N-Acetylgalactosylamino Dendrimer Scavengers for DOTA-Pretargeted Radioimmunotherapy

Abstract

The present disclosure provides compositions and methods for treating cancer. Specifically, the compositions of the present technology include novel clearing agents that can be used in pretargeted radioimmunotherapy.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of and priority to U.S. Provisional Application No. 62 / 697,956, filed on July 13, 2018, the disclosure of which is incorporated herein by reference in its entirety. Technical Field

[0003] The present technology generally relates to compositions comprising novel N-acetylgalactosamine dendrimer scavengers and methods of using the same in pretargeted radioimmunotherapy.

[0004] Government Support Statement

[0005] This invention was made with U.S. Government support under CA86438 awarded by the National Institutes of Health. The U.S. Government has certain rights in this invention. Background Art

[0006] The following description of the technical background of the present invention is only provided to help understand the present invention, and it is not admitted that the description describes or constitutes the prior art for the present invention.

[0007] Radiolabeled agents have been used as vehicles for delivering ionizing radiation to specific disease sites for more than 50 years (Larson SM. Cancer 67: 1253-1260 (1991); Britton KE. Nucl Med Commun. 18: 992-1007 (1997)). A large number of molecules have been considered for targeted delivery of radioisotopes, including radiolabeled antibodies, antibody fragments, alternative scaffolds, and small molecules (Tolmachev V et al. Cancer Res. 67: 2773-2782 (2007); Birchler MT et al., Otolaryngol Head Neck Surg. 136: 543-548 (2007); Reubi JC, Maecke HR. J Nucl Med. 49: 1735-1738 (2008)). Partly due to suboptimal tumor dose and therapeutic index (TI), using antibodies to target toxins to tumors, such as radioimmunotherapy (RIT) with directly conjugated antibodies, has always been challenging. In addition, due to bystander toxicity of normal tissues, dose escalation is not feasible, so this therapy results in limited anti-tumor effects. In addition, antibodies exhibit a long half-life in the blood, resulting in a low tumor to background ratio. Antibody fragments and other smaller binding scaffolds exhibit faster blood clearance, but result in high kidney and / or liver uptake. Compared with antibodies and antibody fragments, radiolabeled small molecule ligands generally exhibit faster blood clearance and lower background, but due to relatively low affinity for the desired target, poor specificity is generally achieved.

[0008] In pretargeted radioimmunotherapy (PRIT), a non-radioactive bifunctional antibody specific for both a tumor antigen and a small molecule hapten is administered and allowed to localize to one or more tumors. After adequate blood clearance of the antibody, the radiolabeled small molecule is administered and captured by the pretargeted antibody. However, many small peptides and metal chelated haptens used in the PRIT system exhibit significant systemic retention, which results in undesirable background activity, limiting the signal-to-background ratio for imaging; and contributes to nonspecific radiation, limiting the maximum tolerated dose for therapeutic applications (Orcutt et al., Mol Imaging Biol 13:215-221 (2011)).

[0009] Therefore, there is a need for novel molecules that allow for (a) highly efficient pretargeted radioimmunotherapy of solid tumors in vivo and (b) rapid clearance of radiolabeled small molecules from non-tumor tissues. Summary of the invention

[0010] In one aspect, the present invention provides a compound, which is

[0011]

[0012]

[0013]

[0014]

[0015] or a pharmaceutically acceptable salt and / or solvate thereof, wherein X 1 , X 2 , X 3 , X 4 , X 5 , X 6 , X 7 , X 8 , X 9 , X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 , X 17 , X 18 , X 19 and X 20 Each independently is H or a lone pair of electrons (i.e., providing an oxygen anion); Y 1 , Y 2 , Y 3 , Y 4 and Y 5 are each independently O or S; x is 1, 2 or 3; and y is 1, 2, 3 or 4.

[0016] In any of the embodiments herein, the compound may be a scavenger, the scavenger being

[0017]

[0018]

[0019]

[0020]

[0021] or a pharmaceutically acceptable salt and / or solvate thereof, wherein M 1 、M 2 、M 3 、M 4 and M 5 Each independently is Lu 3+ Sc 3+ , Ga3+ , Y 3+ 、In 3+ ,La 3+ 、Ce 3+ 、Eu 3+ , Tb 3+ or Gd 3+ ;X 1 , X 2 , X 3 , X 4 , X 5 , X 6 , X 7 , X 8 , X 9 , X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 , X 17 , X 18 , X 19 and X 20 Each independently is H or a lone pair of electrons (i.e., providing an oxygen anion); Y 1 , Y 2 , Y 3 , Y 4 and Y 5 are each independently O or S; x is 1, 2 or 3; and y is 1, 2, 3 or 4.

[0022] In any of the embodiments herein, M 1 、M 2 、M 3 、M 4 and M 5 In any embodiment herein, Y 1 , Y 2 , Y 3 , Y 4 and Y 5 and x may each independently be S. In any embodiment herein, x may be 1 or 2. In any embodiment herein, y may be 2 or 3.

[0023] In a related aspect, a composition is provided, comprising one or more of any embodiments of the compounds described above and a pharmaceutically acceptable carrier.

[0024] In another aspect, the present disclosure provides a method for increasing the sensitivity of a tumor to radiation therapy in a subject diagnosed with cancer, the method comprising (a) administering to the subject an effective amount of an anti-DOTA bispecific antibody, wherein the anti-DOTA bispecific antibody is configured to localize to a tumor expressing a tumor antigen target; (b) administering to the subject an effective amount of a clearing agent of the present technology; and (c) administering to the subject an effective amount of a radiolabeled DOTA hapten, wherein the DOTA hapten is configured to form a complex with the anti-DOTA bispecific antibody.

[0025] In another aspect, the present disclosure provides a method for treating cancer in a subject in need thereof, the method comprising (a) administering to the subject an effective amount of an anti-DOTA bispecific antibody, wherein the anti-DOTA bispecific antibody is configured to localize to a tumor expressing a tumor antigen target; (b) administering to the subject an effective amount of a clearing agent of the present technology; and (c) administering to the subject an effective amount of a radiolabeled DOTA hapten, wherein the DOTA hapten is configured to form a complex with the anti-DOTA bispecific antibody. The method for treating cancer may also include sequentially, separately or simultaneously administering to the subject at least one chemotherapeutic agent selected from the group consisting of nitrogen mustards, ethyleneimine derivatives, alkyl sulfonates, nitrosoureas, gemcitabine, triazenes, folic acid analogs, anthracyclines, taxanes, COX-2 inhibitors, pyrimidine analogs, purine analogs, antibiotics, enzyme inhibitors, epipodophyllotoxins, platinum coordination complexes, vinca alkaloids, substituted ureas, methylhydrazine derivatives, adrenocortical inhibitors, hormone antagonists, endostatin, paclitaxel, camptothecin, SN-38, doxorubicin, doxorubicin analogs, antimetabolites, alkylating agents, antimitotic agents, antiangiogenic agents, tyrosine kinase inhibitors, mTOR inhibitors, heat shock protein (HSP90) inhibitors, proteasome inhibitors, HDAC inhibitors, pro-apoptotic agents, methotrexate and CPT-11.

[0026] Additionally or alternatively, in some embodiments of the methods disclosed herein, the tumor antigen target is selected from GPA33, HER2 / neu, GD2, MAGE-1, MAGE-3, BAGE, GAGE-1, GAGE-2, MUM-1, CDK4, N-acetylglucosamine transferase, p15, gp75, β-catenin, ErbB2, cancer antigen 125 (CA-125), carcinoembryonic antigen (CEA), RAGE, MART (melanoma antigen), MUC-1, MUC-2, MUC-3, MUC-4, MUC-5ac, MUC-16, MUC-17, tyrosinase, Pmel 17 (gp100), GnT-V intron V sequence (N-acetylglucosamine transferase V intron V sequence), prostate cancer psm, PRAME (melanoma antigen), β-catenin, EBNA (Epstein-Barr virus nuclear antigen) 1-6, p53, lung resistance protein (LRP) Bcl-2, prostate specific antigen (PSA), Ki-67, CEACAM6, colon specific antigen-p (CSAp), HLA-DR, CD40, CD74, CD138, EGFR, EGP-1, EGP-2, VEGF, PlGF, insulin-like growth factor (ILGF), tenascin, platelet-derived growth factor, IL-6, CD20, CD19, PSMA, CD33, CD123, MET, DLL4, Ang-2, HER3, IGF-1R, CD30, TAG-72, SPEAP, CD45, L1-CAM, Lewis Y (Le y ) antigens, E-cadherin, V-cadherin and EpCAM.

[0027] Additionally or alternatively, in some embodiments of the methods disclosed herein, the anti-DOTA bispecific antibody, the clearing agent, and / or the radiolabeled DOTA hapten is administered intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intradermally, intraperitoneally, transtracheally, subcutaneously, intracerebroventricularly, orally, or intranasally.

[0028] Additionally or alternatively, in some embodiments of the methods disclosed herein, the cancer is selected from breast cancer, colorectal cancer, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, brain cancer, lung cancer, stomach cancer or gastric cancer, pancreatic cancer, thyroid cancer, kidney cancer or renal cancer, prostate cancer, melanoma, sarcoma, carcinoma, Wilms tumor, endometrial cancer, glioblastoma, squamous cell carcinoma, astrocytoma, salivary gland cancer, vulvar cancer, penile cancer, and head and neck cancer. The brain cancer can be a pituitary adenoma, meningioma, neuroblastoma, or craniopharyngioma.

