Pharmaceutical composition for treating and / or preventing cancer

Abstract

Provided is a conjugate obtained by binding a thiol-containing maytansinoid and an antibody or a fragment thereof having immunological reactivity with an amino acid sequence represented by any one of even SEQ ID NOs: 2 to 30 or a CAPRIN -1 protein having an amino acid sequence having 80% or more sequence identity with the antibody. The conjugate can be used as an antibody-drug complex (ADC) for treating and / or preventing cancer.

Description

Title of Invention: PHARMACEUTICAL COMPOSITION FOR TREATING AND / OR PREVENTING CANCER Technical Field

[0001] The present invention relates to a conjugate of an antibody against a CAPRIN-1 protein or a fragment thereof and a thiol-containing maytansinoid, and medical use thereof as an agent for treatment and / or prevention of a cancer. Background Art

[0002] Various antibody drugs targeting specific antigenic proteins on cancer cells are applied as therapeutic drugs for cancers with fewer adverse reactions to cancer treatment because of their cancer specificity. For example, cytoplasmic-activation and proliferation-associated protein 1 (CAPRIN-1) is expressed on the cell membrane surface of many solid cancers, and antibodies against this CAPRIN-1 protein have been known to be promising in medical use for the treatment and / or prevention of cancers (Patent Literature 1).

[0003] In recent years, studies have been made to enhance the pharmacological effects of the antibody drugs against cancers. Particularly, efforts have been directed toward the development of antibody-drug-conjugates (ADCs), in which an antitumor drug is conjugated to an antibody. A practical example of an ADC using, as a drug, a thiol-containing maytansinoid, that is, an antitumor compound that inhibits microtubule polymerization and thereby inhibits mitosis, is ELAHERE (registered trademark) (mirvetuximab soravtansine-gynx), in which DM4, a thiol-containing maytansinoid, is conjugated to mirvetuximab, that is, an antibody against folic acid receptor-a (FRa receptor). ELAHERE has demonstrated an objective response rate (ORR) of 31.7% and a median duration of response (DOR) of 6.9 months in a single-arm study in patients with FRa-positive, platinum-resistant epithelial ovarian cancer, fallopian tube cancer, or primary peritoneal cancer with prior bevacizumab treatment, and is currently used in certain treatments for such cancer patients (Non Patent Literature 1). In addition, with respect to KADCYLA (registered trademark) (ado-trastuzumab emtansine (TDM1)), in which DM1, a thiol-containing maytansinoid, is conjugated to trastuzumab, that is, an antibody against HER2, it has been shown that, in a comparison between a combination therapy with lapatinib and capecitabine and TDM1 monotherapy in patients with HER2-positive advanced breast cancer previously treated with a taxane-based anticancer agent, the median progression-free survival was 9.6 months for TDM1, whereas it was 6.4 months for the combination therapy with lapatinib and capecitabine, and the median overall survival was 30.9 months for TDM1, whereas it was 25.1 months for the combination therapy with lapatinib and capecitabine (Non Patent Literature 2). Citation List Patent Literature

[0004] Patent Literature 1: WO 2010 / 016526 Non Patent Literature

[0005] Non Patent Literature 1: HIGHLIGHTS OF PRESCRIBING INFORMATION (Reference ID: 5077370) (https: / / www.accessdata.fda.gov / drugsatfda_docs / label / 2022 / 761310s000lbl.pdf) Non Patent Literature 2: The New England Journal of Medicine 2012 Nov 8;367(19):1783-91 Summary of Invention Technical Problem

[0006] As described above, although ADCs employing thiol-containing maytansinoids for the purpose of enhancing the pharmacological effects of antibody drugs against cancers have already been put into practical use, the successful cases have been limited to those directed to only some cancers, and their antitumor effects have also been limited. Furthermore, as shown in the Examples of the present specification, a conjugate of trastuzumab, which is marketed as an antibody drug for cancers, and a thiol-containing maytansinoid exhibited only a slight enhancement of antitumor activity compared with trastuzumab alone. In the present invention, it has been newly found that ADC technology employing a thiol-containing maytansinoid is not a technology that markedly enhances the antitumor effects of all antibody drugs against cancers.

[0007] Accordingly, an object of the present invention is to create an ADC employing a thiol-containing maytansinoid that exhibits an excellent antitumor effect. Solution to Problem

[0008] The present inventor has conducted diligent studies and consequently completed the present invention by finding that: a conjugate of an antibody or a fragment thereof against a CAPRIN-1 protein and a thiol-containing maytansinoid exerts a much stronger antitumor effect than that of the antibody against the CAPRIN-1 protein or the fragment thereof used alone; and the effect of enhancing the antitumor effect by conjugating the antibody against the CAPRIN-1 protein or the fragment thereof to the thiol-containing maytansinoid is much superior to the effect of enhancing the antitumor effect by conjugating an existing antibody drug for a cancer to the thiol-containing maytansinoid.

[0009] Specifically, the present invention has the following features (1) to (12).

[0010] (1) A conjugate of an antibody or a fragment thereof linked to a thiol-containing maytansinoid, wherein the antibody or the fragment thereof has immunological reactivity with a CAPRIN-1 protein having an amino acid sequence represented by any of even-numbered SEQ ID NOs among SEQ ID NOs: 2 to 30 or an amino acid sequence having 80% or more sequence identity to the amino acid sequence. (2) The conjugate according to (1), wherein the antibody or the fragment thereof has immunological reactivity with a partial polypeptide of the CAPRIN-1 protein, wherein the partial polypeptide has an amino acid sequence represented by any of SEQ ID NOs: 31 to 35 and 296 to 299, 308, and 309 or an amino acid sequence having 80% or more sequence identity to the amino acid sequence.

[0012] (3) The conjugate according to (1) or (2), wherein the antibody is a monoclonal antibody or a polyclonal antibody.

[0013] (4) The conjugate according to any of (1) to (3), wherein the antibody or the fragment thereof is any of the following (A) to (M): (A) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 36, 37, and 38 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 40, 41, and 42 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (B) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 44, 45, and 46 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 48, 49, and 50 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (C) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 52, 53, and 54 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 56, 57, and 58 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (D) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 60, 61, and 62 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 64, 65, and 66 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (E) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 170, 171, and 172 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 173, 174, and 175 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (F) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 176, 177, and 178 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 179, 180, and 181 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (G) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 182, 183, and 184 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 185, 186, and 187 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (H) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 188, 189, and 190 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 191, 192, and 193 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (I) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 146, 147, and 148 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 149, 150, and 151 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (J) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 272, 273, and 274 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 275, 276, and 277 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (K) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 290, 291, and 292 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 293, 294, and 295 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, (L) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 301, 302, and 303 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 305, 306, and 307 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, and (M) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 134, 135, and 136 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 137, 138, and 139 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein.

[0014] (5) The conjugate according to any of (1) to (4), wherein the antibody or the fragment thereof is any of the following (a) to (al): (a) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 39 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 43, (b) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 51, (c) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 55 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 59, (d) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 63 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 67, (e) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 68 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 69, (f) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 70 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 71, (g) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 72 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 73, (h) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 74 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 75, (i) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 76 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 77, (j) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 78 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 79, (k) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 80 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 81, (l) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 82 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 83, (m) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 84 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 85, (n) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 86 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 87, (o) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 88 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 89, (p) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 90 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 91, (q) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 92 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 93, (r) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 94 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 95, (s) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 96 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 97, (t) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 98 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 99, (u) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 100 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 101, (v) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 102 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 103, (w) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 104 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 105, (x) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 106 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 107, (y) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 108 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 109, (z) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 110 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 111, (aa) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 112 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 113, (ab) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 114 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 115, (ac) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 116 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 117, (ad) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 118 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 119, (ae) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 120 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 121, (af) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 122 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 123, (ag) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 124 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 125, (ah) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 126 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 127, (ai) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 128 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 129, (aj) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 130 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 131, (ak) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 132 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 133, and (al) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 300 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 304.

[0015] (6) The conjugate according to any of (1) to (5), wherein the antibody is a human antibody, a humanized antibody, a chimeric antibody, or a single-chain antibody.

[0016] (7) The conjugate according to any of (1) to (6), wherein the antibody or the fragment thereof is linked to the thiol-containing maytansinoid via a linker.

[0017] (8) The conjugate according to any of (1) to (7), wherein the thiol-containing maytansinoid is DM1, DM4, or a derivative thereof. (9) A pharmaceutical composition for treatment and / or prevention of a cancer, comprising the conjugate according to any of (1) to (8) as an active ingredient.

[0019] (10) The pharmaceutical composition according to (9), wherein the cancer is a cancer expressing a CAPRIN-1 protein on the cell membrane surface.

[0020] (11) The pharmaceutical composition according to (9) or (10), wherein the cancer is breast cancer, kidney cancer, pancreatic cancer, colorectal cancer, lung cancer, brain tumor, gastric cancer, uterine cancer, ovarian cancer, prostate cancer, bladder cancer, esophageal cancer, leukemia, lymphoma, liver cancer, gallbladder cancer, bile duct cancer, sarcoma, mastocytoma, melanoma, adrenocortical carcinoma, Ewing's tumor, Hodgkin's lymphoma, mesothelioma, multiple myeloma, testicular cancer, thyroid cancer, head and neck cancer, or urothelial carcinoma.

[0021] (12) A method for treating and / or preventing a cancer, comprising administering the conjugate according to any of (1) to (8), or the pharmaceutical composition according to any of (9) to (11) to a subject. Advantageous Effects of Invention

[0022] The conjugate according to the present invention not only exerts a much stronger antitumor effect than that of an antibody or a fragment thereof against a CAPRIN-1 protein used alone but also is superior in antitumor effect to a known conjugate of an antibody drug for a cancer and a thiol-containing maytansinoid. In addition, the effect of enhancing the antitumor effect by conjugating the antibody against the CAPRIN-1 protein or the fragment thereof and the thiol-containing maytansinoid is much superior to the effect of enhancing the antitumor effect by conjugating an existing antibody drug for a cancer and the thiol-containing maytansinoid. Thus, the conjugate according to the present invention is effective for the treatment or prevention of a cancer. Description of Embodiments

[0023] The antitumor activity of the conjugate of an antibody or a fragment thereof against a CAPRIN-1 protein (hereinafter, referred to as "anti-CAPRIN-1 antibody") and a thiol-containing maytansinoid of the present invention can be evaluated, as described below, by examining the inhibition of tumor growth in vitro or in vivo (specifically in a tumor-bearing animal).

[0024] In the present invention, the "conjugate" refers to an antibody or a fragment thereof linked to a drug via a covalent bond. The antibody and the drug may be linked via a linker.

[0025] The anti-CAPRIN-1 antibody that is a constituent of the conjugate according to the present invention may be a monoclonal antibody or a polyclonal antibody but is preferably a monoclonal antibody. The anti-CAPRIN-1 antibody may also be any type of antibody as long as the conjugate of the present invention can exert antitumor activity, and may be a recombinant antibody, a human antibody, a humanized antibody, a chimeric antibody, or a non-human animal antibody.

[0026] The thiol-containing maytansinoid that forms the conjugate of the present invention is one of maytansine derivatives that target microtubules and inhibit cell proliferation by inhibiting mitosis.

[0027] The subject who is the target to be treated and / or prevented for cancer according to the present invention is a mammal such as a human, a pet animal, livestock, or a sport animal, and a preferred subject is a human.

[0028] Hereinafter, the anti-CAPRIN-1 antibody, the thiol-containing maytansinoid, the conjugate of the anti-CAPRIN-1 antibody and the thiol-containing maytansinoid, the pharmaceutical composition comprising the conjugate, and the method for treating and / or preventing a cancer using the conjugate, according to the present invention will be described.

[0029] <Anti-CAPRIN-1 antibody> Among CAPRIN-1 proteins having an amino acid sequence represented by any of even-numbered SEQ ID NOs among SEQ ID NOs: 2 to 30 and having immunological reactivity with the anti-CAPRIN-1 antibody used in the present invention, the amino acid sequences represented by SEQ ID NOs: 6, 8, 10, 12, and 14 are the amino acid sequences of canine CAPRIN-1 proteins; the amino acid sequence represented by SEQ ID NOs: 2 and 4 are the amino acid sequences of human CAPRIN-1 proteins; the amino acid sequence represented by SEQ ID NO: 16 is the amino acid sequence of a bovine CAPRIN-1 protein; the amino acid sequence represented by SEQ ID NO: 18 is the amino acid sequence of an equine CAPRIN-1 protein; the amino acid sequences represented by SEQ ID NOs: 20 to 28 are the amino acid sequences of mouse CAPRIN-1 proteins; and the amino acid sequence represented by SEQ ID NO: 30 is the amino acid sequence of a chicken CAPRIN-1 protein.

[0030] The anti-CAPRIN-1 antibody used in the present invention means an antibody or an antigen-binding fragment thereof that has immunological reactivity with the full-length CAPRIN-1 protein or with a fragment thereof as an antigen. Here, "immunological reactivity" means a property whereby the antibody specifically binds, in vivo, to the CAPRIN-1 protein or a partial polypeptide thereof. Specific examples include those that have immunological reactivity with a variant of the CAPRIN-1 protein having 80% or more, preferably 90% or more, more preferably 95% or more, and even more preferably 99% or more sequence identity to the amino acid sequence represented by any of even-numbered SEQ ID NOs among SEQ ID NOs: 2 to 30. In this context, the term "% sequence identity" means a percentage (%) of the number of identical amino acids (or bases) to the total number of amino acids (or bases) when two sequences are aligned so that the maximum degree of similarity can be achieved with or without introducing a gap.

[0031] The anti-CAPRIN-1 antibody used in the present invention may be a monoclonal antibody or a polyclonal antibody.

[0032] The polyclonal antibody having immunological reactivity with the full-length CAPRIN-1 protein or the fragment thereof (anti-CAPRIN-1 polyclonal antibody) can be obtained, for example, by immunizing a mouse, a human antibody-producing mouse, a rat, a rabbit, a chicken, or the like with a naturally occurring CAPRIN-1 protein, a fusion protein thereof with GST or the like, or a partial peptide thereof and obtaining serum therefrom. The obtained serum is applied to ammonium sulfate precipitation, protein A, protein G, a DEAE ion-exchange column, an affinity column linked to a CAPRIN-1 protein or a partial peptide, or the like, and thus the antibody can be obtained.

[0033] As for the full-length CAPRIN-1 protein or the fragment thereof to be used in the immunization, the base sequences and amino acid sequences of CAPRIN-1 and homologs thereof are available, for example, by accessing GenBank (NCBI, USA) and using an algorithm such as BLAST or FASTA (Karlin and Altschul, Proc. Natl. Acad. Sci. USA, 90: 5873-5877, 1993; and Altschul et al., Nucleic Acids Res. 25: 3389-3402, 1997). In addition, a method for preparing the CAPRIN-1 protein is available with reference to WO 2014 / 012479, or can be carried out using, for example, cells expressing the CAPRIN-1 protein.

