumor or neoplasm is selected from acute myeloid leukemia (AML), multiple myeloma, non-Hodgkin's lymphoma, Hodgkin's lymphoma, chronic lymphocytic leukemia (CLL), glioblastoma, myelodysplastic syndrome (MDS), and chronic myelogenous leukemia (CML).

33. The composition of any one of claims 27 to 32, wherein the tumor or neoplasm is AML.

34. The composition of any one of claims 27 to 33, wherein the subject is a human.

35. 13. A kit for reducing tumor burden in a subject, for treating and / or preventing a tumor or neoplasm in a subject, and / or for increasing or extending the survival of a subject having a tumor or neoplasm, comprising the antibody or antigen-binding fragment thereof described in any one of claims 1 to 12.

36. 36. The kit of claim 35, further comprising written instructions for using the antibody or antigen-binding fragment thereof to treat and / or prevent a tumor or neoplasm in a subject and / or to increase or prolong the survival of a subject having a tumor or neoplasm.

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