[0029] Additionally or alternatively, in some embodiments of the methods disclosed herein, the radiolabeled DOTA hapten comprises one or more of the following: Proteus-DOTA, S-2-(R-aminobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (DOTA-Bn), DOTA-Bn-biotin, BAD (((S)-2-(4-(2-bromo)-acetamido)-benzyl)-DOTA), NBD ((S)-2-(4-nitro The radiolabeled DOTA hapten may be labeled with a radionuclide selected from the group consisting of: 213 Bi, 211 At 225 Ac, 152 Dy, 212 Bi, 223 Ra, 219 Rn, 215 Po, 211 Bi, 221 Fr. 217 At 255 Fm, 86 Y. 90 Y. 89 Sr. 165 Dy, 186 Re, 188 Re, 177 Lu, 67 Cu, 111 In, 67 Ga, 51 Cr, 58 Co. 99m Tc, 103m Rh, 195m Pt, 119 Sb, 161 Ho, 189m Os, 192 Ir, 201 Tl, 203 Pb, 68 Ga, 227 Th and 64 Cu.

[0030] Additionally or alternatively, in some embodiments of the methods disclosed herein, the level of radioactivity emitted by the radiolabeled DOTA hapten-anti-DOTA bispecific antibody complex is detected between 4 and 24 hours after administration of the radiolabeled DOTA hapten. The level of radioactivity emitted by the complex can be expressed as a percentage of the injected dose per gram of tissue (% ID / g). The reference value can be calculated by measuring the level of radioactivity present in non-tumor (normal) tissue and calculating the average radioactivity level ± standard deviation present in non-tumor (normal) tissue. In some embodiments, the reference value is a standard uptake value (SUV). See Thie JA, J Nucl Med. 45(9): 1431-4 (2004). The therapeutic effectiveness of this complex can be determined by calculating the area under the curve (AUC) tumor: AUC normal tissue ratio. In some embodiments, the complex has an AUC tumor:AUC normal tissue ratio of about 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 55:1, 60:1, 65:1, 70:1, 75:1, 80:1, 85:1, 90:1, 95:1, or 100:1. Additionally or alternatively, in some embodiments of the methods disclosed herein, the ratio of radioactivity levels between tumor and normal tissue is about 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 55:1, 60:1, 65:1, 70:1, 75:1, 80:1, 85:1, 90:1, 95:1, or 100:1.

[0031] Also disclosed herein are kits containing components suitable for treating cancer in patients. In one aspect, the kit comprises a scavenger of the present invention and instructions for use. The kit of the present invention may also comprise at least one anti-DOTA BsAb and / or a DOTA hapten optionally labeled with one or more radionuclides. Examples of suitable radionuclides include, but are not limited to 213 Bi, 211 At 225 Ac, 152 Dy, 212 Bi, 223 Ra, 219 Rn, 215 Po, 211 Bi, 221 Fr. 217 At 255 Fm, 86 Y.90 Y. 89 Sr. 165 Dy, 186 Re, 188 Re, 177 Lu, 67 Cu, 111 In, 67 Ga, 51 Cr, 58 Co. 99m Tc, 103m Rh, 195m Pt, 119 Sb, 161 Ho, 189m Os, 192 Ir, 201 Tl, 203 Pb, 68 Ga, 227 Th and 64 Cu. Additionally or alternatively, in some embodiments, the at least one anti-DOTA BsAb binds to a tumor antigen target selected from the group consisting of GPA33, HER2 / neu, GD2, MAGE-1, MAGE-3, BAGE, GAGE-1, GAGE-2, MUM-1, CDK4, N-acetylglucosamine transferase, p15, gp75, β-catenin, ErbB2, cancer antigen 125 (CA-125), carcinoembryonic antigen (CEA), RAGE, MART (melanoma antigen), MUC-1, MUC-2, MUC-3, MUC-4, MUC-5ac, MUC-16, MUC-17, tyrosinase, Pmel 17 (gp100), GnT-V intron V sequence (N-acetylglucosamine transferase V intron V sequence), prostate cancer psm, PRAME (melanoma antigen), β-catenin, EBNA (Epstein-Barr virus nuclear antigen) 1-6, p53, lung resistance protein (LRP) Bcl-2, prostate specific antigen (PSA), Ki-67, CEACAM6, colon specific antigen-p (CSAp), HLA-DR, CD40, CD74, CD138, EGFR, EGP-1, EGP-2, VEGF, PlGF, insulin-like growth factor (ILGF), tenascin, platelet-derived growth factor, IL-6, CD20, CD19, PSMA, CD33, CD123, MET, DLL4, Ang-2, HER3, IGF-1R, CD30, TAG-72, SPEAP, CD45, L1-CAM, Lewis Y (Le y ) antigens, E-cadherin, V-cadherin and EpCAM. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 The scavengers disclosed herein have been shown to have an effect on the circulation of 131 Effect of I-BsAb. Normal athymic (tumor-free) mice were injected intravenously at t = 0 131 I-BsAb (19-21 μCi; 250 μg, 1.19 nmol) was then injected intravenously with vehicle (saline) or dendrimer scavenger (25 μg; 2.76 nmol) at t=24 hours. Serial blood sampling was performed at various time points from t=1-28 hours. Data are presented as mean ± standard error of the mean.

[0033] Figure 2 The scavenger CCA-16-DOTA-Y of the present invention is shown. 3+ The achieved tumor pretargeting results.

[0034] Figure 3 CCA-16-DOTA-Y is shown 3+ Dose-dependent effect on tumor uptake. **P<0.01 compared with the 25 μg group.

[0035] Figure 4 Shown is a comparison of tumor pretargeting results achieved with embodiments of clearing agents of the present technology compared to no clearing agent (vehicle) and with a 500 kD dextran-DOTA hapten conjugate clearing agent.

[0036] Figure 5(A) shows the tracer pretargeting in athymic nude mice bearing SW1222 tumors at 24 h post-injection. 225 Ac]Proteus-DOTA (n=3) or [ 111 Comparison of the biodistribution of In]Proteus-DOTA (n=5). After intravenous injection (via the lateral tail vein) of huA33-C825 antibody (0.25 mg, 1.19 nmol), scavenger and radiolabeled DOTA-hapten, animals were euthanized 24 h later for organ collection and radioactivity determination. *P<0.05, ***P<0.001. Data are presented as mean ± SEM.

[0037] FIG. 5(B) shows the absolute tumor uptake of pretargeted radiolabeled DOTA hapten at 24 h post-injection, plotted as a function of the molar amount of tracer administered; n=1-7 for each data point.

[0038] Figure 6(A) shows the results of the injection of 172pmol / 1.67MBq [45μCi] of 111In] Proteus-DOTA (n = 4) or 790 pmol / 7.66 MBq [207 μCi] [ 111 Results of tumor pretargeting achieved in animals treated with In] Proteus-DOTA (n=1).

[0039] Figure 6(B) shows the results of the anti-GPA33-DOTA-PRIT assay. 225 Ac]Proteus-DOTA and [ 111 In] Statistical comparison of Proteus-DOTA pretargeting.

[0040] Figure 7 Shown is the effect of injection of pretargeted [ 111 In] SPECT / CT image approximately 24 h after Proteus-DOTA. SW1222 xenografts can be clearly delineated in the flank. Image-based region of interest analysis of the tumor, kidney (left side), heart, and liver revealed active concentrations (expressed as mean ± 1SD; percent injected dose per gram) of 6.89 ± 4.68, 0.46 ± 0.47, 0.20 ± 0.24, and 0.22 ± 0.27, respectively.

[0041] Figure 8 The structure of Proteus-DOTA is shown (chemical formula: C 50 H 80 LqCy 11 O 19 S 3- ; Exact mass: 1345.48; Molecular weight: 1346.28). The boxed portion of the molecule is the non-radioactive benzyl-DOTA(Lu) hapten, which is expressed as K d =10 pM is recognized by the anti-DOTA-hapten antibody single chain variable fragment C825. The empty three-arm DOTA portion of the molecule can accommodate a variety of radioactive metals related to therapy and / or imaging, including 225 Ac, 68 Ga and 64 Cu.

[0042] Fig. 9 The results show that the huA33-C825 (0.25 mg / mouse) and the dendrimer scavenger CCA-16-DOTA-Y 3+ (25 μg; 2.76 nmol) and 3.7 MBq (20 pmol) 177 Lu-aminobenzyl DOTA ([ 177 After PRIT with Lu]LuDOTA-Bn, the biodistribution assay was used to determine the expression of177 Lu activity. n = 5 animals / group; data presented as %ID / g, mean ± 1 SD.

[0043] Fig.10 showed that after injection of huA33-C825 PRIT+[ 177 Attenuation-corrected saturation of SW1222 tumors and selected normal tissues 1 to 48 hours after the addition of Lu]LuDOTA-Bn (3.7 MBq, 20 pmol) and the dendrimer scavenger CCA-16-DOTA-Y3+ (25 μg; 2.76 nmol). 177 Lu activity biodistribution curve. Data are presented as %ID / g, mean ± 1SD.

[0044] Fig.11 The dendrimer scavenger CCA-16-DOTA-Y was used for pretargeting in nude mice bearing subcutaneous GPA33(+)SW1222 tumors. 3+ of[ 177 The therapeutic index was defined as the estimated tumor / normal tissue absorbed dose ratio. DETAILED DESCRIPTION

[0045] It should be appreciated that certain aspects, modes, embodiments, variations and features of the inventive methods are described below in varying levels of detail in order to provide a substantial understanding of the inventive technology.