[0034] The monoclonal antibody having immunological reactivity with the full-length CAPRIN-1 protein or the fragment thereof (anti-CAPRIN-1 monoclonal antibody) can be obtained, for example, by: administering SK-BR-3 (breast cancer cells expressing CAPRIN-1 protein) or the full-length CAPRIN-1 protein or the fragment thereof, or the like to a mouse for immunization; fusing spleen cells separated from the mouse with myeloma cells; and selecting a clone producing anti-CAPRIN-1 monoclonal antibodies from among the obtained fusion cells (hybridomas). The antibody produced from the hybridoma thus selected can be prepared in the same way as the method for purifying the polyclonal antibody mentioned above.

[0035] The antibody used in the present invention includes human antibodies, humanized antibodies, chimeric antibodies, and non-human animal antibodies.

[0036] The human antibody can be obtained by: sensitizing EB virus-infected human lymphocytes with the protein, protein-expressing cells, or lysates thereof; fusing the sensitized lymphocytes with human-derived myeloma cells such as U266 cells; and obtaining an antibody having immunological reactivity with the full-length CAPRIN-1 protein or the fragment thereof from the obtained fusion cells.

[0037] The humanized antibody, also called reshaped human antibody, is an engineered antibody. The humanized antibody is constructed by grafting complementarity-determining regions of an antibody derived from an immunized animal onto complementarity-determining regions of a human antibody. A genetic engineering technique commonly used for constructing humanized antibodies is also well-known. Specifically, for example, DNA sequences designed to link complementarity-determining regions of a mouse or rabbit antibody, to framework regions of a human antibody are synthesized by PCR from several prepared oligonucleotides having overlapping terminal portions. The obtained DNAs are ligated with DNAs encoding human antibody constant regions. The resulting ligation products are incorporated into expression vectors, which are then transferred to hosts for antibody production to obtain the antibody of interest (see, EP 239400 and WO 96 / 02576). The framework regions of a human antibody connected via the complementarity-determining regions are selected so that the complementarity-determining regions form a favorable antigen-binding site. If necessary, an amino acid in the framework regions of antibody variable regions may be substituted so that the complementarity-determining regions of a reshaped human antibody form an appropriate antigen-binding site (Sato K. et al., Cancer Research 1993, 53: 851-856). In addition, these framework regions may be replaced with framework regions derived from various human antibodies (see WO 99 / 51743).

[0038] Antibodies are typically heteromultimeric glycoproteins each comprising at least two heavy chains and two light chains. The antibodies are each composed of two identical light chains and two identical heavy chains. Each heavy chain has a heavy chain variable region at one end, followed by a series of constant regions. Each light chain has a light chain variable region at one end, followed by a series of constant regions. The variable regions contain certain variable regions called complementarity-determining regions (CDRs) and impart binding specificity to the antibody. Portions relatively conserved in the variable regions are called framework regions (FRs). The complete heavy chain and light chain variable regions each contain four FRs connected via three CDRs (CDR1 to CDR3).

[0039] The sequences of human-derived heavy chain and light chain constant regions and variable regions are available from NCBI (USA; GenBank, UniGene, etc.). For example, the following sequences may be referred to: Accession No. J00228 for a human IgG1 heavy chain constant region; Accession No. J00230 for a human IgG2 heavy chain constant region; Accession Nos. V00557, X64135, X64133, etc., for a human light chain k constant region; and Accession Nos. X64132, X64134, etc., for a human light chain X constant region.

[0040] A chimeric antibody is an antibody prepared from a combination of sequences derived from different animals and may be, for example, an antibody composed of heavy chain variable region and light chain variable region of a mouse antibody and constant regions of heavy chain and light chain variable regions of a human antibody. The chimeric antibody can be prepared using a method known in the art and is obtained, for example, by: ligating DNAs encoding the antibody V regions to DNAs encoding the human antibody C regions; incorporating the resulting ligation products into expression vectors; and transferring the vectors into hosts for antibody production.

[0041] The non-human animal antibody is obtained by immunizing an animal with a sensitizing antigen according to a method known in the art and, as a general method, by intraperitoneally, intracutaneously, or subcutaneously injecting a sensitizing antigen to an animal such as a mouse. For the injection of the sensitizing antigen, the antigen is mixed in an appropriate amount with various adjuvants including CFA (complete Freund's adjuvant), and the mixture is administered to the animal a plurality of times. The animal is immunized and then verified to contain anti-CAPRIN-1 antibodies in serum. The serum can be obtained and applied, as mentioned above, to ammonium sulfate precipitation, protein A, protein G, a DEAE ion-exchange column, an affinity column bound with a CAPRIN-1 protein or a partial peptide, or the like to obtain the non-human animal antibody. In the case of obtaining a monoclonal antibody from a nonhuman animal, immune cells can be collected from an immunized animal and subjected to cell fusion with myeloma cells to obtain the monoclonal antibody. The cell fusion of the immune cells with the myeloma cells may be carried out according to a method known in the art (see Kohler, G. and Milstein, C. Methods Enzymol. (1981) 73, 3-46).

[0042] The antibody used in the present invention may also be obtained as a recombinant antibody produced using a genetic engineering technique by cloning genes of the antibody from a hybridoma; incorporating the antibody genes into appropriate vectors; and transferring the vectors into hosts (see Carl, A.K., Borrebaeck, James, W. Larrick, THERAPEUTIC MONOCLONAL ANTIBODIES, Published in the United Kingdom by MACMILLAN PUBLISHERS LTD, 1990).

[0043] The anti-CAPRIN-1 antibody used in the present invention may be an antibody in which an amino acid in a variable region (e.g., FR) or a constant region is substituted with another amino acid. The amino acid substitution is the substitution of one or more, for example, less than 15, less than 10, 8 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less amino acids, preferably 1 to 9 amino acids. The substituted antibody is preferably an antibody that has the property of specifically binding to the antigen and binding affinity for the antigen equivalent to or higher than those of the corresponding unsubstituted antibody and causes no rejection when applied to humans.

[0044] The anti-CAPRIN-1 antibody used in the present invention is expected to have a stronger antitumor effect when it has the higher binding affinity for the CAPRIN-1 protein on cancer cell surface. Its association constant (affinity constant) Ka (kon / koff) is preferably at least 107 M-1, at least 108 M-1, at least 5 x 108 M-1, at least 109 M-1, at least 5 x 109 M-1, at least 1010 M-1, at least 5 x 1010 M-1, at least 1011 M-1, at least 5 x 1011 M-1, at least 1012 M-1, or at least 1013 M-1.

[0045] The binding activity of the anti-CAPRIN-1 antibody used in the present invention against effector cells may be improved by substituting 1, 2 or several amino acids in the heavy chain constant region of the antibody or by removing fucose linked to N-acetylglucosamine in a N-glycoside-linked sugar chain attached to the heavy chain constant region. Such an antibody may only have the amino acid substitution or may be in a composition comprising a fucosylated antibody.

[0046] The antibody in which 1, 2 or several amino acids in the heavy chain constant region are substituted may be prepared with reference to, for example, WO 2004 / 063351, WO 2011 / 120135, U.S. Patent No. 8388955, WO 2011 / 005481, U.S. Patent No. 6737056, and / or WO 2005 / 063351.

[0047] The antibody lacking fucose added to N-acetylglucosamine in a N-glycoside-linked sugar chain in the heavy chain constant region has been removed, or cells producing the antibody, may be prepared with reference to U.S. Patent No. 6602684, European Patent No. 1914244, and / or U.S. Patent No. 7579170. A composition of the antibody lacking fucose added to N-acetylglucosamine in a N-glycoside-linked sugar chain attached to the heavy chain constant region, and the antibody having the fucose, or cells producing the composition may be prepared with reference to, for example, U.S. Patent No. 8642292.

[0048] Methods for preparing and purifying the anti-CAPRIN-1 polyclonal antibody, the anti-CAPRIN-1 monoclonal antibody, and the antibody used in the present invention, and a method for preparing the CAPRIN-1 protein or the partial polypeptide thereof to be used in immunization may be carried out with reference to WO 2010 / 016526, WO 2011 / 096517, WO 2011 / 096528, WO 2011 / 096519, WO 2011 / 096533, WO 2011 / 096534, WO 2011 / 096535, WO 2013 / 018886, WO 2013 / 018894, WO 2013 / 018892, WO 2013 / 018891, WO 2013 / 018889, WO 2013 / 018883, WO 2013 / 125636, WO 2013 / 125654, WO 2013 / 125630, WO 2013 / 125640, WO 2013 / 147169, WO 2013 / 147176 and WO 2015 / 020212.

[0049] Specific examples of the anti-CAPRIN-1 antibody used in the present invention include anti-CAPRIN-1 antibodies disclosed in WO 2010 / 016526, WO 2011 / 096517, WO 2011 / 096528, WO 2011 / 096519, WO 2011 / 096533, WO 2011 / 096534, WO 2011 / 096535, WO 2013 / 018886, WO 2013 / 018894, WO 2013 / 018892, WO 2013 / 018891, WO 2013 / 018889, WO 2013 / 018883, WO 2013 / 125636, WO 2013 / 125654, WO 2013 / 125630, WO 2013 / 125640, WO 2013 / 147169, WO 2013 / 147176 and WO 2015 / 020212 mentioned above. Preferred examples of the anti-CAPRIN-1 antibody include the following.

[0050] An antibody or a fragment thereof having immunological reactivity with the amino acid sequence represented by SEQ ID NO: 2 or SEQ ID NO: 4 or a partial polypeptide of the CAPRIN-1 protein having an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more, still further preferably 99% or more) sequence identity to the amino acid sequence represented by SEQ ID NO: 2 or SEQ ID NO: 4.

[0051] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 31 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 36, 37, and 38 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 40, 41, and 42 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein; an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 140, 141, and 142 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 143, 144, and 145 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein; or an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 164, 165, and 166 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 167, 168, and 169 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 39 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 43; an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 70 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 71; or an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 78 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 79.

[0052] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 33 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 60, 61, and 62 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 64, 65, and 66 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein; more preferably an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 63 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 67.

[0053] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 32 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 52, 53, and 54 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 56, 57, and 58 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 55 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 59.

[0054] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 34 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 170, 171, and 172 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 173, 174, and 175 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein; or an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 176, 177, and 178 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 179, 180, and 181 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 80 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 81; or an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 82 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 83.

[0055] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 35 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 182, 183, and 184 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 185, 186, and 187 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein; or an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 188, 189, and 190 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 191, 192, and 193 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 84 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 85; or an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 86 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 87.

[0056] An antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 44, 45, and 46 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementaritydetermining regions of SEQ ID NOs: 48, 49, and 50 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. Preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 51.

[0057] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 296 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 146, 147, and 148 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 149, 150, and 151 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 72 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 73.

[0058] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 297 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 272, 273, and 274 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 275, 276, and 277 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 114 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 115.

[0059] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 298 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 290, 291, and 292 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 293, 294, and 295 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 120 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 121.

[0060] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 299 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 301, 302, and 303 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 305, 306, and 307 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 300 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 304.

[0061] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 308 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 134, 135, and 136 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 137, 138, and 139 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 68 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 69.

[0062] An antibody or a fragment thereof having immunological reactivity with a partial polypeptide of the CAPRIN-1 protein having the amino acid sequence represented by SEQ ID NO: 309 or an amino acid sequence having 80% or more (preferably 85% or more, more preferably 90% or more, further preferably 95% or more) sequence identity to the amino acid sequence. Preferably, an antibody or a fragment thereof which comprises a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 134, 135, and 136 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 137, 138, and 139 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein. More preferably, an antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 68 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 69.

[0063] Additionally, the following anti-CAPRIN-1 antibodies are preferably used.

[0064] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 68 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 69.

[0065] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 70 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 71.

[0066] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 72 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 73.

[0067] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 74 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 75.

[0068] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 76 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 77.

[0069] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 78 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 79.

[0070] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 80 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 81.

[0071] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 82 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 83.

[0072] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 84 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 85.

[0073] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 86 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 87.

[0074] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 88 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 89.

[0075] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 90 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 91.

[0076] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 92 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 93.

[0077] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 94 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 95.

[0078] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 96 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 97.

[0079] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 98 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 99.

[0080] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 100 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 101.

[0081] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 102 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 103.

[0082] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 104 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 105.

[0083] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 106 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 107.

[0084] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 108 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 109.

[0085] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 110 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 111.

[0086] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 112 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 113.

[0087] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 114 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 115.

[0088] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 116 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 117.

[0089] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 118 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 119.

[0090] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 120 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 121.

[0091] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 122 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 123.

[0092] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 124 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 125.

[0093] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 126 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 127.

[0094] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 128 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 129.

[0095] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 130 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 131.

[0096] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 132 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 133.

[0097] An antibody or a fragment thereof comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 300 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 304.

[0098] In Examples described later, conjugates with a thiol-containing maytansinoid were prepared using the polyclonal antibody or the monoclonal antibody against the full-length CAPRIN-1 protein or a polypeptide of a portion of a region thereof expressed on the cell membrane surface of cancer cells, and verified to have a strong antitumor effect.

[0099] <Thiol-containing maytansinoid> The thiol-containing maytansinoid is one of the maytansine derivatives that target microtubules and inhibit cell proliferation by inhibiting mitosis, and specific examples include DM1 (N(2')-deacetyl-N(2')-(3-mercapto-1-oxopropyl)-maytansine, CAS No. 139504-50-0), DM4 (N(2')-deacetyl-N2-(4-mercapto-4-methyl-1-oxopentyl)-maytansine, CAS No. 79984096-3), and derivatives thereof. In the present invention, DM1, DM4, and their derivatives are preferably used.

[0100] Specific examples of DM1 derivatives (including specific examples of conjugated compounds containing a linker) include N(2')-deacetyl-N2'-[3-(3-carboxy-1-methyl-propyldithio)-1-oxopropyl]-maytansine and compounds described in U.S. Patent No. 5,208,020.