[0046] In practicing the methods of the present invention, many conventional techniques in molecular biology, protein biochemistry, cell biology, microbiology, and recombinant DNA are used. See, for example, Sambrook and Russell, eds. (2001) Molecular Cloning: A Laboratory Manual, 3rd edition; Ausubel et al., eds. (2007) Current Protocols in Molecular Biology; Methods in Enzymology (Academic Press, Inc., New York) series; MacPherson et al. (1991) PCR 1: A Practical Approach (IRL Press at Oxford University Press); MacPherson et al. (1995) PCR 2: A Practical Approach; Harlow and Lane, eds. (1999) Antibodies, A Laboratory Manual; Freshney (2005) Culture of Animal Cells: A Manual of Basic Technique, 5th edition; Gait, ed. (1984) Oligonucleotide Synthesis; U.S. Pat. No. 4,683,195; Hames and Higgins, eds. (1984) Nucleic Acid Hybridization; Anderson (1999) Nucleic Acid Hybridization; Hames and Higgins, eds. (1984) Transcription and Translation; Immobilized Cells and Enzymes (IRL Press (1986)); Perbal (1984) A Practical Guide to Molecular Cloning; Miller and Calos, eds. (1987) Gene Transfer Vectors for Mammalian Cells (Cold Spring Harbor Laboratory); Makrides, ed. (2003) Gene Transfer and Expression in Mammalian Cells; Mayer and Walker, eds. (1987) Immunochemical Methods in Cell and Molecular Biology (Academic Press, London); and Herzenberg et al., eds. (1996) Weir's Handbook of Experimental Immunology.

[0047] Clearing agents (CAs) are a class of compounds designed to rapidly remove targeted biomolecules from the circulation during pretargeting of radioisotopes to tumor antigens. Dextran-based clearing agents are commonly used in high therapeutic index DOTA-based pretargeted radioimmunotherapy (DOTA PRIT). Although highly effective as clearing agent scaffolds, dextran poses dosing challenges due to its inherent polydispersity and kinetics of enzymatic degradation in vivo.

[0048] The DOTA-PRIT platform disclosed herein requires a three-step pretargeting strategy, including administration of (1) an IgG-single chain variable fragment (scFv) bispecific antibody construct (IgG-scFv) comprising an antibody sequence with high affinity for an anti-tumor antigen antibody (IgG portion) and an anti-DOTA-hapten single chain variable fragment scFv (e.g., "C825"); (2) a clearing agent of the present technology, which is used to rapidly reduce circulating BsAb after sufficient time has been given for the BsAb to accumulate in antigen-positive tumors; and (3) a radiolabeled DOTA hapten composition (e.g., 177 Lu-DOTA-Bn).

[0049] The compositions of the present technology include novel N-acetylgalactosamine dendrimer scavengers that can be used in PRIT (e.g., alpha particle radioimmunotherapy). The scavenger compositions disclosed herein exhibit enhanced blood clearance of DOTA-PRIT radiolabeled bispecific antibodies (BsAbs) and improved therapeutic index (TI) during DOTA-PRIT. For example, upon administration of a single dose of an excess of the dendrimer scavenger of the present technology (injected 131 Within 5 minutes, the molar ratio of I-BsAb to CA was 1:2.3. 131 I activity decreased by 64%, from a baseline of 6.7% ID / g to 2.4% ID / g. DOTA-PRIT studies in mouse xenograft models of human colorectal cancer showed that injection of 177 Lu activity after 24 hours 177 Lu-DOTA-Bn CA dose-dependent tumor-to-blood uptake ratios (e.g., mean tumor-to-blood ratios were 2.9, 26, and 59 for 0 μg (vehicle), 15 μg, or 20 μg of dendrimer CA, respectively). A 25 μg dose of dendrimer CA resulted in a mean tumor-to-blood ratio of 76, which is almost identical to the previously optimized dextran CA dosing (mean tumor-to-blood ratio of 77). Overall, these results demonstrate that the dendrimer scavengers of the present technology are suitable for high TIDOTA-PRIT.

[0050] definition

[0051] Unless otherwise defined, all technical and scientific terms used herein generally have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention belongs. As used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural indicators unless the context clearly dictates otherwise. For example, reference to "a cell" includes combinations of two or more cells, and so on. Generally, the nomenclature used herein and the laboratory procedures in cell culture, molecular genetics, organic chemistry, analytical chemistry, and nucleic acid chemistry and hybridization described below are those well known and commonly used in the art.

[0052] As used herein, the term "about" with respect to a number is generally deemed to include numbers that fall within 1%, 5%, or 10% in either direction (greater than or less than) of the stated number, unless the context indicates otherwise or it is otherwise obvious (except where such a number is below 0% or above 100% of the possible value).

[0053] Pharmaceutically acceptable salts of the compounds described herein are within the scope of the present invention and include acid or base addition salts that retain the desired pharmacological activity and are not biologically undesirable (e.g., the salts are not overly toxic, sensitizing, or irritating and are bioavailable). When the compound of the present invention has a basic group (e.g., such as an amino group), a pharmaceutically acceptable salt may be formed with an inorganic acid (e.g., hydrochloric acid, hydroboric acid, nitric acid, sulfuric acid, and phosphoric acid), an organic acid (e.g., alginic acid, formic acid, acetic acid, benzoic acid, gluconic acid, fumaric acid, oxalic acid, tartaric acid, lactic acid, maleic acid, citric acid, succinic acid, malic acid, methanesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, and p-toluenesulfonic acid) or an acidic amino acid (e.g., aspartic acid and glutamic acid). When the compound of the present invention has an acidic group (e.g., such as a carboxylic acid group), it may form a salt with a metal (e.g., an alkali metal and an alkaline earth metal (e.g., Na + , Li + , K + , Ca 2+ Mg 2+ 、Zn 2+ )), ammonia or organic amines (e.g., dicyclohexylamine, trimethylamine, triethylamine, pyridine, picoline, ethanolamine, diethanolamine, triethanolamine) or basic amino acids (e.g., arginine, lysine and ornithine). Such salts can be prepared in situ during the isolation and purification of the compound, or by separately reacting the purified compound (in its free base or free acid form) with a suitable acid or base, respectively, and isolating the salt thus formed.

[0054] As used herein, "administering" an agent or drug to a subject includes any route of introducing or delivering a compound to a subject to perform its intended function. Administration can be by any suitable route, including oral, intranasal, parenteral (intravenous, intramuscular, intraperitoneal or subcutaneous), rectal or topical. Administration includes self-administration and administration by another person.

[0055] As used herein, the term "antibody" collectively refers to immunoglobulins or immunoglobulin-like molecules, including, for example, but not limited to, IgA, IgD, IgE, IgG, and IgM, combinations thereof, and similar molecules produced during an immune response in any vertebrate, such as mammals (such as humans, goats, rabbits, and mice), as well as non-mammalian species (such as shark immunoglobulins). As used herein, "antibodies" (including "whole immunoglobulins") and "antigen-binding fragments" specifically bind to a target molecule (or a group of highly similar target molecules) while substantially excluding binding to other molecules (e.g., the binding constant for the target molecule is about 10 times the binding constant for other molecules in a biological sample). 3 M -1 times, about 10 4 M -1 times or about 10 5 M -1 The term "antibody" also includes genetically engineered forms such as chimeric antibodies (e.g., humanized murine antibodies), heteroconjugate antibodies (e.g., bispecific antibodies). See also Pierce Catalog and Handbook, 1994-1995 (Pierce Chemical Co., Rockford, IL); Kuby, J., Immunology, 3rd ed., WH Freeman & Co., New York, 1997.

[0056] More specifically, an antibody refers to a polypeptide ligand that specifically recognizes and binds to an antigen epitope and contains at least a light chain immunoglobulin variable region or a heavy chain immunoglobulin variable region. Antibodies are composed of heavy chains and light chains, each of which has a variable region, called a heavy chain variable region (V H ) region and light chain variable (V L ) area. H Area and V LThe regions are jointly responsible for binding to the antigen recognized by the antibody. Typically, immunoglobulins have heavy (H) chains and light (L) chains interconnected by disulfide bonds. There are two types of light chains, namely λ (lambda) and κ (kappa). There are five main heavy chain categories (or isotypes): IgM, IgD, IgG, IgA and IgE, which determine the functional activity of the antibody molecule. Each heavy chain and light chain contains a constant region and a variable region (the region is also referred to as a "domain"). Combined, the heavy chain variable region and the light chain variable region specifically bind to the antigen. The light chain variable region and the heavy chain variable region contain a "framework" region interrupted by three hypervariable regions (also referred to as "complementarity determining regions" or "CDRs"). The scope of the framework region and CDR has been defined (see Kabat et al., Sequences of Proteins of Immunological Interest, US Department of Health and Human Services, 1991, which is hereby incorporated by reference). The Kabat database is currently maintained online. The sequences of the framework regions of different light or heavy chains are relatively conserved within species. The framework region of an antibody (i.e., the combined framework region of the constituent light and heavy chains) predominantly adopts a β-sheet conformation, and the CDR forms a loop connecting the β-sheet structure, and in some cases the loop forms a part of the β-sheet structure. Therefore, the framework region plays a role in forming a scaffold that positions the CDR in the correct orientation through interchain non-covalent interactions.

[0057] The CDRs are primarily responsible for binding to the antigenic epitope. The CDRs of each chain are usually referred to as CDR1, CDR2, and CDR3, are numbered sequentially starting from the N-terminus, and are usually also identified according to the chain in which a particular CDR is located. H CDR3 is located in the variable domain of the heavy chain of the antibody in which it is found, while V L CDR1 is the CDR1 from the variable domain of the light chain of the antibody in which it is found. An antibody that binds to a target protein (e.g., GPA33) or molecule (e.g., DOTA or a DOTA hapten) will have a specific V H Area and V L Region sequence, therefore has specific CDR sequence. Antibodies with different specificities (i.e., different binding sites for different antigens) have different CDRs. Although CDRs vary from antibody to antibody, only a limited number of amino acid positions in CDRs are directly involved in antigen binding. These positions in CDRs are called specificity determining residues (SDRs). Examples of antibodies include monoclonal antibodies, polyclonal antibodies, humanized antibodies, chimeric antibodies, recombinant antibodies, multispecific antibodies, bispecific antibodies, and antibody fragments. Antibodies specifically bind to antigens.