[0101] Specific examples of DM4 derivatives (including specific examples of conjugated compounds containing a linker) include Ravtansine (also known as the IUPAC name: (1S,2R,3S,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl)                               (2S)-2-[methyl-(4-methyl-4- sulfanylpentanoyl)amino]propanoate, CAS:1194029-15-6, WOB38VS2NI, UNII-WOB38VS2NI,                    N2'-Deacetyl-N2'-(4-mercapto-4-methyl-1-oxopentyl)- maytansine(14S,16S,32S,33S,2R,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinana-3(2,3)-oxirana-8(1,3)-benzenacyclotetradecaphane-10,12-dien-4-yl N-(4-mercapto-4-methylpentanoyl)-N-methyl-L-alaninate), Maytansinoid DM4 (also known as the IUPAC name: [(1S,3S,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,2,5,9,16-pentamethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl](2S)-2-[methyl-(4-methyl-4-sulfanylpentanoyl)amino]propanoat), 799840-96-3, SCHEMBL21954026, HY-100503, CS-0019630), [(16Z,18Z)-11-chloro-21-hydroxy-12,20-dimethoxy-2,2,5,9,16-pentamethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl] 2-[methyl-(4-methyl-4-sulfanylpentanoyl)amino]propanoate, [(16E,18Z)-11-chloro-21-hydroxy-12,20-dimethoxy-2,2,5,9,16-pentamethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl] 2-[methyl-(4-methyl-4-sulfanylpentanoyl)amino]propanoate, Compound CID:131859751, Compound CID:134693255, DM4-SPDB, DM4-SPDB-L-Lysine, Soravtansine (DM4-Sulfo-TBA), S-Me-DM4, Slfo-SPDB-DM4, SPDB-DM4, DM4-Sme, Soravtansine-lysine, D-Lin-K-DM4, N2'-(4- ((3-Carboxypropyl)dithio)-4-methyl-1-oxopentyl)-N2'-deacetylmaytansine (also known as OX9S087G5M or DBA-DM4), DM4-SMCC, DBA-DM4, SSP-DM4, Maytansinoid DM4 impurity 2-d6, Sulfo-PDBA-DM4, DM4-SPDP (see Carlsson et al., Biochem. J., 173:723-737 (1978)), DM4 (hydrate), Maytansinoid DM4 impurity 5-d6, Maytansinoid DM4 impurity 3-d6, Maytansinoid DM4-d6, Sulfo-PDBA-DM4, DM4-d6 or a derivative thereof. Other examples include compounds described in U.S. Patent No. 5,208,020 and U.S. Patent No. 7,276,497, as well as DM4 containing a non-cleavable linker as described in WO 2005 / 037992. Non-cleavable linkers include those that undergo acid-induced cleavage, light-induced cleavage, peptidase-induced cleavage, esterase-induced cleavage, and reducing compound-induced cleavage of a disulfide bond.

[0102] <Conjugate of anti-CAPRIN-1 antibody and thiol-containing maytansinoid> In the present invention, the mode of binding between the anti-CAPRIN-1 antibody and the thiol-containing maytansinoid in the conjugate of the anti-CAPRIN-1 antibody and the thiol-containing maytansinoid is not particularly limited as long as the antitumor activity against a cancer can be maintained, and preferably is a mode of binding in which a linker structure is formed between the anti-CAPRIN-1 antibody and the thiol-containing maytansinoid.

[0103] In this context, the linker means a compound capable of linking the antibody or a fragment thereof to the drug. Any of various linkers known in the art may be used, or the drug may be chemically modified in the structure so as to function as a linker and then conjugated to the antibody (including the compounds described in the section <Thiol-containing maytansinoid> above).

[0104] The details of the type of the linker and the binding method can be basically in accordance with a method known in the art (see, for example, Greg T. Hermanson Bioconjugate Techniques, Third Edition, WO 2004 / 010957, and WO 2014 / 012479).

[0105] In an embodiment of the present invention, examples of reactive groups attached to the anti-CAPRIN-1 antibody, the thiol-containing maytansinoid, and the linker include the following.

[0106] Examples of the reactive group attached to the amino acid sequence of the antibody or a glycoprotein modifying an amino acid include primary amine (c-amino), carboxyl, thiol (sulfhydryl), carbonyl (ketone or aldehyde), and hydroxyl unless a special chemical modification is introduced. The primary amine exists at the N-terminus of a polypeptide or the side chain of a lysine residue and is positively charged under physiological conditions. The primary amine usually exists outside of the protein and can therefore be used in binding without denaturing the structure of the protein. The carboxyl exists at the C-terminus of a polypeptide or the side chain of aspartic acid or glutamic acid. The sulfhydryl exists at the side chain of cysteine and forms a disulfide bond that maintains the higher-order structure of the protein. The ketone or aldehyde is generated in a glycoprotein by the oxidation of glycosyl with sodium metaperiodate.

[0107] The conjugate of the present invention is prepared by binding the drug to the linker bound to the reactive group of the antibody, binding the thiol-containing maytansinoid to the linker bound to the drug, or directly binding the drug to the antibody.

[0108] Examples of the reactive groups attached to the thiol-containing maytansinoid and the linker include the following.

[0109] Reactive group capable of reacting with the amine: N-hydroxysuccinimide (NHS) ester, N-sulfosuccinimidyl ester, imido ester, pentafluorophenyl ester, hydroxymethylphosphine, isothiocyanate, isocyanate, acyl azide, N-hydroxyl ester, sulfonyl chloride, aldehyde, glyoxal, epoxide, oxirane, carbonate, aryl, carbodiimide, and carboxylic anhydride.

[0110] Reactive group capable of reacting with the carboxyl and the amine: carbodiimide, diazoalkane, diazoacetyl compounds, and carbonyldiimidazole.

[0111] Reactive group capable of reacting with the thiol: maleimide, haloacetamide, pyridyl disulfide, thiosulfone, vinyl sulfone, haloacetyl, aziridine, acryloyl, and aryl.

[0112] Reactive group capable of reacting with the aldehyde: hydrazide and alkoxyamine. Reactive group capable of reacting with the hydroxyl: epoxy, oxirane, carbonyldiimidazole, N,N'-disuccinimidyl carbonate, N-hydroxysuccinimidyl chloroformate, and isocyanate.

[0113] Reactive group capable of reacting with the hydroxyl: isocyanate.

[0114] Photoreactive reactive group: diaziridine, aryl azide, aryl, benzophenol, and diazo compounds.

[0115] A compound containing a dithiopyridyl group capable of forming a disulfide bond.

[0116] Specific examples of the linker having said reactive group include the following.

[0117] As a linker having the same reactive group ends, a linker having N-hydroxysuccinimide ester as a reactive group (e.g., Disuccinimidyl Glutarate (DSG), disuccinimidyl suberate (DSS), bis(sulfosuccinimidyl)suberate (BS3), tris-(succinimidyl)aminotriacetate (TSAT), PEGylated bis(sulfosuccinimidyl)suberate (BS(PEG)5, BS(PEG)9), dithiobis (succinimidyl propionate) (DSP), 3,3'-dithiobis (sulfosuccinimidyl propionate) (DTSSP), ethylene glycol bis (succinimidyl succinate) (EGS), Sulfo-ethylene glycol bis (succinimidyl succinate (Sulfo-EGS), Dimethyl adipimidate-2HCl (DMA), Dimethyl pimelimidate-2HCl (DMP), Dimethyl suberimidate-2HCl (DMS), Dimethyl3,3'-dithiobispropionimidate-2HCl (DTBP), 1,5-difluoro-2,4-dinitrobenzene (DFDNB), Disuccinimidyl tartrate (DST), Bis[2-(Succinimidooxycarbonyloxy)ethyl]Sulfone (BSOCOES)) and a linker having maleimide as a reactive group (e.g., Bismaleimidoethane (BMOE), 1,4-bismaleimidobutane (BMB), Bismaleimidohexane (BMH), Tris(2-maleimidothyl)amine (TMEA), 1,8-bismaleimido-(PEG)2 (BM(PEG)2), 1,8-bismaleimido-(PEG)3 (BM(PEG)3), and Dithiobismaleimidoethane (DTME)) may be used.

[0118] As a linker having different reactive group ends, a linker having NHS ester and maleimide as reactive groups (e.g., AMAS, BMPS, GMBS, Sulfo-MBS, MBS, Sulfo-MBS, SMCC, Sulfo-SMCC, EMCS, Sulfo-EMCS, SMPB, Sulfo-SMPB, SMPH, LC-SMCC, Sulfo-KMUS, SM(PEG)2, SM(PEG)4, SM(PEG)6, SM(PEG)8, SM(PEG)12, and SM(PEG)24), a linker having NHS ester and pyridyldithiol as reactive groups (e.g., SPDP, LC-SPDP, Sulfo-LC-SPDP, SMPT, (PEG)4-SPDP, and PEG12-SPDP), a linker having NHS ester and haloacetyl as reactive groups (e.g., SIA, SBA, SBAP, SIAB, and Sulfo-SIAB), a linker having NHS ester and aryl azide as reactive groups (e.g., ANB-NOS, Sulfo-SANPAH, and ATFB), a linker having NHS ester and diaziridine as reactive groups (e.g., SDA, Sulfo-SDA, LC-SDA, SDAD, and Sulfo-SDAD), a linker having carbodiimide as reactive groups (e.g., DCC, EDC, EDAC, NHS, and Sulfo-NHS), a linker having maleimide and hydrazide as reactive groups (e.g., BMPH, EMCH, MPBH, and KMUH), a linker having pyridyldithiol and hydrazide as a reactive group (e.g., PDPH), a linker having isocyanate and maleimide as reactive groups (e.g., PMPI), and a linker having NHS ester and psoralen as reactive groups (e.g., SPB) may be used.

[0119] As other linkers, a linker containing a polypeptide, for example, Fmoc-Ala-Ala-Asn-PAB, Fmoc-Ala-Ala-Asn(Trt)-PAB, Fmoc-PEG3-Ala-Ala-Asn(Trt)-PAB, Fmoc-PEG4-Ala-Ala-Asn(Trt)-PAB, Fmoc-Ala-Ala-Asn-PAB-PNP, Fmoc-Ala-Ala-Asn(Trt)-PAB-PNP, Fmoc-PEG3-Ala-Ala-Asn(Trt)-PAB-PNP, Azide-PEG4-Ala-Ala-Asn(Trt)-PAB-PNP, Mal-PEG4-Ala-Ala-Asn(Trt)-PAB-PNP, Fmoc-Val-Cit-PAB-OH, Val-Cit-PAB-OH, Fmoc-Val-Cit-PAB-PNP, MC-Val-Cit-PAB, MC-Val-Cit-PAB-PNP may be used.

[0120] In addition, Bis-PEG-acid, PEG Acid (e.g., Acid-PEG-TEMPO, Amino-PEG-acid, Amino-PEG-CH2CO2H, Aminoxy-PEG-acid, Azido-PEG-acid, Carboxy-PEG-sulfonic acid, Fmoc-N-amido-PEG-acid, Fmoc-N-amido-PEG-CH2CO2H, Fmoc-aminooxy-PEG-acid, Hydroxy-PEG-acid, Hydroxy-PEG-CH2CO2H, m-PEG-acid, m-PEG-(CH2)3-acid, Methoxytrityl-N-PEG-acid, N-methyl-N-(t-Boc)-PEG-acid, Propargyl-PEG-acid, Propargyl-PEG-CH2CO2H, Propargyl-PEG-(CH2)3-acid, t-Boc-N-amido-PEG-acid, t-Boc-N-amido-PEG-CH2CO2H, t-Boc-Aminooxy-PEG-acid, Acid-PEG-PFP ester, Miscellaneous PEG acid), PEG PFP ester (e.g., Acid-PEG-PFP ester, Bis-PEG-PFP ester), Bis-PEG-NHS, PEG Aldehyde (e.g., m-PEG-aldehyde, m-PEG-benzaldehyde, Ald-PEG-acid, Ald-PEG-amine, Ald-PEG-azide, Ald-PEG-NH-Boc, Ald-PEG-NHS ester, Ald-PEG-TFP ester, Ald-PEG-t-butyl ester), PEG Tosylate (e.g., Azido-PEG-Tos, Hydroxy-PEG-Tos, m-PEG-Tos, t-Boc-Aminooxy-PEG-Tos, Trifluoroethyl-PEG-Tos, Tos-PEG-acid, Tos-PEG-CH2CO2H, Tos-PEG-alkyne, Tos-PEG-t-butyl ester, Tos-PEG-CH2CO2tBu, Tos-PEG-Tos, S-acetyl-PEG6-Tos, N-Tos-N-(t-butoxycarbonyl)-aminooxy-PEG4-Tos, Ms-PEG-Ms, Ms-PEG-t-butyl ester, PEG-Ms, Propargyl-PEG-Ms), Boc-PEG (e.g., Amino-PEG-t-Boc-Hydrazide, Azido-PEG-t-Boc-Hydrazide, Boc-NH-PEG-NH-Boc, Bromoacetamido-PEG-Boc-amine, m-PEG-ONHBoc, Mal-Alkyl-t-Boc-amine, N-Boc-PEG-alcohol, N-Boc-PEG-bromide, N-methyl-N-(t-Boc)-PEG-acid, t-Boc-N-amido-PEG-acid, t-Boc-N-amido-PEG-CH2CO2H, t-Boc-N-Amido-PEG-amine, t-Boc-N-amido-PEG-azide, t-Boc-N-amido-PEG-NHS ester, t-Boc-N-amido-PEG-sulfonic acid), PEG NHS ester (e.g., Acid-PEG-NHS ester, Azido-PEG-NHS ester, Bis-PEG-NHS, Fmoc-PEG-NHS ester, m-PEG-NHS ester, m-PEG-NHS Carbonate, Mal-PEG-NHS ester, Propargyl-PEG-NHS ester, t-Boc-N-amido-PEG-NHS ester, t-Butoxycarbonyl-PEG-NHS ester), Fmoc-PEG (e.g., Fmoc-N-amido-PEG-acid, Fmoc-NH-PEG-CH2CO2H, Fmoc-PEG-NHS ester), Biotin PEG (e.g., Biotin PEG-acid, Biotin PEG-alcohol, Biotin PEG-alkyne, Biotin PEG-amine, Biotin PEG-azide, Biotin PEG-DBCO, Biotin PEG-hydrazide, Biotin-PEG-Mal, Biotin-PEG-NHS, Biotin-EDA-PEG-NHS, Biotin-PEG-oxyamine, Biotin-PEG-PFP, Biotin-EDA-PEG-PFP, Biotin-PEG-Tetrazine, Biotin-PEG-TFP, Azide-SS-biotin, Biotin-PEG3-SS-azide, DBCO-S-S-PEG3-Biotin, Dde Biotin-PEG4-Alkyne, Dde Biotin-PEG4-Azide, Dde Biotin-PEG4-DBCO, Diazo Biotin-PEG3-Alkyne, Diazo Biotin-PEG3-Azide, Diazo Biotin-PEG3-DBCO, Diol Biotin-PEG3-Alkyne, Diol Biotin-PEG3-Azide, PC Biotin-PEG3-Alkyne, PC-Biotin-PEG4-PEG4-Alkyne, PC-Biotin-PEG4-PEG4-Alkyne, PC Biotin-PEG3-Azide, PC-Biotin-PEG4-PEG3-Azide, PC-Biotin-PEG4-NHS carbonate, PC DBCO-PEG3-Biotin, WSPC Biotin-PEG3-DBCO, Fmoc-Lys (biotin-PEG)-OH, Fmoc-N-amido-(PEG-biotin)-acid, TAMRA-Azide-PEG-Biotin), PEG Phosphonate, Aminooxy PEG (e.g., Aminooxy-PEG-acid, Aminooxy-PEG-alcohol, Aminooxy-PEG-azide, Aminooxy-PEG-bromide, Aminooxy-PEG-methane, Aminooxy-PEG-Propargyl, Aminooxy-PEG-t-butyl ester, Aminooxy-PEG-Thiol, Bis-(Aminooxy)-PEG, t-Boc-Aminooxy-PEG-acid, t-Boc-Aminooxy-PEG-alcohol, t-Boc-Aminooxy-PEG-amine, t-Boc-Aminooxy-PEG-Azide, t-Boc-Aminooxy-PEG-Bromide, t-Boc-aminooxy-PEG-Methane, t-Boc-aminooxy-PEG-Propargyl, t-Boc-aminooxy-PEG-S-Ac, t-Boc-Aminooxy-PEG-Thiol, t-Boc-Aminooxy-PEG-Tos, Fmoc-aminooxy-PEG-acid, Trifluoroethyl-PEG-Aminooxy), Alkyne PEG (e.g., endo-BCN-PEG, exo-BCN-PEG, Propargyl-PEG-acid, Propargyl-PEG-CH2CO2H, Propargyl-PEG-(CH2)3-acid, Propargyl-PEG-(CH2)3-methyl ester, Propargyl-PEG-Acrylate, Propargyl-PEG-alcohol, Propargyl-PEG-amine, Propargyl-PEG-methylamine, Aminooxy-PEG-Propargyl, Propargyl-PEG-azide, Propargyl-PEG-bromide, Propargyl-PEG-Maleimide, Propargyl-PEG-Ms, Propargyl-PEG-NHS ester, Propargyl-PEG-sulfonic acid, Propargyl-PEG-t-butyl ester, Propargyl-PEG-CH2CO2tBu, Propargyl-PEG-thiol, Propargyl-PEG-5-nitrophenyl carbonate, t-Boc-aminooxy-PEG-Propargyl, Bis-Propargyl-PEG, m-PEG-Propargyl), Azido PEG (e.g., Azido-PEG-acid, Azido-PEG-CH2CO2H, Azido-PEG-(CH2)3-methyl ester, Azido-PEG-Acrylate, Azido-PEG-alcohol, Azido-PEG-(CH2)3OH, Azido-PEG-amine, Azido-PEG-azide, Azido-PEG-Maleimide, Azido-PEG-methylamine, Azido-PEG-methyl ester, Azido-PEG-NHS ester, Azido-PEG-CH2CO2-NHS, Azido-PEG-oxazolidin-2-one, Azido-PEG-PFP ester, Azido-PEG-phosphonic acid, Azido-PEG-phosphonic acid ethyl ester, Azido-PEG-sulfonic acid, Azido-PEG-t-Boc-Hydrazide, Azido-PEG-t-butyl ester, Azido-PEG-CH2CO2-t-butyl ester, Azido-PEG-TFP ester, Azido-PEG-Tos, Aminooxy-PEG-azide, Bromo-PEG-azide, Bromoacetamido-PEG-azide, Carboxyrhodamine 110-PEG-Azide, Isothiocyanato-PEG-Azide, Isothiocyanato-PEG-Azide, m-PEG-azide, Propargyl-PEG-azide, TAMRA-PEG-Azide, t-Boc-N-Amido-PEG-Azide, t-Boc-Aminooxy-PEG-Azide, Thiol-PEG-Azide, Trifluoroethyl-PEG-Azide, Azido-PEG-amino acid, Azido-PEG4-4-nitrophenyl carbonate, S-Acetyl-PEG3-Azido, Azide, Trityl-PEG10-Azide), Alkyne PEG, DBCO-PEG, BCN-PEG, Propargyl-PEG, Bis-PEG-acid, Bis-PEG-NHS, Bis-PEG-PFP, Bis-Propargyl-PEG, Amine-PEG-Amine, Azido-PEG-azide, Bromo-PEG, or Mal PEG may be used.

[0121] In another embodiment, polyethylene glycols (PEGs) described in WO 2015 / 057699 and WO 2017 / 165851 may be used to obtain conjugates that are expected to provide better in vivo kinetics.

[0122] The linker between the anti-CAPRIN-1 antibody and the thiol-containing maytansinoid may be composed of a single type or composed of a plurality of types.

[0123] In another embodiment, a charged linker described in WO 2014 / 186403 is used.

[0124] Methods for preparing the conjugate of the present invention include a method that utilizes the s-amino group at the lysine side chain of the antibody or a fragment thereof for conjugation, and a method that utilizes thiol formed by reducing a cysteine residue constituting a disulfide bond of the antibody for conjugation.

[0125] In the case of utilizing a s-amino group at the lysine residue of the antibody, for example, a method is used which involves reacting active ester (e.g., N-hydroxysuccinimide ester) therewith to form an amide bond. In this case, since the antibody or a fragment thereof contains many lysine residues, the binding reaction proceeds nonspecifically.

[0126] In the case of utilizing thiol constituting a disulfide bond and present at the side chain of cysteine in the antibody or a fragment thereof, a method in which thiol is formed from the disulfide bond on the antibody or a fragment thereof using a reducing agent such as mercaptoethanol and reacted with maleimide or a-haloamide is used. For example, a method using sulfone phenyloxadiazole, a 4-cyanoethynyloxy derivative, or the like is used for stabilizing a thiol-mediated bond. These bonds are stable for a longer time than the bond based on the conjugate addition reaction of cysteine with maleimide. In addition, a linker having an amino group near an imide group may also be used because an imide ring by adding a thiol group to maleimide is opened by hydrolysis so that stability is improved owing to the resulting amide bond. The thiols of cysteines form a disulfide bond in the antibody or a fragment thereof, and thus an alternative method involves binding the drug via two thiols by binding in between. As an example, a cross-linked bond may be formed using a linker having two disulfide bond sites that can be formed from an amide group having two sulfones at the P position, or dibromomaleimide.

[0127] The conjugate of the present invention may be prepared by a method using, for example, the THIOMAB (trademark) technique (see Nature Biotechnology 26, 925-932 (2008)), the ThioBridge (trademark) technique, (see Methods Mol Bio. 2078: 113-129 (2020)), or the RESPECT (trademark) technique (see MAbs. 9(6): 907-915 (2017)), which is a method capable of introducing a determined number of thiol groups at a particular site of an antibody or a fragment thereof.

[0128] The conjugate of the present invention may be formed, for example, by reducing the antibody or a fragment thereof using a reducing agent dithiothreitol (DTT) in a phosphate buffer to obtain an antibody having a reactive thiol group, which is then conjugated with the thiol-containing maytansinoid. The conjugate can be obtained by adding a thiol group to primary amine at the lysine residue of the antibody by the introduction of a Traut's reagent (2-iminothiolane or N-succinimidyl S-acetylthioacetate (SATA)), instead of the method using a reducing agent.

[0129] The amount of the thiol added to the antibody or a fragment thereof can be determined, for example, by mixing a sample solution containing 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) and an SH group with a phosphate buffer solution (pH 8.0) and distilled water, adding a solution of DTNB dissolved in a phosphate buffer, a Good's buffer, or a Tris buffer to the mixture, and incubating the resulting for a given time, followed by the measurement of absorbance at 412 nm (see G.L. Ellman, Arch. Biochem. Biophys., 82, 70 (1959)).

[0130] The thiol group added by the cleavage of the disulfide bond of the antibody through reduction treatment is preferably treated (capped) in order to prevent the formation of a disulfide bond again. For example, N-ethylmaleimide (NEM) or 2-iodoacetamide (IAA) may be employed for the capping.

[0131] The formation of a conjugate using a thiol group added to the antibody or a fragment thereof can be carried out by known methods. Specifically, the binding can be carried out using, for example, a linker reagent having a maleimide group or a bromoacetamide group as a linker reagent specifically binding to the thiol group of the reduced antibody. For example, N-succinimidyl-4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (SMCC) is used as the linker having a maleimide group. In this case, when an amino group is present in the drug, the conjugate can be obtained by forming an amide bond with the N-succinimide group of SMCC.

[0132] In another embodiment, the conjugate can be obtained by first forming an amide bond using SMCC with an amino group present in the drug, and then reacting the maleimide group of the SMCC in the drug with a thiol group added to the antibody or a fragment thereof.

[0133] In another embodiment, the conjugate may be formed using two linkers. For example, a primary amino group present at the lysine residue of the antibody or a fragment thereof is bound to the N-succinimide group of SATA (N-succinimidyl-S-acetylthioacetate) via an amide bond to add a thiol group to the antibody or a fragment thereof. SMCC is reacted with the drug containing an amino group or having an amino group added thereto according to an ordinary method to form an amide bond with the N-succinimide group of the SMCC. Then, the maleimide group of the SMCC on the drug can be reacted with the thiol group of the SATA on the antibody or a fragment thereof to obtain the conjugate.

[0134] In another further embodiment, examples of the preparation of the conjugate include a method using maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (MC-Val-Cit-PAB) as a linker. MC-val-Cit-PAB (mc-vc-PAB) is a linker cleavable by intracellular protease (e.g., cathepsin B). A thiol group is added to the antibody or a fragment thereof dissolved in a phosphate buffer using DTT or the like. Meanwhile, the thiol-containing maytansinoid having an amino group is reacted with the benzyloxycarbonyl (PAB) in MC-Val-Cit-PAB to prepare a thiol-containing maytansinoid bound with MC-val-Cit-PAB, which can then be reacted with the thiol-added antibody or a fragment thereof described above to obtain the conjugate.

[0135] In another further embodiment, the conjugate is included in which SATA is bound to a primary amino group at the lysine residue of the antibody or a fragment thereof to add a thiol group thereto, and succinimidyl 3-(2-pyridyldithio)propionate (SPDP) is reacted with the amino group of the drug to form an amide bond with the N-succinimide group of the SPDP.

[0136] As the linker used in the present invention, a linker cleavable under intracellular conditions can be used, and the use of such a linker allows a substance having antitumor activity that contains the drug or the drug together with a portion of the linker, to be released inside the cell. For example, the linker is a linker cleaved by peptidase or protease in a cell. Preferably, the linker is a linker cleaved by lysosome, endosomal protease, cathepsin B, cathepsin D, or plasmin. Examples thereof include a linker comprising a polypeptide (Val-Cit, Phe-Leu, or Gly-Phe-Leu-Gly) cleavable by cathepsin B. More specifically, the linker described in U.S. Patent No. 6,214,345 may be used.

[0137] In still another embodiment, as a method for improving the stability, solubility, and metabolism in blood of the conjugate of the present invention and the binding of the drug to the antibody or a fragment thereof in the conjugate, a linker having glucuronic acid (preferably a p-D-glucuronide) as described, for example, in WO 2007 / 011968 can be used. Alternatively, the methods described in WO 2013 / 173337, WO 2015 / 095755, WO 2015 / 123679, and WO 2018 / 031690 may be used.

[0138] In another further embodiment, for example, methods described in WO 2006 / 65533 and WO 2018 / 160683 may be used to site-specifically bind the drug to the antibodies or fragments thereof.

[0139] In still another embodiment, a conjugate in which two or more drugs containing a thiol-containing maytansinoid are bound to an anti-CAPRIN-1 antibody can be obtained by a method described in WO 2018 / 112253.

[0140] In order to obtain the conjugate of the present invention, for example, a peak fraction of a higher molecular weight compared to the antibody before the binding of the linker may be collected by the application of gel filtration chromatography or the like. The method described in, for example, WO 2013 / 049410 may be used to detect the mass of the conjugate while maintaining an intact bivalent antibody.

[0141] The number of thiol-containing maytansinoid molecules bound per molecule of the antibody in the conjugate of the present invention can be quantified by known methods, such as mass spectrometry, ELISA, electrophoresis, and chromatographic techniques including HPLC.

[0142] <Antitumor effect of conjugate> The conjugate of the present invention has antitumor activity in vitro or in vivo. The antitumor activity means reduction, elimination, growth inhibition, apoptosis, necrosis, or killing of target cancer cells. Accordingly, the antitumor effect of the conjugate of the present invention may be determined by examining its antitumor activity against a cancer.

[0143] In order to determine the antitumor effect in vivo, the antitumor activity can be evaluated by: administering the conjugate to an organism having a cancer; and examining the size of the cancer over time via measuring the size of the tumor after the administration. The antitumor effect of the present invention can also be evaluated by examining a survival rate. Alternatively, the antitumor effect of the present invention may be evaluated by examining the ability to produce a cytokine or a chemokine. The antitumor effect of the conjugate of the present invention can be further determined by examining the prevention of a cancer, the prevention of metastasis, or the prevention of recurrence.

[0144] The conjugate of the present invention is expected to have a stronger antitumor effect, when the conjugate has higher binding affinity for the CAPRIN-1 protein on cancer cell surface. Its association constant (affinity constant) Ka (kon / koff) is preferably at least 107 M-1, at least 108 M-1, at least 5 x 108 M-1, at least 109 M-1, at least 5 x 109 M-1, at least 1010 M-1, at least 5 x 1010 M-1, at least 1011 M-1, at least 5 x 1011 M-1, at least 1012 M-1, or at least 1013 M-1.

[0145] The ability of the conjugate of the present invention to bind to CAPRIN-1 protein can be identified through the use of binding assay using, for example, surface plasmon resonance (SPR), ELISA, Western blot, immunofluorescence, or flow cytometry.

[0146] The conjugate of the present invention enhances the antitumor effect compared to the anti-CAPRIN-1 antibody alone, as described above. The rate of the enhancement is preferably 30% or more, more preferably 40% or more, further preferably 50% or more, still further preferably 55% or more, even further preferably 60% or more, furthermore preferably 65% or more, most preferably 70% or more. The rate of enhancement in antitumor effect by the conjugate of the present invention with respect to the anti-CAPRIN-1 antibody alone can be calculated by administering their respective effective amounts to tumor-bearing mice under the same conditions, and comparing tumor volumes 10 days or later after the start of the administration.

[0147] <Pharmaceutical composition, method for treating and / or preventing cancer> The target of the pharmaceutical composition for the treatment and / or prevention of a cancer of the present invention is not particularly limited as long as the target is cancer (cells) expressing the CAPRIN-1 protein.

[0148] The terms "tumor" and "cancer" used in the present specification mean malignant neoplasm and are used interchangeably with each other.

[0149] The cancer targeted in the present invention may be any cancer expressing the CAPRIN-1 protein on the cell membrane surface. The cancer is preferably breast cancer, kidney cancer, pancreatic cancer, colorectal cancer, lung cancer, brain tumor, gastric cancer, uterine cancer, ovarian cancer, prostate cancer, bladder cancer, esophageal cancer, leukemia, lymphoma, liver cancer, gallbladder cancer, sarcoma, mastocytoma, melanoma, adrenocortical carcinoma, Ewing's tumor, Hodgkin's lymphoma, mesothelioma, multiple myeloma, testicular cancer, thyroid cancer, head and neck cancer, or urothelial carcinoma as described above.