[0058] "Bispecific antibodies" are antibodies that can bind to two different antigens simultaneously. Bispecific antibodies (BsAb) and bispecific antibody fragments (BsFab) can have at least one arm that specifically binds to, for example, a tumor-associated antigen (e.g., GPA33) and at least another arm that specifically binds to a targetable conjugate carrying a therapeutic or diagnostic agent (e.g., a DOTA hapten associated with a radionuclide). A variety of different bispecific antibody structures are known in the art. In some embodiments, each binding moiety in a bispecific antibody comprises V from a different monoclonal antibody. H and / or V L In some embodiments, the bispecific antibody comprises an immunoglobulin molecule having a V region containing the CDRs from a first monoclonal antibody. H and / or V L and antibody fragments (e.g., Fab, F(ab'), F(ab')2, Fd, Fv, dAB, scFv, etc.) having a V region containing the CDRs from a second monoclonal antibody H and / or V L district.

[0059] As used herein, the term "diabodies" refers to small antibody fragments with two antigen-binding sites contained in the same polypeptide chain in conjunction with a light chain variable domain (V L ) connected to the heavy chain variable domain (V H )(V H V L ). By using a linker that is too short to allow pairing between the two domains on the same chain, the domains are forced to pair with the complementary domains of another chain and create two antigen-binding sites. Diabodies are more fully described in, for example, EP 404,097; WO 93 / 11161; and 30 Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993).

[0060] As used herein, the term "single-chain antibody" or "single-chain Fv (scFv)" refers to the two domains V and Fv fragments. L and V H The single-chain antibody molecule may contain a polymer having multiple individual molecules, such as a dimer, trimer or other polymer. v The two domains of the fragment V L and V H They are encoded by separate genes, but they can be joined by synthetic linkers using recombinant methods so that they can become a single protein chain, in which V L and V HThe domains pair to form a monovalent molecule (called a single-chain F v (scF v )). Bird et al. (1988) Science 242:423-426 and Huston et al. (1988) Proc. Natl. Acad Sci. USA 85:5879-5883. Such single-chain antibodies can be prepared by recombinant techniques or by enzymatic or chemical cleavage of intact antibodies.

[0061] As used herein, the term "intact antibody" or "intact immunoglobulin" means an antibody or immunoglobulin having at least two heavy (H) chain polypeptides and two light (L) chain polypeptides interconnected by disulfide bonds. Each heavy chain consists of a heavy chain variable region (abbreviated herein as HCVR or V H ) and a heavy chain constant region. The heavy chain constant region is composed of three domains: CH1, CH2, and CH3. Each light chain consists of a light chain variable region (abbreviated herein as LCVR or V L ) and the light chain constant region. The light chain constant region consists of a domain C L Composition. H and V L The V region can be further subdivided into regions of high variability, termed complementarity determining regions (CDRs), interspersed with more conserved regions, termed framework regions (FRs). H and V L It is composed of three CDRs and four FRs, arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens. The constant region of an antibody can mediate the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system.

[0062] As used herein, "antigen" refers to a molecule to which an antibody can selectively bind. The target antigen can be a protein (e.g., an antigenic peptide), a carbohydrate, a nucleic acid, a lipid, a hapten, or other naturally occurring or synthetic compound. Antigens can also be administered to an animal subject to generate an immune response in the subject.

[0063] As used herein, the term "antigen-binding fragment" refers to a fragment of the entire immunoglobulin structure that has a portion of the polypeptide responsible for binding to an antigen. Examples of antigen-binding fragments that can be used in the technology of the present invention include scFv, (scFv)2, scFvFc, Fab, Fab' and F(ab')2; diabodies; linear antibodies; single-chain antibody molecules; and multispecific antibodies formed from antibody fragments.

[0064] "Binding affinity" refers to the strength of the total non-covalent interaction between a single binding site of a molecule (e.g., an antibody) and the binding partner of that molecule (e.g., an antigen). The affinity of a molecule X for its partner Y can generally be expressed by the dissociation constant (K d ) is represented by. Affinity can be measured by standard methods known in the art (including those described herein). Low affinity complexes contain antibodies that generally tend to dissociate easily from the antigen, while high affinity complexes contain antibodies that generally tend to remain bound to the antigen for a longer period of time.

[0065] As used herein, a "scavenger" is an agent that binds to excess bifunctional antibodies present in the blood compartment of a subject to promote rapid clearance via the kidneys. The use of the scavenger prior to administration of a DOTA-based radiotherapeutic promotes a better tumor-to-background ratio in the PRIT system.

[0066] As used herein, a "control" is an alternative sample used in an experiment for comparison purposes. A control can be "positive" or "negative." For example, where the purpose of an experiment is to determine the efficacy relevance of a therapeutic agent for treating a particular type of disease or condition, a positive control (a compound or composition known to exhibit the desired therapeutic effect) and a negative control (a subject or sample that does not receive therapy or receives a placebo) are typically employed.

[0067] As used herein, the term "effective amount" of a composition is an amount sufficient to achieve the desired preventive or therapeutic effect, such as an amount that results in a reduction in symptoms associated with the disease being treated (e.g., a disease associated with the target polypeptide or a medical condition (e.g., breast cancer, colorectal cancer, brain cancer, etc.)). The amount of the composition of the present technology administered to the subject will depend on the extent, type, and severity of the disease and the characteristics of the individual, such as overall health, age, sex, weight, and tolerance to drugs. A skilled person will be able to determine an appropriate dosage based on these and other factors. The composition of the present technology may also be administered in combination with one or more additional therapeutic compounds.

[0068] As used herein, the term "epitope" means an antigenic determinant capable of specific binding to an antibody. An epitope usually consists of chemically active surface groups of a molecule and usually has specific three-dimensional structural characteristics as well as specific charge characteristics.

[0069] As used herein, the term "sample" refers to a clinical sample or isolated microorganism obtained from a subject. In certain embodiments, a sample is obtained from a biological source (i.e., a "biological sample"), such as a tissue, body fluid, or microorganism collected from a subject. Sample sources include, but are not limited to, mucus, sputum, bronchoalveolar lavage fluid (BAL), bronchial washings (BW), whole blood, body fluids, cerebrospinal fluid (CSF), urine, plasma, serum, or tissue.

[0070] As used herein, the term "separate" therapeutic use refers to the simultaneous or substantially simultaneous administration of at least two active ingredients by different routes.

[0071] As used herein, the term "sequential" therapeutic use refers to the administration of at least two active ingredients at different times, with the same or different routes of administration. More specifically, sequential use refers to the entire administration of one active ingredient before the administration of the other one or more active ingredients begins. Thus, one active ingredient can be administered a few minutes, hours, or days before the administration of the other one or more active ingredients. In this case, there is no simultaneous treatment.

[0072] As used herein, the term "simultaneous" therapeutic use refers to the administration of at least two active ingredients by the same route and at the same time or substantially at the same time.

[0073] As used herein, "specifically binds" refers to a molecule (e.g., an antibody) recognizing and binding to another molecule (e.g., an antigen), but not substantially recognizing and binding to other molecules. As used herein, the term "specifically binds to," "specifically binds to," or "specifically binds to" a particular molecule (e.g., an antigen or an epitope on an antigen) can be, for example, determined by the K of the molecule to which it binds. d About 10 -4 M, 10 -5 M, 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 M, 10 -10 M, 10 -11 M or 10 -12 M to show.

[0074] As used herein, the terms "subject," "individual," or "patient" are used interchangeably and refer to a single organism, vertebrate, mammal, or human. In certain embodiments, the individual, patient, or subject is a human.

[0075] As used herein, the term "therapeutic agent" is intended to mean a compound that, when present in an effective amount, produces a desired therapeutic effect on a subject in need thereof.

[0076] As used herein, "treating" or "treatment" encompasses treating a disease or disorder described herein in a subject, such as a human, and includes: (i) inhibiting the disease or disorder, i.e., preventing its development; (ii) relieving the disease or disorder, i.e., causing regression of the disorder; (iii) slowing the progression of the disorder; and / or (iv) inhibiting, relieving, or slowing the progression of one or more symptoms of the disease or disorder. "Treating cancer" means that symptoms associated with the cancer are, for example, alleviated, relieved, cured, or in a state of remission.

[0077] It is also understood that the various treatment modalities for diseases as described herein are intended to mean "substantially," which includes complete treatment but less than complete treatment, and in which some biologically or medically relevant result is achieved. The treatment can be continuous, extended treatment for chronic diseases, or a single or several administrations for acute condition treatment.

[0078] Use of scavengers in pretargeted radioimmunotherapy (PRIT)

[0079] The therapeutic index (TI) of radioimmunotherapy (RIT) should be maximized to effectively treat solid tumors (Larson SM et al., Nature Reviews Cancer 15:347-60 (2015)). RIT with radiolabeled IgG antibodies has a low TI due to the slow pharmacokinetics of the IgG carrier and is therefore generally ineffective at tolerable doses.

[0080] An alternative is to use a pre-targeting method (PRIT) for RIT. During PRIT, the slow antibody-mediated tumor targeting step is separated from the administration of radioactivity. Pre-targeting is a multi-step process to solve the slow blood clearance of antibodies targeting tumors, which contributes to the undesirable toxicity to normal tissues (such as bone marrow). During pre-targeting, radionuclides or other diagnostic agents or therapeutic agents are attached to carriers with more favorable pharmacokinetics (e.g., low molecular weight compounds with rapid renal clearance and low normal tissue uptake and systemic clearance (Orcutt KD et al., Molecular imaging and biology 13: 215-21 (2011)), such as DOTA-haptens), and the dose of normal tissues is minimized while targeting tumors. In order to target radioactivity to tumors, circulating therapeutic agents (e.g., DOTA-haptens) are captured by BsAbs located in tumors or otherwise effectively cleared via renal pathways.

[0081] Scavengers can rapidly reduce the concentration of circulating biomolecules by forming large complexes in the circulation that are recognized by the reticuloendothelial system (RES) or targeted to the hepatic asialoglycoprotein receptor (ASPGR) using appropriate sugar haptens (Rossin R et al., Journal Nuclear Medicine 54: 1989-95 (2013)). The TI of DOTA-PRIT is highly variable depending on scavenger administration because circulating BsAb retains the ability to bind to the radiolabeled DOTA hapten. In order to maximize tumor targeting of the radiolabeled DOTA hapten, a saturating dose of BsAb is typically used to ensure that the maximum concentration of the anti-DOTA antibody domain is used to bind to the subsequently administered radiolabeled DOTA hapten. This step poses the challenge of removing excess circulating unbound BsAb to maximize TI prior to administration of the cytotoxic radiolabeled DOTA hapten. Although long time intervals can be used to allow for endogenous clearance of the BsAb, this may affect the ability of the BsAb to bind the radioactive hapten at the tumor because the BsAb may be degraded and / or internalized depending on the molecular pharmacology of the BsAb-antigen complex.

[0082] 500kD dextran-DOTA hapten conjugate (Orcutt KD et al., Molecular Cancer Therapeutics 11: 1365-72 (2012)) has been previously used for DOTA-based pre-targeting of carcinoembryonic antigen. Dextran CA is designed to bind to the anti-DOTA (M)-scFv domain of circulating BsAb via the DOTA (Y) portion displayed on the dextran scaffold, and remove unbound BsAb from the blood via recognition and catabolism by the reticuloendothelial system (RES). Due to its large size, the binding of dextran CA to tumor-associated BsAb is limited by poor intratumoral extravasation.

[0083] Although highly effective, the use of glucan CA has disadvantages. As a naturally occurring glucose polymer, the glucan scaffold is inherently polydisperse, thus bringing challenges related to reproducible batch production and in vivo use. In addition, the enzymatic degradation of RES glucan-1,6-glucosidase may cause its hapten fragment to be introduced into the circulation, thereby possibly competing with the radioactive hapten to be bound by tumor-BsAb. During clinical streptavidin-biotin PRIT, similar problems were also found in the case of albumin-based CA (Knox SJ et al., Clinical Cancer Research 6: 406-14 (2000); Breitz HB et al., Journal Nuclear Medicine 41: 131-40 (2000)).

[0084] Composition of the present invention

[0085] In one aspect, the present technology provides a compound, which is

[0086]

[0087]

[0088]

[0089]

[0090] or a pharmaceutically acceptable salt and / or solvate thereof, wherein X 1 , X 2 , X 3 , X 4 , X 5 , X 6 , X 7 , X 8 , X 9 , X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 , X 17 , X 18 , X 19 and X 20 Each independently is H or a lone pair of electrons (i.e., providing an oxygen anion); Y 1 , Y 2 , Y 3 , Y 4 and Y 5 are each independently O or S; x is 1, 2 or 3; and y is 1, 2, 3 or 4.

[0091] In any of the embodiments herein, the compound may be a scavenger (also referred to herein as a "dendrimer scavenger" of the present technology, a "dendrimer CA" of the present technology, etc.), wherein the scavenger is

[0092]

[0093]

[0094]

[0095]

[0096] or a pharmaceutically acceptable salt and / or solvate thereof, wherein M1 、M 2 、M 3 、M 4 and M 5 Each independently is Lu 3+ Sc 3+ , Ga 3+ , Y 3+ 、In 3+ ,La 3+ 、Ce 3+ 、Eu 3+ , Tb 3+ or Gd 3+ ;X 1 , X 2 , X 3 , X 4 , X 5 , X 6 , X 7 , X 8 , X 9 , X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 , X 17 , X 18 , X 19 and X 20 Each independently is H or a lone pair of electrons (i.e., providing an oxygen anion); Y 1 , Y 2 , Y 3 , Y 4 and Y 5 are each independently O or S; x is 1, 2 or 3; and y is 1, 2, 3 or 4.

[0097] In any of the embodiments herein, M 1 、M 2 、M 3 、M 4 and M 5 In any embodiment herein, Y 1 , Y 2 , Y 3 , Y 4 and Y 5 and x may each independently be S. In any embodiment herein, x may be 1 or 2. In any embodiment herein, y may be 2 or 3.

[0098] In a related aspect, a composition is provided, the composition comprising one or more of any embodiments of the compound as described above and a pharmaceutically acceptable carrier (unless more specific terms are used, such carriers, excipients, fillers, etc. will be collectively referred to as "pharmaceutically acceptable carriers"). The composition can be used in the methods and imaging described herein. The technology of the present invention also provides pharmaceutical compositions and drugs, which contain one or more of any embodiments of the compound as described above and a pharmaceutically acceptable carrier. Such pharmaceutical compositions can be packaged in unit dosage forms. The pharmaceutical compositions and drugs can be prepared by the following method: mixing one or more compounds of the technology of the present invention, their pharmaceutically acceptable salts and / or solvates with pharmaceutically acceptable carriers, excipients, binders, diluents, etc. Such compositions can be, for example, in the form of granules, powders, tablets, capsules, syrups, suppositories, injections, emulsions, elixirs, suspensions or solutions. The compositions of the present invention can be formulated for a variety of administration routes, such as by oral, parenteral or rectal administration. Parenteral or systemic administration includes, but is not limited to, subcutaneous, intravenous, intraperitoneal and intramuscular injections. The following dosage forms are given as examples and should not be construed as limiting the present technology.

[0099] For oral, buccal and sublingual administration, powder, suspension, granule, tablet, pill, capsule, soft capsule and caplet are acceptable as solid dosage form. These can be prepared, for example, by the following method: one or more compounds of the present invention technology or its pharmaceutically acceptable salt or tautomer are mixed with at least one additive (such as starch or other additives). Suitable additives are sucrose, lactose, cellulose sugar, mannitol, maltitol, dextran, starch, agar, alginate, chitin, chitosan, pectin, tragacanth, gum arabic, gelatin, collagen, casein, albumin, synthetic or semi-synthetic polymer or glyceride. Optionally, oral dosage form can contain other ingredients that contribute to administration, such as inert diluent or lubricant (such as magnesium stearate) or preservative (such as paraben or sorbic acid) or antioxidant (such as ascorbic acid, tocopherol or cysteine), disintegrant, binder, thickener, buffer, sweetener, flavoring agent or fragrance. Tablets and pills can be further processed with suitable coating materials known in the art.

[0100] Liquid dosage forms for oral administration may be in the form of pharmaceutically acceptable emulsions, syrups, elixirs, suspensions and solutions, which may contain inert diluents (such as water). Pharmaceutical formulations and drugs may be prepared as liquid suspensions or solutions using sterile liquids (such as, but not limited to, oils, water, alcohols and combinations of these). Pharmaceutically suitable surfactants, suspending agents, emulsifiers may be added for oral or parenteral administration.

[0101] As mentioned above, suspensoids can include oils. Such oils include, but are not limited to, peanut oil, sesame oil, cottonseed oil, corn oil and olive oil. Suspension preparations can also contain fatty acid esters, such as ethyl oleate, isopropyl myristate, fatty acid glycerides and acetylated fatty acid glycerides. Suspension formulations can include alcohols, such as, but not limited to, ethanol, isopropyl alcohol, hexadecanol, glycerol and propylene glycol. Ethers (such as, but not limited to, poly (ethylene glycol), petroleum hydrocarbons (such as mineral oil and petrolatum)) and water can also be used in suspension formulations.

[0102] Injectable dosage forms generally include aqueous suspensions or oil suspensions, which can be prepared using suitable dispersants or wetting agents and suspending agents. Injectable forms can be in the solution phase or in the form of a suspension prepared with a solvent or diluent. Acceptable solvents or vehicles include sterile water, Ringer's solution or isotonic aqueous saline solutions. Isotonic solutions will be understood to be isotonic with the subject. Alternatively, sterile oils can be used as solvents or suspending agents. Typically, the oil or fatty acid is non-volatile, including natural or synthetic oils, fatty acids, monoglycerides, diglycerides or triglycerides.

[0103] For injection, the pharmaceutical formulation and / or drug may be a powder suitable for reconstitution with an appropriate solution as described above. Examples of these include, but are not limited to, freeze-dried, rotary-dried or spray-dried powders, amorphous powders, granules, precipitates or microparticles. For injection, the formulation may optionally contain stabilizers, pH regulators, surfactants, bioavailability regulators and combinations of these.

[0104] In addition to those representative dosage forms described above, pharmaceutically acceptable excipients and carriers are generally known to those skilled in the art and are therefore included in the technology of the present invention. Such excipients and carriers are described in, for example, "Remingtons Pharmaceutical Sciences" Mack Pub. Co., New Jersey (1991), which is incorporated herein by reference.

[0105] As described below, the formulations of the present technology can be designed to be short-acting, fast-releasing, long-acting and sustained-releasing. Therefore, the pharmaceutical formulations can also be formulated for controlled release or slow release.

[0106] The composition of the present invention may also include, for example, micelles or liposomes, or some other encapsulated forms, or may be administered in a prolonged release form to provide extended storage and / or delivery. Therefore, the pharmaceutical formulation and the drug may be compressed into pellets or cylinders and implanted intramuscularly or subcutaneously as a reservoir injection or as an implant (such as a stent). Such implants may be made of known inert materials such as silicone and biodegradable polymers.

[0107] The specific dosage can be adjusted according to the subject's disease condition, age, weight, overall health, sex and diet, the dosage interval, administration route, excretion rate and combination of the drug. Any of the above dosage forms containing an effective amount are all within the scope of routine experiments, and therefore are also within the scope of the present invention.

[0108] Therapeutic methods of the present invention

[0109] In one aspect, the present disclosure provides a method for increasing the sensitivity of a tumor to radiation therapy in a subject diagnosed with cancer, the method comprising (a) administering to the subject an effective amount of an anti-DOTA bispecific antibody, wherein the anti-DOTA bispecific antibody is configured to localize to a tumor expressing a tumor antigen target; (b) administering to the subject an effective amount of a clearing agent of the present technology; and (c) administering to the subject an effective amount of a radiolabeled DOTA hapten, wherein the DOTA hapten is configured to form a complex with the anti-DOTA bispecific antibody.