[0150] More specifically, examples of these cancers include, but are not limited to, breast adenocarcinoma, composite type breast adenocarcinoma, malignant mammary mixed tumor, intraductal papillary adenocarcinoma, recurrent metastatic breast cancer, lung adenocarcinoma, non-small cell lung cancer (NSCLC), squamous non-small cell lung cancer, squamous cell cancer, small-cell cancer, large-cell cancer, glioma that is a tumor of neuroepithelial tissue, glioblastoma, neuroblastoma, ependymoma, neuronal tumor, embryonal neuroectodermal tumor, schwannoma, neurofibroma, meningioma, chronic lymphocytic leukemia, lymphoma, gastrointestinal lymphoma, digestive lymphoma, small-cell-to-medium-cell lymphoma, cecal cancer, ascending colon cancer, descending colon cancer, transverse colon cancer, sigmoid colon cancer, rectal cancer, epithelial ovarian cancer, germ cell tumor, stromal cell tumor, pancreatic ductal carcinoma, invasive pancreatic ductal carcinoma, pancreatic adenocarcinoma, metastatic adenocarcinoma, acinar cell carcinoma, adenosquamous carcinoma, giant cell tumor, intraductal papillary-mucinous neoplasm, mucinous adenocarcinoma, pancreatoblastoma, isletcell adenoma, Frants tumor, serous cystadenocarcinoma, solid-pseudopapillary cancer, gastrinoma, glucagonoma, insulinoma, multiple endocrine neoplasia type-1 (Wermer's syndrome), nonfunctional islet cell tumor, somatostatinoma, VIPoma, uterine cervix cancer, uterine body cancer, fibrosarcoma, sarcoma of bones or joints, Ewing's sarcoma, Wilms tumor, hepatoblastoma, soft tissue sarcoma, acute leukemia, chronic leukemia, spinal cord tumor, malignant soft tissue tumor, teratoma group tumor, and head and neck cancer including hypopharynx cancer, oropharynx cancer, tongue cancer, nasopharyngeal cancer, oral cavity cancer, lip cancer, sinus cancer, laryngeal cancer, and recurrent brain tumor.

[0151] The subjects (patients) to be targeted are preferably mammals, for example, mammals including primates, pet animals, livestock, and sport animals and are particularly preferably humans, dogs, and cats.

[0152] In the case of using the conjugate used in the present invention in a pharmaceutical composition, the pharmaceutical composition can be formulated by a method known to those skilled in the art. For example, the pharmaceutical composition may be used in the form of a parenterally injectable agent of an aseptic solution or suspension with water or any other pharmaceutically acceptable liquid. For example, the conjugate may be formulated in a unit dosage form required for generally accepted pharmaceutical practice, by mixing with pharmacologically acceptable carriers or media, specifically, sterilized water, physiological saline, a plant oil, an emulsifier, a suspending agent, a surfactant, a stabilizer, a fragrance, an excipient, a binder, or the like, in appropriate combination. The amount of the active ingredient in such a preparation is determined so that an appropriate dose within the prescribed range can be achieved.

[0153] In the case of using the conjugate of the present invention in a pharmaceutical composition, the pharmaceutical composition may be formulated in a lyophilized state, comprising any salts, surfactants, buffers, sugars, cryoprotectants (including some sugars). For example, a pharmaceutical composition comprising excipients containing sucrose, polysorbate 20, polysorbate 80, cyclodextrin, glucose, glycerol, polyethylene glycol, mannitol, sodium chloride, and amino acids as described in WO 2007 / 019232, a solution of which is prepared as a pharmaceutical preparation having a pH of 4 to 8, preferably 4.5 to 7.6 may be used.

[0154] An aseptic composition for injection can be formulated according to conventional pharmaceutical practice using a vehicle such as injectable distilled water. Examples of aqueous solutions for injection include saline, isotonic solutions containing glucose and other auxiliary agents (specifically, D-sorbitol, D-mannose, D-mannitol, or sodium chloride), and these solutions may be used in combination with an appropriate solubilizer, for example, an alcohol (specifically, ethanol, a polyalcohol, propylene glycol or polyethylene glycol), or a nonionic surfactant, (specifically, Polysorbate 80 (TM) or HCO-60). Examples of an oil solution include sesame oil and soybean oil, and may be used in combination with benzyl benzoate or benzyl alcohol as a solubilizer. The solution may also be mixed with a buffer (specifically, a phosphate buffer solution or a sodium acetate buffer solution), a soothing agent (specifically, procaine hydrochloride), a stabilizer (specifically, benzyl alcohol or phenol), and an antioxidant. The injection solutions thus prepared are usually filled into appropriate ampules. Examples of oil solutions include sesame oil and soybean oil, and may be used in combination with benzyl benzoate or benzyl alcohol as a solubilizer. In addition, the solution may be mixed with a buffer (specifically, a phosphate buffer or a sodium acetate buffer), a soothing agent (specifically, procaine hydrochloride), a stabilizer (specifically, benzyl alcohol or phenol), and an antioxidant. The injection solution thus prepared is usually filled into an appropriate ampule.

[0155] The administration is carried out orally or parenterally, preferably parenterally. Specific examples include injectable agents, intranasal administration preparations, transpulmonary administration preparations, and percutaneous administration preparations. Examples of the injectable agents include intravenous injection, intramuscular injection, intraperitoneal injection, subcutaneous injection, and intratumoral administration, through which the pharmaceutical composition can be administered systemically or locally.

[0156] Also, the administration method may be appropriately selected in view of the age, weight, sex, symptoms, or the like of a patient. The dose of a pharmaceutical composition containing the antibody or a polynucleotide encoding the antibody may be selected within a range of, for example, 0.0001 mg to 1000 mg per kg of body weight per dose, for example, 0.5 mg, 1 mg, 2 mg, 3 mg, 5 mg, 10 mg, 20 mg, 50 mg, 75 mg, 100 mg, 200 mg, 500 mg, or 1000 mg per kg of body weight per dose. Alternatively, the dose can be selected within a range of, for example, 0.001 to 100000 mg / body of a patient, though the dose is not necessarily limited to these numeric values.

[0157] Although the dose and the administration method vary depending on the weight, age, sex, symptoms, or the like of a patient, those skilled in the art can appropriately select the dose and the method.

[0158] The pharmaceutical composition for the treatment and / or prevention of a cancer comprising the conjugate of the present invention as an active ingredient can be administered to a subject to treat and / or prevent the aforementioned cancer expressing CAPRIN-1 on the cell membrane surface, preferably breast cancer, kidney cancer, pancreatic cancer, colorectal cancer, lung cancer, brain tumor, gastric cancer, uterine cancer, ovarian cancer, prostate cancer, bladder cancer, esophageal cancer, leukemia, lymphoma, liver cancer, gallbladder cancer, sarcoma, mastocytoma, melanoma, adrenocortical carcinoma, Ewing's tumor, Hodgkin's lymphoma, mesothelioma, multiple myeloma, testicular cancer, thyroid cancer, head and neck cancer, or urothelial carcinoma. Examples

[0159] Hereinafter, the present invention will be specifically described with reference to Examples. However, the scope of the present invention is not intended to be limited by these specific examples.

[0160] (Example 1) Anti-CAPRIN-1 polyclonal antibody In order to obtain anti-CAPRIN-1 polyclonal antibodies having immunological reactivity with the CAPRIN-1 protein to be used in conjugates of the present invention, 1 mg of a recombinant human CAPRIN-1 protein of SEQ ID NO: 2 or SEQ ID NO: 4 produced according to Example 3 of WO 2010 / 016526 was mixed with an equal volume of incomplete Freund's adjuvant (IFA) solution, and this mixture was subcutaneously administered to rabbits four times every 2 weeks. Then, blood was collected to obtain antiserum containing polyclonal antibodies. The obtained antiserum was purified using a protein G carrier (GE Healthcare BioSciences Corp.) to prepare a polyclonal antibody against the CAPRIN-1 protein (anti-CAPRIN-1 polyclonal antibody #1). Additionally, the serum of a rabbit to which an antigen is not administered was purified using a protein G carrier in the same manner as above and used as a rabbit control antibody.

[0161] Additionally, the following polyclonal antibodies #2 to #6 against partial polypeptides of CAPRIN-1 were obtained in the same manner as in the method for preparing the polyclonal antibody against the CAPRIN-1 protein.

[0162] Anti-CAPRIN-1 polyclonal antibody #2 against a partial polypeptide represented by SEQ ID NO: 37 disclosed in WO 2011 / 096528 (SEQ ID NO: 31 of the present specification).

[0163] Anti-CAPRIN-1 polyclonal antibody #3 against a partial polypeptide represented by SEQ ID NO: 5 disclosed in WO 2013 / 018894 (SEQ ID NO: 32 of the present specification).

[0164] Anti-CAPRIN-1 polyclonal antibody #4 against a partial polypeptide represented by SEQ ID NO: 5 disclosed in WO 2013 / 125654 (SEQ ID NO: 33 of the present specification).

[0165] Anti-CAPRIN-1 polyclonal antibody #5 against a partial polypeptide represented by SEQ ID NO: 37 disclosed in WO 2011 / 096533 (SEQ ID NO: 34 of the present specification).

[0166] Anti-CAPRIN-1 polyclonal antibody #6 against a partial polypeptide represented by SEQ ID NO: 37 disclosed in WO 2011 / 096534 (SEQ ID NO: 35 of the present specification).

[0167] (Example 2) Anti-CAPRIN-1 monoclonal antibody The following anti-CAPRIN-1 monoclonal antibodies were used in the conjugate of the present invention. The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2011 / 096528, wherein the antibody comprises CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 36, SEQ ID NO: 37, and SEQ ID NO: 38, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 40, SEQ ID NO: 41, and SEQ ID NO: 42, respectively, and wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by SEQ ID NO: 39, and the amino acid sequence of a light chain variable region represented by SEQ ID NO:43.

[0169] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2015 / 020212, wherein the antibody comprises CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 44, SEQ ID NO: 45, and SEQ ID NO: 46, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 48, SEQ ID NO: 49, and SEQ ID NO: 50, respectively, and wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by SEQ ID NO: 47, and the amino acid sequence of a light chain variable region represented by SEQ ID NO: 51.

[0170] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2011 / 096519, wherein the antibody comprises CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 52, SEQ ID NO: 53, and SEQ ID NO: 54, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 56, SEQ ID NO: 57, and SEQ ID NO: 58, respectively, and wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by SEQ ID NO: 55, and the amino acid sequence of a light chain variable region represented by SEQ ID NO: 59.

[0171] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 125654, wherein the antibody comprises CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 60, SEQ ID NO: 61, and SEQ ID NO: 62, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 64, SEQ ID NO: 65, and SEQ ID NO: 66, respectively, and wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by SEQ ID NO: 63, and the amino acid sequence of a light chain variable region represented by SEQ ID NO: 67.

[0172] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2011 / 096517, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 68 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 69.

[0173] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2011 / 096528, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 70 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 71; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 72 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 73; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 74 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 75; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 76 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 77; or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 78 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 79.

[0174] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2011 / 096533, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 80 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 81, or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 82 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 83.

[0175] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2011 / 096534, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 84 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 85, or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 86 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 87.

[0176] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2010 / 016526, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 88 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 89; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 90 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 91; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 92 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 93; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 94 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 95; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 96 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 97; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 98 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 99; or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 100 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 101.

[0177] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 018894, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 102 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 103, or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 104 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 105.

[0178] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 018892, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 106 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 107.

[0179] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 018891, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 108 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 109.

[0180] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 018889, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 110 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 111.

[0181] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 018883, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 112 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 113.

[0182] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 125636, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 114 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 115.

[0183] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 125654, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 116 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 117, or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 118 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 119.

[0184] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2013 / 125630, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 120 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 121.

[0185] The anti-CAPRIN-1 monoclonal antibody disclosed in WO 2015 / 020212, wherein the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 122 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 123; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 124 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 125; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 126 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 127; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 128 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 129; the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 130 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 131; or the antibody comprises the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 132 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 133.

[0186] A nucleotide sequence was designed to express a heavy chain variable region comprising CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 36, SEQ ID NO: 37, and SEQ ID NO: 38, respectively, which are from one of the abovedescribed anti-CAPRIN-1 monoclonal antibodies, and framework regions comprising those of a human antibody sequence; and the nucleotide sequence was inserted into a mammalian expression vector with an insert encoding a heavy chain constant region of human IgG1. Similarly, a nucleotide sequence was designed to express a light chain variable region comprising CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 40, SEQ ID NO: 41, and SEQ ID NO: 42, respectively, and framework regions comprising those of a human antibody sequence; and the nucleotide sequence was inserted into a mammalian expression vector with an insert encoding a light chain constant region of human IgG1. These two recombinant expression vectors were transferred to mammalian cells according to a conventional method, and a culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #1 (humanized antibody #1) comprising CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 36, SEQ ID NO: 37, and SEQ ID NO: 38, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 40, SEQ ID NO: 41, and SEQ ID NO: 42, respectively, was obtained from the cells.

[0187] Similarly, a nucleotide sequence was designed to express a heavy chain variable region represented by SEQ ID NO: 47 comprising CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 44, SEQ ID NO: 45, and SEQ ID NO: 46, respectively, and framework regions comprising those of a human antibody sequence; and the nucleotide sequence was inserted into a mammalian expression vector with an inserted heavy chain constant region of human IgG1. Similarly, a nucleotide sequence was designed to express a heavy chain variable region represented by SEQ ID NO: 51 comprising CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 48, SEQ ID NO: 49, and SEQ ID NO: 50, respectively, and framework regions comprising those of a human antibody sequence; and the nucleotide sequence was inserted into a mammalian expression vector with an inserted heavy chain constant region of human IgG1. These two recombinant expression vectors were transferred to mammalian cells according to a conventional method, and a culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #2 (humanized antibody #2) comprising CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 44, SEQ ID NO: 45, and SEQ ID NO: 46, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 48, SEQ ID NO: 49, and SEQ ID NO: 50, respectively, was obtained from the cells.

[0188] A culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #3 (humanized antibody #3) comprising CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 52, SEQ ID NO: 53, and SEQ ID NO: 54, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 56, SEQ ID NO: 57, and SEQ ID NO: 58, respectively, was prepared in the same manner.

[0189] A culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #4 (humanized antibody #4) comprising CDR1-3 of heavy chain variable region consisting of the amino acid sequences of SEQ ID NO: 60, SEQ ID NO: 61, and SEQ ID NO: 62, respectively, and CDR1-3 of light chain variable region consisting of the amino acid sequences of SEQ ID NO: 64, SEQ ID NO: 65, and SEQ ID NO: 66, respectively, was prepared in the same manner.

[0190] Culture supernatants containing the following humanized anti-CAPRIN-1 monoclonal antibodies #9 to #41 (humanized antibodies #9 to #41) were prepared in the same manner.

[0191] Humanized anti-CAPRIN-1 monoclonal antibody #9 (humanized antibody #9) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 68 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 69.