[0110] In another aspect, the present disclosure provides a method for treating cancer in a subject in need thereof, the method comprising (a) administering to the subject an effective amount of an anti-DOTA bispecific antibody, wherein the anti-DOTA bispecific antibody is configured to localize to a tumor expressing a tumor antigen target; (b) administering to the subject an effective amount of a clearing agent of the present technology; and (c) administering to the subject an effective amount of a radiolabeled DOTA hapten, wherein the DOTA hapten is configured to form a complex with the anti-DOTA bispecific antibody. The method for treating cancer may also include sequentially, separately or simultaneously administering to the subject at least one chemotherapeutic agent selected from the group consisting of nitrogen mustards, ethyleneimine derivatives, alkyl sulfonates, nitrosoureas, gemcitabine, triazenes, folic acid analogs, anthracyclines, taxanes, COX-2 inhibitors, pyrimidine analogs, purine analogs, antibiotics, enzyme inhibitors, epipodophyllotoxins, platinum coordination complexes, vinca alkaloids, substituted ureas, methylhydrazine derivatives, adrenocortical inhibitors, hormone antagonists, endostatin, paclitaxel, camptothecin, SN-38, doxorubicin, doxorubicin analogs, antimetabolites, alkylating agents, antimitotic agents, antiangiogenic agents, tyrosine kinase inhibitors, mTOR inhibitors, heat shock protein (HSP90) inhibitors, proteasome inhibitors, HDAC inhibitors, pro-apoptotic agents, methotrexate and CPT-11.

[0111] Additionally or alternatively, in some embodiments of the methods disclosed herein, the level of radioactivity emitted by the radiolabeled DOTA hapten-anti-DOTA bispecific antibody complex is detected between 4 and 24 hours after administration of the radiolabeled DOTA hapten. The level of radioactivity emitted by the complex can be expressed as a percentage of the injected dose per gram of tissue (% ID / g). The reference value can be calculated by measuring the level of radioactivity present in non-tumor (normal) tissue and calculating the average radioactivity level ± standard deviation present in non-tumor (normal) tissue. In some embodiments, the reference value is a standard uptake value (SUV). See Thie JA, J Nucl Med. 45(9): 1431-4 (2004). The therapeutic effectiveness of this complex can be determined by calculating the area under the curve (AUC) tumor: AUC normal tissue ratio. In some embodiments, the complex has an AUC tumor:AUC normal tissue ratio of about 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 55:1, 60:1, 65:1, 70:1, 75:1, 80:1, 85:1, 90:1, 95:1, or 100:1. Additionally or alternatively, in some embodiments of the methods disclosed herein, the ratio of radioactivity levels between tumor and normal tissue is about 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 55:1, 60:1, 65:1, 70:1, 75:1, 80:1, 85:1, 90:1, 95:1, or 100:1.

[0112] Additionally or alternatively, in some embodiments of the methods disclosed herein, the tumor antigen target is selected from GPA33, HER2 / neu, GD2, MAGE-1, MAGE-3, BAGE, GAGE-1, GAGE-2, MUM-1, CDK4, N-acetylglucosamine transferase, p15, gp75, β-catenin, ErbB2, cancer antigen 125 (CA-125), carcinoembryonic antigen (CEA), RAGE, MART (melanoma antigen), MUC-1, MUC-2, MUC-3, MUC-4, MUC-5ac, MUC-16, MUC-17, tyrosinase, Pmel 17 (gp100), GnT-V intron V sequence (N-acetylglucosamine transferase V intron V sequence), prostate cancer psm, PRAME (melanoma antigen), β-catenin, EBNA (Epstein-Barr virus nuclear antigen) 1-6, p53, lung resistance protein (LRP) Bcl-2, prostate specific antigen (PSA), Ki-67, CEACAM6, colon specific antigen-p (CSAp), HLA-DR, CD40, CD74, CD138, EGFR, EGP-1, EGP-2, VEGF, PlGF, insulin-like growth factor (ILGF), tenascin, platelet-derived growth factor, IL-6, CD20, CD19, PSMA, CD33, CD123, MET, DLL4, Ang-2, HER3, IGF-1R, CD30, TAG-72, SPEAP, CD45, L1-CAM, Lewis Y (Le y ) antigens, E-cadherin, V-cadherin and EpCAM.

[0113] Additionally or alternatively, in some embodiments of the methods disclosed herein, the anti-DOTA bispecific antibody, the clearing agent, and / or the radiolabeled DOTA hapten is administered intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intradermally, intraperitoneally, transtracheally, subcutaneously, intracerebroventricularly, orally, or intranasally.

[0114] Additionally or alternatively, in some embodiments of the methods disclosed herein, the cancer is selected from breast cancer, colorectal cancer, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, brain cancer, lung cancer, stomach cancer or gastric cancer, pancreatic cancer, thyroid cancer, kidney cancer or renal cancer, prostate cancer, melanoma, sarcoma, carcinoma, Wilms tumor, endometrial cancer, glioblastoma, squamous cell carcinoma, astrocytoma, salivary gland cancer, vulvar cancer, penile cancer, and head and neck cancer. The brain cancer can be a pituitary adenoma, meningioma, neuroblastoma, or craniopharyngioma.

[0115] Additionally or alternatively, in some embodiments of the methods disclosed herein, the radiolabeled DOTA hapten comprises one or more of the following: Proteus-DOTA, S-2-(R-aminobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (DOTA-Bn), DOTA-Bn-biotin, BAD (((S)-2-(4-(2-bromo)-acetamido)-benzyl)-DOTA), NBD ((S)-2-(4-nitro

[00136] Alternatively, any radiolabeled DOTA hapten known in the art may be employed in the methods disclosed herein. The radiolabeled DOTA hapten may be labeled with a radionuclide selected from the group consisting of: 213 Bi, 211 At 225 Ac, 152 Dy, 212 Bi, 223 Ra, 219 Rn, 215 Po, 211 Bi, 221 Fr. 217 At 255 Fm, 86 Y. 90 Y. 89 Sr. 165 Dy, 186 Re, 188 Re, 177 Lu, 67 Cu, 111 In, 67 Ga, 51 Cr, 58 Co. 99m Tc, 103m Rh, 195m Pt, 119 Sb, 161 Ho, 189m Os, 192 Ir, 201 Tl, 203 Pb, 68 Ga, 227 Th and 64 Cu.

[0116] In any of the above embodiments of the methods disclosed herein, the subject is a human. The anti-DOTA bispecific antibody is administered under conditions and for a period of time (e.g., according to a dosing regimen) sufficient for the anti-DOTA bispecific antibody to saturate tumor cells and, after administration of the anti-DOTA bispecific antibody, any unbound anti-DOTA bispecific antibody is removed from the bloodstream. In some embodiments, the radiolabeled DOTA hapten is administered after a period of time that may be sufficient to allow clearance of unbound anti-DOTA bispecific antibody.

[0117] Additionally or alternatively, in some embodiments of the methods disclosed herein, the scavenger and the radiolabeled DOTA hapten are administered without further administration of the anti-DOTA bispecific antibody. For example, in some embodiments, the anti-DOTA bispecific antibody is administered according to a regimen comprising at least one cycle of: (i) administration of an anti-DOTA bispecific antibody (optionally saturating the relevant tumor cells); (ii) administration of a scavenger and a radiolabeled DOTA hapten of the present technology; and (iii) optionally additionally administering the radiolabeled DOTA hapten and / or the scavenger without further administration of the anti-DOTA bispecific antibody. In some embodiments, the method may include multiple such cycles (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more cycles).

[0118] The radiolabeled DOTA hapten can be administered any time between 1 minute to 4 or more days after administration of the anti-DOTA bispecific antibody. For example, in some embodiments, the radiolabeled DOTA hapten is administered 1 minute, 2 minutes, 3 minutes, 4 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes, 35 minutes, 40 minutes, 45 minutes, 50 minutes, 55 minutes, 1 hour, 1.25 hours, 1.5 hours, 1.75 hours, 2 hours, 2.5 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, 6 hours, 6.5 hours, 7 hours, 7.5 hours, 8 hours, 8.5 hours, 9 hours, 9.5 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, or any range thereof, after administration of the anti-DOTA bispecific antibody. Alternatively, the radiolabeled DOTA hapten can be administered any time after 4 or more days after administration of the anti-DOTA bispecific antibody.

[0119] Additionally or alternatively, in some embodiments of the methods disclosed herein, the radiolabeled DOTA hapten can be administered any time between 1 minute to 4 or more days after administration of the clearing agent. For example, in some embodiments, the radiolabeled DOTA hapten is administered 1 minute, 2 minutes, 3 minutes, 4 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes, 35 minutes, 40 minutes, 45 minutes, 50 minutes, 55 minutes, 1 hour, 1.25 hours, 1.5 hours, 1.75 hours, 2 hours, 2.5 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, 6 hours, 6.5 hours, 7 hours, 7.5 hours, 8 hours, 8.5 hours, 9 hours, 9.5 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, or any range thereof, after administration of the clearing agent. Alternatively, the radiolabeled DOTA hapten may be administered any time after 4 or more days after administration of the clearing agent.