[0192] Humanized anti-CAPRIN-1 monoclonal antibody #10 (humanized antibody #10) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 70 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 71.

[0193] Humanized anti-CAPRIN-1 monoclonal antibody #11 (humanized antibody #11) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 72 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 73.

[0194] Humanized anti-CAPRIN-1 monoclonal antibody #12 (humanized antibody #12) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 74 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 75.

[0195] Humanized anti-CAPRIN-1 monoclonal antibody #13 (humanized antibody #13) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 76 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 77.

[0196] Humanized anti-CAPRIN-1 monoclonal antibody #14 (humanized antibody #14) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 78 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 79.

[0197] Humanized anti-CAPRIN-1 monoclonal antibody #15 (humanized antibody #15) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 80 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 81.

[0198] Humanized anti-CAPRIN-1 monoclonal antibody #16 (humanized antibody #16) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 82 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 83.

[0199] Humanized anti-CAPRIN-1 monoclonal antibody #17 (humanized antibody #17) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 84 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 85.

[0200] Humanized anti-CAPRIN-1 monoclonal antibody #18 (humanized antibody #18) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 86 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 87.

[0201] Humanized anti-CAPRIN-1 monoclonal antibody #19 (humanized antibody #19) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 88 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 89.

[0202] Humanized anti-CAPRIN-1 monoclonal antibody #20 (humanized antibody #20) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 90 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 91.

[0203] Humanized anti-CAPRIN-1 monoclonal antibody #21 (humanized antibody #21) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 92 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 93.

[0204] Humanized anti-CAPRIN-1 monoclonal antibody #22 (humanized antibody #22) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 94 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 95.

[0205] Humanized anti-CAPRIN-1 monoclonal antibody #23 (humanized antibody #23) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 96 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 97.

[0206] Humanized anti-CAPRIN-1 monoclonal antibody #24 (humanized antibody #24) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 98 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 99.

[0207] Humanized anti-CAPRIN-1 monoclonal antibody #25 (humanized antibody #25) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 100 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 101.

[0208] Humanized anti-CAPRIN-1 monoclonal antibody #26 (humanized antibody #26) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 102 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 103.

[0209] Humanized anti-CAPRIN-1 monoclonal antibody #27 (humanized antibody #27) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 104 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 105.

[0210] Humanized anti-CAPRIN-1 monoclonal antibody #28 (humanized antibody #28) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 106 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 107.

[0211] Humanized anti-CAPRIN-1 monoclonal antibody #29 (humanized antibody #29) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 108 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 109.

[0212] Humanized anti-CAPRIN-1 monoclonal antibody #30 (humanized antibody #30) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 110 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 111.

[0213] Humanized anti-CAPRIN-1 monoclonal antibody #31 (humanized antibody #31) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 112 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 113.

[0214] Humanized anti-CAPRIN-1 monoclonal antibody #32 (humanized antibody #32) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 114 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 115.

[0215] Humanized anti-CAPRIN-1 monoclonal antibody #33 (humanized antibody #33) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 116 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 117.

[0216] Humanized anti-CAPRIN-1 monoclonal antibody #34 (humanized antibody #34) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 118 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 119.

[0217] Humanized anti-CAPRIN-1 monoclonal antibody #35 (humanized antibody #35) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 120 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 121.

[0218] Humanized anti-CAPRIN-1 monoclonal antibody #36 (humanized antibody #36) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 122 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 123.

[0219] Humanized anti-CAPRIN-1 monoclonal antibody #37 (humanized antibody #37) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 124 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 125.

[0220] Humanized anti-CAPRIN-1 monoclonal antibody #38 (humanized antibody #38) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 126 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 127.

[0221] Humanized anti-CAPRIN-1 monoclonal antibody #39 (humanized antibody #39) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 128 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 129.

[0222] Humanized anti-CAPRIN-1 monoclonal antibody #40 (humanized antibody #40) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 130 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 131.

[0223] Humanized anti-CAPRIN-1 monoclonal antibody #41 (humanized antibody #41) comprising the amino acid sequence of a heavy chain variable region represented by the amino acid sequence of SEQ ID NO: 132 and the amino acid sequence of a light chain variable region represented by the amino acid sequence of SEQ ID NO: 133.

[0224] Additionally, among these anti-CAPRIN-1 monoclonal antibodies, a nucleotide sequence was designed based on humanized antibody #1 to express a heavy chain variable region comprising CDR1-3 consisting of the amino acid sequences of SEQ ID NO: 36, SEQ ID NO: 37, and SEQ ID NO: 38, respectively, and framework region comprising sequences of a human antibody. This nucleotide sequence was inserted into a mammalian expression vector with an inserted heavy chain constant region of human IgG1 in which serine (Ser) at amino acid position 239 by EU numbering is substituted with aspartic acid (Asp), and isoleucine (Ile) at amino acid position 332 by EU numbering is substituted with glutamic acid (Glu). Further, a nucleotide sequence was designed to express the amino acid sequence of a light chain variable region comprising CDR1-3 consisting of the amino acid sequences of SEQ ID NO: 40, SEQ ID NO: 41, and SEQ ID NO: 42, respectively, and framework region comprising sequences of a human antibody, and the nucleotide sequence was inserted into a mammalian expression vector with an inserted light chain constant region of human IgG1. These 2 recombinant expression vectors were transferred to mammalian cells according to a conventional method; and a culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #5 (humanized antibody #5) composed of the full-length heavy chain amino acid sequence consisting of the heavy chain variable region designed above and the heavy chain constant region of human IgG1 in which serine (Ser) at amino acid position 239 by EU numbering is substituted with aspartic acid (Asp), and isoleucine (Ile) at amino acid position 332 by EU numbering is substituted with glutamic acid (Glu), and the full-length light chain amino acid sequence consisting of the light chain variable region designed above and the human light chain constant region, was obtained from the cells.

[0225] A culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #6 (humanized antibody #6) composed of the amino acid sequence of the heavy chain variable region and the amino acid sequence of the light chain variable region of the humanized antibody #2 produced above was prepared in the same manner.

[0226] A culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #7 (humanized antibody #7) composed of the amino acid sequence of the heavy chain variable region and the amino acid sequence of the light chain variable region of the humanized antibody #3 produced above was prepared in the same manner. A culture supernatant containing humanized anti-CAPRIN-1 monoclonal antibody #8 (humanized antibody #8) composed of the amino acid sequence of the heavy chain variable region and the amino acid sequence of the light chain variable region of the humanized antibody #4 produced above was prepared in the same manner.

[0228] Culture supernatants containing each of humanized anti-CAPRIN-1 antibodies #42 to #74 (humanized antibodies #42 to #74) composed of the amino acid sequence of the heavy chain variable region and the amino acid sequence of the light chain variable region of each of the humanized antibodies #9 to #41 produced above were prepared in the same manner.

[0229] The obtained culture supernatants containing each of the humanized anti-CAPRIN-1 monoclonal antibodies #1 to #74 were subjected to purification using Hitrap Protein A Sepharose FF (GE Healthcare) according to a conventional method. The buffer was replaced with PBS(-). The resultant was filtered through a 0.22 gm filter (Merck Millipore Corp.) to prepare the humanized antibodies.

[0230] (Example 3) Preparation of conjugate of anti-CAPRIN-1 antibody and thiol-containing maytansinoid DM4 Conjugates of the anti-CAPRIN-1 polyclonal antibodies #1 to #6 described in Example 1 and the thiol-containing maytansinoid DM4 were prepared. Anti-CAPRIN-1 polyclonal antibody #1 was dissolved in PBS(-) (pH 8.5) at a concentration of 4 mg / ml, and DMA was added to prepare a solution containing 10% DMA. A solution of SPDB-DM4 (CAS No. 1626359-62-3, MedChemExpress) dissolved in DMA at a concentration of 10 mM was added to the above solution containing anti-CAPRIN-1 polyclonal antibody #1 in an amount corresponding to 6 molar equivalents, and the mixture was allowed to react at room temperature for 3 hours. After the reaction, the solution was replaced with PBS (pH 7.4) using a Zeba (trademark) Spin Desalting Column (MWCO 40 k) to remove free SPDB-DM4, thereby obtaining a solution containing anti-CAPRIN-1 polyclonal antibody #1-SPDB-DM4 (Conjugate 1) of the present invention.

[0231] In the same manner, conjugates were also prepared for anti-CAPRIN-1 polyclonal antibodies #2 to #6 using SPDB-DM4, and solutions containing the respective conjugates were obtained (conjugate prepared using anti-CAPRIN-1 polyclonal antibody #2: Conjugate 2; conjugate prepared using anti-CAPRIN-1 polyclonal antibody #3: Conjugate 3; conjugate prepared using anti-CAPRIN-1 polyclonal antibody #4: Conjugate 4; conjugate prepared using anti-CAPRIN-1 polyclonal antibody #5: Conjugate 5; and conjugate prepared using anti-CAPRIN-1 polyclonal antibody #6: Conjugate 6). Furthermore, using the same procedure described above, a solution containing a conjugate with SMDB-DM4 (Control conjugate 1) was obtained for the rabbit control antibody described in Example 1, which does not react with the CAPRIN-1 protein.

[0232] In addition, by the same method as described above, a solution containing a conjugate with SPDB-DM4 (Conjugate 10) was obtained using the humanized antibody #4, which is the anti-CAPRIN-1 monoclonal antibody described in Example 2.

[0233] Furthermore, a conjugate of humanized antibody #1, which is the anti-CAPRIN-1 monoclonal antibody described in Example 2, with SPDB-DM4 (Conjugate 7) was prepared in a conventional manner. Likewise, solutions containing conjugates with SPDB-DM4 were obtained for the humanized antibodies #2, #3, and #5 to #74 (Conjugates 8, 9, and 11 to 80 respectively).

[0234] The above-described conjugates were analyzed by mass spectrometry in a conventional manner to determine the number of DM4 molecules bound per antibody molecule, and the average value was approximately 4 to 7.

[0235] In addition, using the same procedure as described above, a solution containing a conjugate with SPDB-DM4 (Control conjugate 2) was also obtained for the human IgG control antibody, which does not react with the CAPRIN-1 protein.

[0236] (Example 4) Specific reactivity of conjugates against CAPRIN-1 protein and CAPRIN-1-expressing cancer cell Conjugates prepared in Example 3 (anti-CAPRIN-1 antibody-SPDB-DM4 conjugates) were assayed for their specific reactivity with the CAPRIN-1 protein and their reactivity against the cell membrane surface of human cancer cells and mouse cancer cells both expressing the CAPRIN-1 protein.

[0237] The specific reactivity with the CAPRIN-1 protein was determined by ELISA. 1 ugmL CAPRIN-1 protein solution was added at 100 p,L / well to a 96-well plate, and the plate was left at 4°C for 18 hours. Each well was washed with PBS-T three times. Then, a 0.5% bovine serum albumin (BSA) solution was added at 400 pL / well, and the plate was left at room temperature for 3 hours. The solution was removed, and the wells were washed with 400 pL / well of PBS-T three times. Then, each of the solutions containing Conjugates 1 to 6 and Control conjugate 1 was added at 100 pL / well, and the plate was left at room temperature for 2 hours. Each well was washed with PBS-T three times. Then, an HRP-labeled anti-rabbit IgG antibody diluted 5000-fold with PBS was added at 100 pL / well, and the plate was left at room temperature for 1 hour. Each well was washed with PBS-T three times. Then, a TMB substrate solution was added at 100 pL / well, and the plate was left for 15 to 30 minutes for chromogenic reaction. After the color developed, the reaction was terminated by adding 1 N sulfuric acid at 100 pL / well, and the absorbance values at 450 nm and 595 nm were measured using an absorption spectrometer. As a result, Conjugates 1 to 6 exhibited a higher absorbance value than that of Control conjugate 1 used as a negative control, indicating that these conjugates specifically react with the CAPRIN-1 protein.

[0238] Similarly, for the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates 7 to 80) prepared using the humanized antibodies #1 to #74 in Example 3, specific reactivity with the CAPRIN-1 protein was detected using HRP-labeled anti-human antibody by ELISA. As a result, these conjugates exhibited a higher absorbance value than that of human IgG (Sigma- Aldrich Corp) and Control conjugate 2 used as negative controls, indicating that these conjugates specifically react with the CAPRIN-1 protein.

[0239] Next, the reactivity against the cell membrane surface of cancer cells expressing CAPRIN-1 protein on the cell surface was verified by flow cytometry. 2 x 105 cells of human breast cancer cells BT-474 (from ATCC) and mouse breast cancer cells 4T1 (from ATCC) were centrifuged in 1.5 mL microcentrifuge tubes. 100 uL of the solutions containing each of Conjugates 1 to 6 and Control conjugate 1 was then added to separate tubes. The tube was left at 4C for 1 hour. After washing with PBS, Alexa 488-labeled anti-rabbit IgG (H + L) diluted 100-fold with PBS(-) containing 0.5% FBS (0.5% FBS-PBS(-)) was added thereto, and the tube was left at 4°C for 1 hour. After washing with 0.5% FBS-PBS(-), the cells were reacted with BD Horizon Fixable Viability Stain (FVS) Reagents (FVS450, Becton, Dickinson and Company) to stain dead cells, and the fluorescence intensity was then measured using FACSFortessa (trademark) (Becton, Dickinson and Company). As a result, the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates 1 to 6) exhibited higher fluorescence intensity than that of Control conjugate 1 used as a negative control, indicating that these conjugates strongly react against the cell surface of human breast cancer cells BT474 and mouse breast cancer cells 4T1 both expressing CAPRIN-1 protein on the cell surface.

[0240] Similarly, for the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates prepared using humanized antibodies #1 to #74, specific reactivity against the human cancer cells BT474 and the mouse cancer cells 4T1, both expressing the CAPRIN-1 protein on the cell membrane surface, was detected using Alexa 488-labeled anti-human IgG (H+L) antibody. As a result, the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates 7 to 80) prepared using humanized antibodies #1 to #74 exhibited higher fluorescence intensity than Control Conjugate 2 used as a negative control, indicating that these conjugates strongly react against the cell surface of the human breast cancer cells BT474 and the mouse breast cancer cells 4T1 both expressing the CAPRIN-1 protein on the cell surface.

[0241] Similarly, for the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates 7 to 80) prepared using the anti-CAPRIN-1 polyclonal antibody and humanized antibodies #1 to #74 in Example 3, reactivity against various human cancer cells and mouse cancer cells was examined. Against the human cancer cells in which expression of the CAPRIN-1 gene has been confirmed and expression of the CAPRIN-1 protein on the cell membrane surface has been verified, such as the breast cancer cells (BT-474, MDA-MB-453, HCC70, MCF7, MDA-MB-231, SKBR3, T47D), colorectal cancer cells (HT-29, HCT116, Lovo, SW480, DLD-1), lung cancer cells (A549, NCI-H322, NCI-H2228), gastric cancer cells (NCI-N87, NUGC3), uterine cancer cells (HEC-1-A), prostate cancer cells (22Rv1), pancreatic cancer cells (Panc10.5), liver cancer cells (HepG2), ovarian cancer cells (MCAS, SKOV3, and TOV112D), kidney cancer cells (Caki-2), brain tumor cells (U-87MG), bladder cancer cells (T24, UMUC3, RT4, 5637), bile duct cancer cells (KKU213), fibrosarcoma cells (HT-1080), esophageal cancer cells (OE33), leukemia cells (OCI-AML5), lymphoma cells (Ramos), gallbladder cancer cells (TGBC14TKB), melanoma cells (Malme-3M), and head and neck cancer cells (FaDu), and mouse kidney cancer cells (Renca) and mouse breast cancer cells (4T1) in which expression of the CAPRIN-1 gene has been confirmed, all the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates prepared in Example 3 exhibited stronger fluorescence intensity than Control conjugate 1 and Control conjugate 2, both used as negative controls, indicating that these conjugates strongly react against the cell membrane surface of the CAPRIN-1-expressing cancer cells.

[0242] (Example 5) Internalization of DM4 conjugates Internalization of the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates 1 to 80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3, into cancer cells was examined using a confocal laser microscope. The anti-CAPRIN-1 antibody-SPDB-DM4 conjugate was added at 100 ugmL to cancer cells HT29 that had been cultured overnight in a culture plate, and allowed to react on ice for 1 hour. The cells were then reacted at 37°C under 5% CO2 for 1 to 6 hours. After reaction, the cells were washed with PBS and fixed at room temperature using PBS containing 4% paraformaldehyde. After fixation, the cells were washed with PBS and then the cell membrane was permeabilized using PBS containing 0.3% Triton X-100. Following permeabilization of the cell membrane, the cells were washed with PBS and subjected to a blocking treatment with PBS containing 5% BSA. After blocking, Alexa 488-labeled anti-rabbit IgG secondary antibody or anti-human IgG secondary antibody (A11013, Invitrogen) was added thereto at a concentration of 10 pg ml and allowed to react at room temperature for 1 hour. After reaction with the secondary antibody, the cells were washed with PBS, and the thus obtained samples were observed using a confocal laser microscope (Nikon A1R inverted microscopy) to examine internalization of the conjugates of the present invention into cancer cells. As a result of the observation, the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 were clearly present inside the cells, indicating that these conjugates of the present invention underwent internalization. In contrast, no internalization into cancer cells was observed for Control conjugates 1 and 2 prepared in Example 3.

[0243] (Example 6) Antitumor activity of DM4 conjugates The antitumor activity against cancer cells of the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 was evaluated in vitro. Against human cancer cells in which expression of CAPRIN-1 on the cell membrane surface has been confirmed in Example 4, including breast cancer cells (BT-474), colorectal cancer cells (HT-29), lung cancer cells (NCI-H322, NCI-H2228), uterine cancer cells (Ishikawa, HEC-1-A), pancreatic cancer cells (Panc10.5), ovarian cancer cells (MCAS, TOV112D), head and neck cancer cells (FaDu), gastric cancer cells (NU-GC-3), bladder cancer cells (UMUC3), prostate cancer cells (22Rv1), liver cancer cells (HepG2), kidney cancer cells (Caki-2), brain tumor cells (U-87MG), bile duct cancer cells (KKU213), fibrosarcoma cells (HT-1080), gallbladder cancer cells (TGBC14TKB), and melanoma cells (Malme-3M), the antitumor activity was evaluated.

[0244] The cancer cells cultured by standard methods were seeded in a cell-culturable black 96-well plate (96 well optical Btm Plt PolymerBase Black w / Lid Cell Culture Sterile PS 165305, Thermo Fischer Scientific, Inc.) at 90 gL / well with 1,000 to 5,000 cells per well and cultured at 37°C under 5% CO2 for 18 to 24 hours in an incubator. Thereafter, the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 were added at 10 gL / well to have a concentration of 10-2 gg / mL to 102 gg / mL and incubated at 37°C under 5% CO2 in an incubator. Cell survival rates were measured 3 days or 4 days after the start of the incubation using the Cell Titer Glow Luminescent Cell Viability Assay (#7573, Promega). Substrate was added at 100 gL / well, and the plate was shaken for 2 minutes using a plate shaker and left to stand for 10 minutes. Then the luminescence signal intensity was measured using SpectraMax (registered trademark) iD3 (Molecular Devices, LLC.) to calculate the ATP content of surviving cells.

[0245] As a result, when the number of the cancer cells survived in wells without addition of the conjugate were defined as 100%, the survival rate of cancer cells treated with the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates (Conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 of the present invention were 70% or less at conjugate concentrations of 1 gg / mL or higher. These results demonstrate that the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates exhibit antitumor activity against cancer cells.

[0246] (Example 7) Antitumor effect of DM4 conjugates on tumor-bearing mouse (1) For Conjugate 1 and Conjugate 7, which were anti-CAPRIN-1 antibody-SPDB-DM4 conjugates prepared in Example 3 (in both of which approximately 4 molecules of DM4 were bound per antibody molecule), their in vivo antitumor effects were evaluated in tumor-bearing mice. Using NOD-SCID mice transplanted with human-derived cancer cells expressing the CAPRIN-1 protein, the antitumor effects of the conjugates of the present invention were examined.

[0247] Approximately 107 cells of human head and neck cancer cell line FaDu were mixed with Matrigel (SIGMA) and transplanted subcutaneously into each mouse. To tumor-bearing mice in which the average tumor volume had grown to 250 mm3 or more, Conjugate 7 and Conjugate 10 were each administered once at 10 mg / kg via the tail vein. FaDu expresses the CAPRIN-1 protein on the cell membrane surface, and as shown in Example 4, Conjugate 1 and Conjugate 7 each bind to FaDu.

[0248] The tumor sizes of the tumor-bearing mice after the administration were measured using calipers over time, and tumor volumes were calculated according to an ordinary method based on the formula: (Length of the major axis of tumor) x (Length of the minor axis of tumor)2 x 0.5. As a result of the evaluation, all the mice receiving Conjugates 1 and 7 had tumor volumes of less than 7 to 50% at 14 days after the start of the administration when the tumor volume of the negative control was set at 100%.

[0249] Comparable antitumor effects were also obtained by the other anti-CAPRIN-1 antibody-SPDB-DM4 conjugates prepared in Example 3.

[0250] These results demonstrated that the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates prepared in Example 3 exert a markedly stronger antitumor effect than the negative control (untreated group).

[0251] (Example 8) Antitumor effect of DM4 conjugates on tumor-bearing mouse (2) For Conjugate 1 and Conjugate 7, which were anti-CAPRIN-1 antibody-SPDB-DM4 conjugates prepared in Example 3 (in both of which approximately 4 molecules of DM4 were bound per antibody molecule), their in vivo antitumor effects were evaluated in tumor-bearing mice. Using Balb / c nude mice transplanted with human-derived cancer cells expressing the CAPRIN-1 protein, the antitumor effects of the conjugates of the present invention were examined.

[0252] Human gastric cancer cells NUGC3 were mixed with Matrigel (SIGMA) and transplanted subcutaneously into each mouse. To tumor-bearing mice in which the average tumor volume had grown to 100 mm3 or more, Conjugate 1 and Conjugate 7 were each administered at 10 mg / kg via the tail vein. NUGC3 expresses the CAPRIN-1 protein on the cell membrane surface, and as shown in Example 4, Conjugate 1 and Conjugate 7 each bind to NUGC.

[0253] In addition, in accordance with the method described in Example 3, a conjugate of trastuzumab, an antibody that is not an anti-CAPRIN-1 antibody, and SPDB-DM4 (in which approximately 4 molecules of DM4 were bound per antibody molecule) was prepared and administered at the same dose to the above-described tumor-bearing mice as a comparative control. NUGC3 expresses the HER2 protein, which is the target antigen of trastuzumab, on the cell membrane surface, and the trastuzumab-SPDB-DM4 conjugate exhibited reactivity against NUGC3.

[0254] The tumor sizes of the tumor-bearing mice after the administration were measured using calipers over time, and tumor volumes were calculated according to an ordinary method based on the formula: (Length of the major axis of tumor) x (Length of the minor axis of tumor)2 x 0.5. As a result of the evaluation, all the mice receiving Conjugates 1 and 7 had tumor volumes ranging from complete regression (0%) to less than 50% at 20 days or later after the start of the administration when the tumor volume in mice receiving the anti-CAPRIN-1 antibody without DM4 conjugation (comparative control) was set at 100%.

[0255] On the other hand, the mice receiving the trastuzumab-SPDB-DM4 conjugate had a tumor volume of 80% or more when the tumor volume of mice receiving trastuzumab without DM4 conjugation was set at 100%.

[0256] As a result of this evaluation, it was demonstrated that the antitumor effects of the anti-CAPRIN-1 antibody-SPDB-DM4 conjugates are markedly enhanced compared with the anti-CAPRIN-1 antibody without DM4 conjugation (comparative control), whereas the enhancement of the pharmacological effect of the trastuzumab-DM4 conjugate relative to trastuzumab without DM4 conjugation is limited.

[0257] (Example 9) Preparation of conjugate of anti-CAPRIN-1 antibody and thiol-containing maytansinoid DM1 Conjugates of the anti-CAPRIN-1 polyclonal antibodies #1 to #6 described in Example 1 with the thiol-containing maytansinoid DM1 were prepared. Anti-CAPRIN-1 polyclonal antibody #1 was dissolved in PBS(-) (pH 7.2) at a concentration of 2.5 mg / ml. A solution of SMCC-DM1 (CAS No. 1228105-51-8, MedChemExpress) dissolved in DMSO at a concentration of 1 mM was added to the above solution containing anti-CAPRIN-1 polyclonal antibody #1 and 19% (v / v) DMSO in an amount corresponding to 7.5 to 12 molar equivalents, and the mixture was allowed to react at 33°C for 1 hour. After the reaction, the solution was replaced with PBS (pH 7.2) using Illustra NAP-10 Columns (GE Healthcare, Cat. No. 17085402) to remove free SMCC-DM1, thereby obtaining a solution containing anti-CAPRIN-1 polyclonal antibody #1-SMCC-DM1 (DM1 conjugate 1) of the present invention.

[0258] In the same manner, conjugates were also prepared for anti-CAPRIN-1 polyclonal antibodies #2 to #6 using SMCC-DM1, and solutions containing the respective DM1 conjugates were obtained (conjugate prepared using anti-CAPRIN-1 polyclonal antibody #2: DM1 conjugate 2, conjugate prepared using anti-CAPRIN-1 polyclonal antibody #3: DM1 conjugate 3, conjugate prepared using anti-CAPRIN-1 polyclonal antibody #4: DM1 conjugate 4, conjugate prepared using anti-CAPRIN-1 polyclonal antibody #5: DM1 conjugate 5, and conjugate prepared using anti-CAPRIN-1 polyclonal antibody #6: DM1 conjugate 6). Furthermore, in the same manner as described above, a solution containing a conjugate with SMCC-DM1 (DM1 control conjugate 1) was obtained for the rabbit control antibody described in Example 1, which does not react with the CAPRIN-1 protein.

[0259] In addition, by the same method as described above, a solution containing a conjugate with SMCC-DM1 (DM1 conjugate 10) was obtained using the humanized antibody #4, which is the anti-CAPRIN-1 monoclonal antibody described in Example 2.

[0260] Furthermore, a conjugate of humanized antibody #1, which is the anti-CAPRIN-1 monoclonal antibody described in Example 2, with SMCC-DM1 (DM1 conjugate 7) was prepared in a conventional manner. Likewise, solutions containing conjugates with SMCC-DM1 were obtained for the humanized antibodies #2, #3, and #5 to #74 (DM1 conjugate 8, DM1 conjugate 9, and DM1 conjugates 11 to 80 respectively).

[0261] The above-described conjugates were analyzed by mass spectrometry in a conventional manner to determine the number of DM1 molecules bound per antibody molecule, and the average value was approximately 4 to 6.

[0262] In addition, using the same procedure as described above, a solution containing a conjugate with SMCC-DM1 (DM1 control conjugate 2) was also obtained for the human IgG control antibody, which does not react with the CAPRIN-1 protein.

[0263] (Example 10) Specific reactivity of DM1 conjugates against CAPRIN-1 protein and CAPRIN-1-expressing cancer cell DM1 conjugates prepared in Example 9 (anti-CAPRIN-1 antibody-SMCC-DM1 conjugates) were assayed for their specific reactivity with the CAPRIN-1 protein and their reactivity against the cell membrane surface of human cancer cells and mouse cancer cells both expressing the CAPRIN-1 protein.

[0264] The specific reactivity with the CAPRIN-1 protein was determined by ELISA. 1 pg / mL CAPRIN-1 protein solution was added at 100 pL / well to a 96-well plate, and the plate was left at 4°C for 18 hours. Each well was washed with PBS-T three times. Then, a 0.5% bovine serum albumin (BSA) solution was added at 400 pL / well, and the plate was left at room temperature for 3 hours. The solution was removed, and the wells were washed with 400 pL / well of PBS-T three times. Then, each of the solutions containing the conjugates 1 to 6 and the control conjugate 1 was added at 100 pLwell, and the plate was left at room temperature for 2 hours. Each well was washed with PBS-T three times. Then, an HRP-labeled anti-rabbit IgG antibody diluted 5000-fold with PBS was added at 100 pL / well, and the plate was left at room temperature for 1 hour. Each well was washed with PBS-T three times. Then, a TMB substrate solution was added at 100 pL / well, and the plate was left for 15 to 30 minutes for chromogenic reaction. After the color developed, the reaction was terminated by adding 1 N sulfuric acid at 100 pL / well, and the absorbance values at 450 nm and 595 nm were measured using an absorption spectrometer. As a result, the conjugates 1 to 6 exhibited a higher absorbance value than that of the control conjugate 1 used as a negative control, indicating that these conjugates specifically react with the CAPRIN-1 protein.

[0265] Similarly, for the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugates 7 to 80) prepared using the humanized antibodies #1 to #74 in Example 3, specific reactivity with the CAPRIN-1 protein was detected using an HRP-labeled anti-human antibody by ELISA. As a result, these conjugates exhibited a higher absorbance value than those of the human IgG (SIGMA) and DM1 control conjugate 2 used as negative controls, indicating that these conjugates specifically react with the CAPRIN-1 protein.