[0120] Reagent test kit

[0121] The technology of the present invention provides a kit containing components suitable for treating cancer in patients. On the one hand, the kit comprises a scavenger and instructions for use of the technology of the present invention. The kit may also comprise at least one anti-DOTA BsAb. In some embodiments, the at least one anti-DOTA BsAb binds to a tumor antigen target selected from the following: GPA33, HER2 / neu, GD2, MAGE-1, MAGE-3, BAGE, GAGE-1, GAGE-2, MUM-1, CDK4, N-acetylglucosamine transferase, p15, gp75, β-catenin, ErbB2, cancer antigen 125 (CA-125), carcinoembryonic antigen (CEA), RAGE, MART (melanoma antigen), MUC-1, MUC-2, MUC-3, MUC-4, MUC-5ac, MUC-16, MUC-17, tyrosinase, Pmel 17 (gp100), GnT-V intron V sequence (N-acetylglucosamine transferase V intron V sequence), prostate cancer psm, PRAME (melanoma antigen), β-catenin, EBNA (Epstein-Barr virus nuclear antigen) 1-6, p53, lung resistance protein (LRP) Bcl-2, prostate specific antigen (PSA), and Ki-67. Additionally or alternatively, in some embodiments, the at least one anti-DOTA BsAb binds to a tumor antigen target selected from the group consisting of CEACAM6, colon-specific antigen-p (CSAp), HLA-DR, CD40, CD74, CD138, EGFR, EGP-1, EGP-2, VEGF, PlGF, insulin-like growth factor (ILGF), tenascin, platelet-derived growth factor, IL-6, CD20, CD19, PSMA, CD33, CD123, MET, DLL4, Ang-2, HER3, IGF-1R, CD30, TAG-72, SPEAP, CD45, L1-CAM, Lewis Y (Lewis Y y ) antigen, E-cadherin, V-cadherin and EpCAM. The at least one anti-DOTA BsAb can be provided in the form of a pre-filled syringe or autoinjector pen containing a sterile liquid formulation or lyophilized formulation of the antibody (e.g., Kivitz et al., Clin. Ther. 28: 1619-29 (2006)).

[0122] Additionally or alternatively, in some embodiments, the kit further comprises a DOTA hapten, optionally labeled with one or more radionuclides. Examples of DOTA haptens include, but are not limited to, Proteus-DOTA, S-2-(R-aminobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (DOTA-Bn), DOTA-Bn-biotin, BAD (((S)-2-(4-(2-bromo)-acetamido)-benzyl)-DOTA), NBD ((S)-2-(4-nitrobenzyl)-DOTA), DOTA-RGD, DOTA-PEG-E (c(RGDyK))2, DOTA-8-AOC-BBN, p-NO2-Bn-DOTA, DOTA-PESIN, DOTA-biotin-sarcosine (DOTA-biotin), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid mono(N-hydroxysuccinimide ester) (DOTA-NHS), DOTATyrLysDOTA, and the like. Additionally or alternatively, in some embodiments of the kits of the present technology, the one or more radionuclides are selected from 213 Bi, 211 At 225 Ac, 152 Dy, 212 Bi, 223 Ra, 219 Rn, 215 Po, 211 Bi, 221 Fr. 217 At and 255 Additionally or alternatively, in certain embodiments, the one or more radionuclides are selected from 86 Y. 90 Y. 89 Sr. 165 Dy, 186 Re, 188 Re, 177 Lu, 67 Cu, 111 In, 67 Ga, 51 Cr, 58 Co. 99m Tc, 103m Rh, 195m Pt, 119 Sb, 161 Ho, 189m Os, 192 Ir, 201 Tl, 203 Pb, 68 Ga, 227 Th and64 Cu.

[0123] If the kit components are not formulated for oral administration, a device capable of delivering the kit components by some other route may be included. Examples of such devices include syringes (for parenteral administration) or inhalation devices.

[0124] The kit components can be packaged together or separately in two or more containers. In some embodiments, the container can be a vial containing a sterile lyophilized formulation suitable for reconstruction of a scavenger, DOTA hapten and / or anti-DOTA BsAb composition disclosed herein. The kit can also contain one or more buffers suitable for reconstruction and / or dilution of other reagents. Other containers that can be used include but are not limited to pouches, trays, boxes, tubes, etc. The kit components can be aseptically packaged and maintained in the container.

[0125] Example

[0126] Example 1: Exemplary Synthesis of Compounds of the Present Technology

[0127] Overview. DOTA-Bn-isothiocyanate (p-SCN-Bn-DOTA) was purchased from Macrocyclics, Inc. (Plano, TX), and amine-PEG4-DOTA was purchased from CheMatech (Dijon, France). Optima TM Grade hydrochloric acid was purchased from ThermoFisher Scientific (Waltham, MA). Chelex-100 resin (200-400 mesh) was purchased from Bio-Rad Laboratories (Hercules, CA). A PD-10 gel filtration size exclusion column (containing 8.3 mL of Sephadex TM G-25 resin / column) was purchased from GE Healthcare Life Sciences (Pittsburgh, PA). All other reagents and synthetic grade chemicals were purchased from Sigma-Aldrich (St. Louis, MO) and used without further purification. All solvents used for HPLC analysis (HPLC grade) and compound purification were also purchased from Thermo Fisher Scientific (Waltham, MA). All buffers and solutions were prepared using ultrapure water (resistivity of 18 MΩ-cm).

[0128] All liquid chromatography mass spectrometry (LC / MS) data were obtained using a Waters Autopure system (Milford, MA) including the following instruments: 2767 Sample Manager, 2545 Binary Gradient Module, System Fluid Manager, 2424 Evaporative Light Scattering Detector, 2998 Photodiode Array Detector, 3100 Mass Detector. HPLC solvents (Solvent A, 0.05% TFA in water; Solvent B, 0.05% TFA in acetonitrile) were filtered prior to use. The analytical method was solvent B from 5% to 25% in 10 min at a flow rate of 1.2 mL / min. Analytical columns: Waters XBridge BEH300 (Milford, MA), C4, 3.5 μm, 4.6 x 50 mm and C18, 4 μm, 4.6 x 50 mm. Preparative method: solvent B from 5% to 25% in 30 min at a flow rate of 20 mL / min. Preparative column: Waters XBridge Prep (Milford, MA) C18, 4 μm, optimal bed density, 19 x 150 mm.

[0129] All NMR data were obtained using a Bruker AV500 or AV600 instrument (Bruker, Billerica, MA) at ambient temperature. The following abbreviations are used: singlet (s), broad singlet (bs), doublet (d), triplet (t), quartet (q), quintet (p), doublet of doublets (dd), multiplet (m).

[0130] All experiments involving molecules with high metal complexing ability (such as DOTA) were carried out in glassware, which was pre-washed with metal-free HCl, rinsed with high purity water (e.g., glass distilled water), and oven dried. Chromatography was carried out on a manually filled glass column to avoid leaching or extracting metal from the metal column wall when loading complexing agents. Reverse phase purification was carried out on a clean, metal-free glass column manually filled with loose C-18 silica gel. The water content in the final complex was not measured.

[0131] CCA-16-methylester-NHBoc (described below) and 5-((tert-butoxycarbonyl)amino)pentyl-2-acetamido-2-deoxy-1-thio-3,4,6-tri-O-acetyl-α-D-galactopyranoside (“mono-peracetyl-sugar-NHBoc”) were prepared according to similar methods and protocols described in Yoo, B. et al. “N-AcetylgalactosaminoDendrons as Clearing Agents to Enhance Liver Targeting of Model Antibody-Fusion Protein,” Bioconjugate Chem. 2013, 24, 2088-2103 (and accompanying supporting information) and Cheal, SM et al. “Evaluation of Glycodendron and Synthetically Modified DextranClearing Agents for Multistep Targeting of Radioisotopes for Molecular Imaging and Radioimmunotherapy,” Mol. Pharmaceutics 2014, 11, 400-16 (and accompanying supporting information), each of which is incorporated herein by reference. The following provides relevant characterization data for CCA-16-methylester-NHBoc and mono-peracetyl-sugar-NHBoc.

[0132] CCA-16-methyl ester-NHBoc:

[0133]

[0134] 1 H NMR(600MHz,D2O)δ:3.68,3.66(2s,48H,OCH3),3.30-3.26(m,32H),3.22-3.19(m,32H),2.83( bs,NCH3),2.35-2.25(m,64H),1.69-1.51(m,128H),1.44(s,9H,C(CH3)3),1.34-1.29(m,64H). 13C NMR(125MHz,D2O)δ:174.10,173.87,172.10,51.57,51.48,47.86,47.79,45.78,45.70,33.95,33.84,33.83,33 .05,32.89,29.21,28.90,28.50,27.70,27.48,27.46,26.99,26.84,26.55,26.42,25.23,25.13,24.68,24.64.

[0135] Mono-peracetyl-sugar-NHBoc (5-((tert-butoxycarbonyl)amino)pentyl-2-acetamido-2-deoxy-1-thio-3,4,6-tri-O-acetyl-α-D-galactopyranoside): 1 H NMR (600MHz, D2O) δ: 5.57 (d, 1H, J = 7.4Hz), 5.49 (d, 1H, J = 4.4Hz), 5.38 (d, 1H, J = 2.4Hz), 5.05 (d d,1H,J=2.4Hz,J=7.4Hz),4.80-4.74(m,1H),4.58(bs,1H),4.54(t,1H,J=5.4Hz),4.13(dd,1H,J =9.6Hz,J=5.4Hz),4.09(dd,1H,J=9.6Hz,J=5.4Hz),3.14-3.08(m,2H),2.66-2.55(m,2H),2.16 ,2.05,2.01,1.98(4s,12H,COCH3),1.67-1.61(m,2H),1.44(s,9H,C(CH3)3),1.41-1.38(m,2H). 13 C NMR(125MHz,D2O)δ:171.00,170.39,170.28,170.15,155.98,84.93,68.54,67.34,6 7.24,61.83,48.33,40.38,30.97,29.62,29.28,28.43,25.94,23.34,20.75,20.71.

[0136] Exemplary Synthesis

[0137] CCA-16-Acid:

[0138]

[0139] To CCA-16-methyl ester-NHBoc (2.1g, 0.54mmol) in MeOH (60mL), add NaOH (10N, 16mL) and water (16mL).The mixture obtained is stirred at room temperature for 1h.Then with 2.0N HCl, pH is adjusted to 5.0, and solution is loaded in separating funnel.Extract (150mL x 5) with DCM / tert-butyl alcohol (3 / 1, v / v), and the organic layer merged is through Na2SO4 short-term drying.After filtering through diatomite bed, the filtrate is evaporated under reduced pressure, and residue is further dried 2h under high vacuum, to provide 2.0g colorless thick oil.Product has enough purity to be directly presented to next step.