[0266] Next, the reactivity against the cell membrane surface of cancer cells expressing CAPRIN-1 protein on the cell surface was verified by flow cytometry. 2 x 105 cells of human breast cancer cells BT-474 (from ATCC) and mouse breast cancer cells 4T1 (from ATCC) were centrifuged in 1.5 ml microcentrifuge tubes. 100 pL of the solutions containing each of DM1 conjugates 1 to 6 and DM1 control conjugate 1 was then added to separate tubes. The tube was left at 4°C for 1 hour. After washing with PBS, Alexa 488-labeled anti-rabbit IgG (H + L) diluted 100-fold with PBS(-) containing 0.5% FBS (0.5% FBS-PBS(-)) was added thereto, and the tube was left at 4°C for 1 hour. After washing with 0.5% FBS-PBS(-), the cells were reacted with BD Horizon Fixable Viability Stain (FVS) Reagents (FVS450, Becton, Dickinson and Company) to stain dead cells, and the fluorescence intensity was then measured using FACSFortessa (trademark) (Becton, Dickinson and Company). As a result, the anti-CAPRIN- 1 antibody-SMCC-DM1 conjugates (conjugates 1 to 6) exhibited higher fluorescence intensity than that of the control conjugate 1 used as a negative control, indicating that these conjugates strongly react against the cell surface of the human breast cancer cells BT474 and mouse breast cancer cells 4T1 both expressing CAPRIN-1 protein on the cell surface.

[0267] Similarly, for the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates prepared using the humanized antibodies #1 to #74, specific reactivity against human cancer cells BT474 and mouse cancer cells 4T1, both expressing the CAPRIN-1 protein on the cell membrane surface, was detected using Alexa 488-labeled anti-human IgG (H + L) antibody. As a result, the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugate 7 to DM1 conjugate 80) prepared using humanized antibodies #1 to #74 exhibited higher fluorescence intensity than that of DM1 control conjugate 2 used as a negative control, indicating that these conjugates strongly react against the cell surface of the human breast cancer cells BT474 and mouse breast cancer cells 4T1 both expressing CAPRIN-1 protein on the cell surface.

[0268] Similarly, for the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugates 7 to 80) prepared using the anti-CAPRIN-1 polyclonal antibody and humanized antibodies #1 to #74 in Example 3, reactivity against various human cancer cells and mouse cancer cells was examined. Against the human cancer cells in which expression of the CAPRIN-1 gene has been confirmed and expression of the CAPRIN-1 protein on the cell membrane surface has been verified, such as the breast cancer cells (BT-474, MDA-MB-453, HCC70, MCF7, MDA-MB-231, SKBR3, T47D), colorectal cancer cells (HT-29, HCT116, Lovo, SW480, DLD-1), lung cancer cells (A549, NCI-H322, NCI-H2228), gastric cancer cells (NCI-N87, NUGC3), uterine cancer cells (HEC-1-A), prostate cancer cells (22Rv1), pancreatic cancer cells (Panc10.5), liver cancer cells (HepG2), ovarian cancer cells (MCAS, SKOV3, and TOV112D), kidney cancer cells (Caki-2), brain tumor cells (U-87MG), bladder cancer cells (T24, UMUC3, RT4, 5637), bile duct cancer cells (KKU213), fibrosarcoma cells (HT-1080), esophageal cancer cells (OE33), leukemia cells (OCI-AML5), lymphoma cells (Ramos), gallbladder cancer cells (TGBC14TKB), melanoma cells (Malme-3M), and head and neck cancer cells (FaDu), and mouse kidney cancer cells (Renca) and mouse breast cancer cells (4T1) in which expression of the CAPRIN-1 gene has been confirmed, all the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates prepared in Example 3 exhibited stronger fluorescence intensity than those of DM1 control conjugate 1 and DM1 control conjugate 2, both used as negative controls, indicating that these conjugates strongly react against the cell membrane surface of the CAPRIN-1 expressing cancer cells.

[0269] (Example 5) Internalization of DM1 conjugates Internalization of the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugates 1 to 80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3, into cancer cells was examined using a confocal laser microscope. The anti-CAPRIN-1 antibody-SMCC-DMl conjugate was added at 100 ag mL to the cancer cells HT29 that had been cultured overnight in a culture plate, and allowed to react on ice for 1 hour. The cells were then reacted at 37°C under 5% CO2 for 1 to 6 hours. After reaction, the cells were washed with PBS and fixed at room temperature using PBS containing 4% paraformaldehyde. After fixation, the cells were washed with PBS and then the cell membrane was permeabilized using PBS containing 0.3% Triton X-100. Following permeabilization of the cell membrane, the cells were washed with PBS and subjected to a blocking treatment with PBS containing 5% BSA. After blocking, Alexa 488-labeled antirabbit IgG secondary antibody or anti-human IgG secondary antibody (A11013, Invitrogen) was added thereto at a concentration of 10 gg / ml and allowed to react at room temperature for 1 hour. After reaction with the secondary antibody, the cells were washed with PBS, and the thus obtained samples were observed using a confocal laser microscope (Nikon A1R inverted microscopy) to examine internalization of the conjugates of the present invention into cancer cells. As a result of the observation, the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 were clearly present inside the cells, indicating that these conjugates of the present invention underwent internalization. In contrast, no internalization into cancer cells was observed for DM1 control conjugates 1 and 2 prepared in Example 3.

[0270] (Example 6) Antitumor activity of DM1 conjugates The antitumor activity against cancer cells of the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (Conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 was evaluated in vitro. Against human cancer cells in which expression of CAPRIN-1 on the cell membrane surface has been confirmed in Example 4, including breast cancer cells (BT-474), colorectal cancer cells (HT-29), lung cancer cells (NCI-H322, NCI-H2228), uterine cancer cells (Ishikawa, HEC-1-A), pancreatic cancer cells (Panc10.5), ovarian cancer cells (MCAS, TOV112D), head and neck cancer cells (FaDu), gastric cancer cells (NU-GC-3), bladder cancer cells (UMUC3), prostate cancer cells (22Rv1), liver cancer cells (HepG2), kidney cancer cells (Caki-2), brain tumor cells (U-87MG), bile duct cancer cells (KKU213), fibrosarcoma cells (HT-1080), gallbladder cancer cells (TGBC14TKB), and melanoma cells (Malme-3M), the antitumor activity was evaluated.

[0271] The cancer cells cultured by standard methods were seeded in a cell-culturable black 96-well plate (96 well optical Btm Plt PolymerBase Black w / Lid Cell Culture Sterile PS 165305, Thermo Fischer Scientific, Inc.) at 90 gL / well with 1,000 to 5,000 cells per well and cultured at 37°C under 5% CO2 for 18 to 24 hours in an incubator. Thereafter, the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 were added at 10 gL / well to have a concentration of 10-2 gg / mL to 102 gg / mL and incubated at 37°C under 5% CO2 in an incubator. Cell survival rates were measured 3 days or 4 days after the start of the incubation using the Cell Titer Glow Luminescent Cell Viability Assay (#7573, Promega). Substrate was added at 100 gL / well, and the plate was shaken for 2 minutes using a plate shaker and left to stand for 10 minutes. Then the luminescence signal intensity was measured using SpectraMax (registered trademark) iD3 (Molecular Devices, LLC.) to calculate the ATP content of surviving cells.

[0272] As a result, when the number of the cancer cells survived in wells without addition of the conjugate were defined as 100%, the survival rate of cancer cells treated with the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates (DM1 conjugates #1 to #80) prepared using the anti-CAPRIN-1 polyclonal antibodies and humanized antibodies #1 to #74 in Example 3 of the present invention were 80% or less at conjugate concentrations of 5 pg / mL or higher. These results demonstrate that the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates exhibit antitumor activity against cancer cells.

[0273] (Example 7) Antitumor effect of DM1 conjugates on tumor-bearing mouse For DM1 conjugate 1 and DM1 conjugate 7, which were anti-CAPRIN-1 antibody-SMCC-DM1 conjugates prepared in Example 3 (in both of which approximately 4 molecules of DM1 were bound per antibody molecule), their in vivo antitumor effects were evaluated in tumor-bearing mice. Using NOD-SCID mice transplanted with human-derived cancer cells expressing the CAPRIN-1 protein, the antitumor effects of the conjugates of the present invention were examined.

[0274] Human colorectal cancer cell line HT29 were mixed with Matrigel (SIGMA) and transplanted subcutaneously into each mouse. To tumor-bearing mice in which the average tumor volume had grown to 50 mm3 or more, DM1 conjugate 1 and DM1 conjugate 7 were each administered at 10 mg / kg via the tail vein. HT29 expresses the CAPRIN-1 protein on the cell membrane surface, and, as shown in Example 4, DM1 conjugate 1 and DM1 conjugate 7 each bind to HT29.

[0275] In addition, in accordance with the method described in Example 3, a conjugate of trastuzumab, which is not an anti-CAPRIN-1 antibody, with SMCC-DM1 (in which approximately 4 molecules of DM1 were bound per antibody molecule) was prepared and administered at the same dose to the above-described tumor-bearing mice as a comparative control. HT29 expresses the HER2 protein, which is the target antigen of trastuzumab, on the cell membrane surface, and the trastuzumab-SMCC-DM1 conjugate exhibited reactivity against HT29.

[0276] The tumor sizes of the tumor-bearing mice after the administration were measured using calipers over time, and tumor volumes were calculated according to an ordinary method based on the formula: (Length of the major axis of tumor) x (Length of the minor axis of tumor)2 x 0.5. As a result of the evaluation, all the mice receiving DM1 conjugates 1 and 7 had tumor volumes of less than 65% at 14 days or later after the start of the administration when the tumor volume of the anti-CAPRIN-1 antibody without DM1 conjugation (comparative control) was set at 100%.

[0277] On the other hand, the mice receiving the trastuzumab-SMCC-DM1 conjugate had tumor volumes of 80% or more when the tumor volume of mice receiving trastuzumab without DM1 conjugation was set at 100%.

[0278] As a result of this evaluation, it was demonstrated that the antitumor effects of the anti-CAPRIN-1 antibody-SMCC-DM1 conjugates are markedly enhanced compared with the anti-CAPRIN-1 antibody without DM1 conjugation (comparative control), whereas the enhancement of the pharmacological effect of the trastuzumab-DM1 conjugate relative to trastuzumab without DM1 conjugation is limited.

Claims

1. A conjugate of an antibody or a fragment thereof linked to a thiol-containing maytansinoid, wherein the antibody or the fragment thereof has immunological reactivity with a CAPRIN-1 protein having an amino acid sequence represented by any of even-numbered SEQ ID NOs among SEQ ID NOs: 2 to 30 or an amino acid sequence having 80% or more sequence identity to said amino acid sequence.

2. The conjugate according to claim 1, wherein the antibody or the fragment thereof has immunological reactivity with a partial polypeptide of the CAPRIN-1 protein, wherein the partial polypeptide has an amino acid sequence represented by any of SEQ ID NOs: 31 to 35 and 296 to 299, 308, and 309 or an amino acid sequence having 80% or more sequence identity to said amino acid sequence.

3. The conjugate according to claim 1, wherein the antibody is a monoclonal antibody or a polyclonal antibody.

4. The conjugate according to claim 1, wherein the antibody or the fragment thereof is any of the following (A) to (M):(A) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 36, 37, and 38 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 40, 41, and 42 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(B) an antibody or a fragment thereof comprising a heavy chain variable region comprisingcomplementarity-determining regions of SEQ ID NOs: 44, 45, and 46 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 48, 49, and 50 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(C) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 52, 53, and 54 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 56, 57, and 58 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(D) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 60, 61, and 62 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 64, 65, and 66 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(E) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 170, 171, and 172 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 173, 174, and 175 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(F) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 176, 177, and 178 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 179, 180, and 181 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(G) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 182, 183, and 184 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 185, 186, and 187 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(H) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 188, 189, and 190 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 191, 192, and 193 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(I) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 146, 147, and 148 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 149, 150, and 151 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(J) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 272, 273, and 274 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 275, 276, and 277 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(K) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 290, 291, and 292 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 293, 294, and 295 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein,(L) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 301, 302, and 303 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 305, 306, and 307 (CDR1, CDR2, and CDR3, respectively), and having immunological reactivity with the CAPRIN-1 protein, and(M) an antibody or a fragment thereof comprising a heavy chain variable region comprising complementarity-determining regions of SEQ ID NOs: 134, 135, and 136 (CDR1, CDR2, and CDR3, respectively), and a light chain variable region comprising complementarity-determining regions of SEQ ID NOs: 137, 138, and 139 (CDR1, CDR2, and CDR3, respectively), and havingimmunological reactivity with the CAPRIN-1 protein.

5. The conjugate according to claim 1, wherein the antibody or the fragment thereof is any of the following (a) to (al):(a) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 39 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 43,(b) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 51,(c) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 55 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 59,(d) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 63 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 67,(e) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 68 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 69,(f) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 70 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 71,(g) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 72 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 73,(h) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 74 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 75,(i) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 76 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 77,(j) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 78 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 79,(k) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 80 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 81,(l) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 82 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 83,(m) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 84 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 85,(n) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 86 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 87,(o) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 88 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 89,(p) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 90 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 91,(q) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 92 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 93,(r) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 94 and a light chain variable region comprising the aminoacid sequence of SEQ ID NO: 95,(s) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 96 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 97,(t) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 98 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 99,(u) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 100 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 101,(v) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 102 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 103,(w) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 104 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 105,(x) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 106 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 107,(y) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 108 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 109,(z) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 110 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 111,(aa) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 112 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 113,(ab) an antibody or a fragment thereof, comprising a heavy chain variable region comprising theamino acid sequence of SEQ ID NO: 114 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 115,(ac) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 116 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 117,(ad) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 118 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 119,(ae) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 120 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 121,(af) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 122 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 123,(ag) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 124 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 125,(ah) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 126 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 127,(ai) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 128 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 129,(aj) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 130 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 131,(ak) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 132 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 133, and(al) an antibody or a fragment thereof, comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 300 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 304.

6. The conjugate according to claim 1, wherein the antibody is a human antibody, a humanized antibody, a chimeric antibody, or a single-chain antibody.

7. The conjugate according to claim 1, wherein the antibody or the fragment thereof is linked to the thiol-containing maytansinoid via a linker.

8. The conjugate according to claim 1, wherein the thiol-containing maytansinoid is DM1, DM4, or a derivative thereof.

9. A pharmaceutical composition for treatment and / or prevention of a cancer, comprising the conjugate according to any one of claims 1 to 8 as an active ingredient.

10. The pharmaceutical composition according to claim 9, wherein the cancer is a cancer expressing a CAPRIN-1 protein on the cell membrane surface.

11. The pharmaceutical composition according to claim 10, wherein the cancer is breast cancer, kidney cancer, pancreatic cancer, colorectal cancer, lung cancer, brain tumor, gastric cancer, uterine cancer, ovarian cancer, prostate cancer, bladder cancer, esophageal cancer, leukemia, lymphoma, liver cancer, gallbladder cancer, bile duct cancer, sarcoma, mastocytoma,melanoma, adrenocortical carcinoma, Ewing's tumor, Hodgkin's lymphoma, mesothelioma, multiple myeloma, testicular cancer, thyroid cancer, head and neck cancer, or urothelial carcinoma.

12. A method for treating and / or preventing a cancer, comprising administering the conjugate according to any one of claims 1 to 8 or the pharmaceutical composition according to any one of claims 9 to 11 to a subject.