[0140] CCA-16-sugar-NHBoc:

[0141]

[0142] CCA-16-acid (2.0g, 0.54mmol) is diluted in DMF (80mL), then treated with HATU (4.0g, 10.5mmol) and 5-amino-1-pentyl-α-thiotetraacetylgalactosamine (5.6g, 9.9mmol). The resulting mixture is stirred at room temperature for 20min, then DIPEA (8.0mL) is added. After 1h, DCM (200mL) is added, and the reaction mixture is washed with water (2X100mL). The organic layer is then dried over Na2SO4 for a short time, filtered through a diatomaceous earth bed, and the filtrate is concentrated under vacuum. CCA-16-sugar-NHBoc (4.9g, 85% yield) is obtained by flash column chromatography purification using 0% to 15% MeOH (v / v) gradient in DCM. 1H NMR (600MHz, D2O) δ: 7.95 (bs, 12H, NH), 5.64-5.61 (m, 16H), 5.47-5.46 (d, 16H, J = 2.8Hz), 5.06 (dd, 16H, J = 11 .8Hz),4.64-4.60(m,32H),4.18-4.12(m,32H),3.35(m,32H),3.21-3.19(m,36H),3.02,2.88(2s,3H,NCH3), 2.86-2.84(bs,4H),2.70-2.61(m,32H),2.41-2.38(m,36H),2.24(m,32H),2.21,2.06,1.98,1.97(4s,192H, COCH3),1.69-1.64(m,128H,1.59-1.53(m,70H),1.48(s,9H,OC(CH3),1.48-1.46(m,32),1.40-1.33(m,70H). 13 C NMR(125MHz,D2O)δ:174.60,174.38,174.30,173.41,172.23,172.14,170.66,170 .34,84.07,68.23,67.23,66.80,61.78,53.46,45.67,39.01,38.88,35.73,35.63 ,35.58,32.59,32.51,30.03,29.06,28.66,28.54,27.57,27.08,26.37,26.28,26.22,26.10,25.83,25.45,25.36,25.18,25.10,21.26,21.22,19.44,19.37,19.24.

[0143] CCA-16-peracetyl-α-thiogalactosamine-NH2·TFA:

[0144]

[0145] 20mg CCA-16-peracetyl-α-thiogalactosamine-NHBoc (1.9 μmol) in 20%TFA (v / v) in 0.8mL of dichloromethane is stirred at room temperature for 60min, and then the mixture is evaporated under reduced pressure to remove TFA and DCM. Monitor the reaction progress by LCMS, use 5%-95% (v / v) acetonitrile (containing 0.05%TFA) gradient operation 10min in water on the Autopure System (Waters) using C4 (Xbridge, Waters, 4.6X50mm). Ensure detection by accompanying mass spectrometer, diode array and evaporative light scattering detection. Residue (20mg) is further dried overnight under high vacuum, and directly used in next step.

[0146] CCA-16-peracetyl-α-thiogalactosamine-NHCSNMe-Bn-DOTA:

[0147]

[0148] The crude ammonium salt (CCA-16-peracetyl-α-thiogalactosamine-NH2·TFA) and P-SCN-Bn-DOTA (2.5 mg, 3.8 μmol, 2 equivalents) were dissolved in DMF (0.3 mL) and Et3N (6 μmL) was added. The resulting homogeneous mixture was stirred at room temperature for 4 h, and the volatiles were removed under high vacuum at room temperature to give a foam with sufficient purity for the next step. LCMS evaluation was run on the same system as described in the previous step, but the difference was that an XBridgeC18 column (4.6×150 mm) was used and a 40%-95% (v / v) acetonitrile (containing 0.05% TFA) gradient in water was used.

[0149] CCA-16-α-Thio-peracetylgalactosamine-DOTA-Y 3+ :

[0150]

[0151] The crude material CCA-16-peracetyl-α-thiogalactosamine-NHCSNMe-Bn-DOTA from the previous step was added to a solution of YCl3·6H2O (6 mg) in 0.05M HCl (0.2 mL) and 0.5M NH4OAc (0.2 mL). The mixture was shaken for 4 h. Attempts to purify by HPLC at this stage were unsuccessful. However, analytical monitoring showed that the reaction was completed with reasonable purity. LCMS was run on an XBridge C18 column using a 5%-95% (v / v) acetonitrile in water (both containing 0.05% TFA) gradient system for 10 min. The residue was presented to the next and final step. ESI-MS (m / z): [M+6H] 6+ Calculated value 1,847.55, Observed value 1,847.86

[0152] CCA-16-DOTA-Y 3+ :

[0153]

[0154] Crude CCA-16-α-thio-peracetylgalactosamine-DOTA-Y 3+ Dissolved in MeOH (0.5mL), and degassed NaOH (0.2N, 400 μL) was added to the solution. After 30 min, the reaction mixture was evaporated to dryness and dissolved in a minimum amount of water, and loaded onto HPLC (XBridge C18, acetonitrile gradient in water 20%-70% (v / v) (both containing 0.05% TFA), 10 min). Appropriate fractions were combined and lyophilized to obtain a total of 22 mg of the target compound in a white foam, and the overall yield from CCA-16-methyl ester-NHBoc was 64%. 1 H NMR(600MHz,D2O)δ:7.21-7.24(m,4H,Ph),5.43(d,16H,J=5.2Hz,H 糖 -1),4.27(dd,16H,J=5.2Hz,J=11.2Hz,H 糖 -2),4.17-4.14(m,16H,H 糖 -5),3.96(d,16H,J=2.7Hz,H 糖 -4),3.75(dd,16H,J=2.7Hz,J=11.2Hz,H 糖 -3),3.68(d,32H,J=6.0Hz,H 糖-6),3.23(bs,64H),3.15-3.08(m,36H),3.00 and 2.85(2s,3H,NCH3),2.58-2.47(m,34H),2.30(b s,32H),2.16-2.13(m,34H),1.96(s,48H,NHCOCH3),1.55-1.30(m,198H),1.23-1.15(m,102H). 13 C NMR(125MHz,D2O)δ:175.94,174.24,163.04,162.81,117.31,115.38,83.84,71.62,68.46,67.77,61.07,50.29, 48.22,45.86,39.19,35.78,32.74,30.26,28.77,28.24,27.13,26.90,26.22,25.86,25.80,25.51,25.39,22.09. ESI-MS(m / z):[M+5H] 5+ Calculated 1,812.2, observed 1,813.1; [M+6H] 6+ Calculated value 1,510.3, observed value 1,511.2.

[0155] Example 2: Exemplary Experimental Materials and Methods for Biological Studies

[0156] Materials. Anti-GPA33 DOTA-PRIT BsAb (huA33-C825) was prepared as previously described in WO 2016 / 130539. The GPA33(+) human colorectal cancer cell line SW1222 was obtained from the Ludwig Institute for Cancer Immunotherapy (New York, NY) and maintained in minimal essential medium supplemented with 10% heat-inactivated fetal bovine serum, 2.0 mM glutamine, 100 units / mL penicillin, and 100 μg / mL streptomycin.

[0157] Radioiodination of BsAbs. 131I was received from Nordion (Ottawa, Ontario, Canada). Pre-coated iodogen tubes were obtained from Thermo Fisher Scientific (Waltham, Massachusetts). Pre-filled Sephadex G-25 columns (PD10 columns) were obtained from GE Healthcare Life Sciences (Pittsburgh, Pennsylvania). Aliquots of anti-GPA33 BsAb were radioiodinated according to the IODO-GEN method (Salacinski PR et al., Analytical biochemistry 117: 136-46 (1981); Harlow E, Lane D. USING ANTIBODIES: A LABORATORY MANUAL. Cold Spring Harbor, New York (1999)), followed by gel filtration purification using commercial pre-filled PD10 columns (composed of Sephadex G-25 resin). Radiochemical purity > 98% was determined using size exclusion high pressure liquid chromatography with external radiation detection. In short, in order to prepare 131 I-huA33-C825, 100 μg was mixed with 80 μL of 0.2 M sodium phosphate (pH 7.4) in a pre-coated iodogen tube. Next, 400 μCi of 131 I. After 5 minutes at room temperature, the reaction was transferred to a new tube containing 50 μL of iodogen stop buffer [10 mg / mL tyrosine (saturated), 10% glycerol, 0.1% xylene cyanol blue in PBS] and purified using a PD10 column pre-equilibrated and eluted with added saline + 1% (w / v) BSA.

[0158] Animal Model. Female athymic nude mice (6-8 weeks old) were obtained from Harlan (Indianapolis, IN) or Envigo (Huntington, UK). Mice were allowed to acclimate for a minimum of 1 week. 3+ right 131 Normal (tumor-free) mice were used to investigate the effect of blood clearance of I-BsAb. For the DOTA-PRIT biodistribution experiment, a group of mice bearing subcutaneous (sc) SW1222 human colorectal cancer xenografts were used. To establish SW1222 tumors, 5.0×10 cells were injected subcutaneously in the lower flank with 200 μL of a 1:1 mixture of culture medium and reconstituted basement membrane (BD Matrigel, Collaborative Biomedical Products Inc., Bedford, Massachusetts). 6Mice were inoculated with 100 cells and established tumors (100-300 mm3) were observed within 7-10 days.

[0159] 131 Blood clearance of I-BsAb. 131 I-BsAb was mixed with additional carrier antibody to achieve 250 μg (1.19 nmol) per dose and antibody tracer was prepared for injection. All blood sampling and injections were performed via the tail vein. 250 μg (1.19 nmol) was administered intravenously to normal mice (n=15) at t=0. 131 I-BsAb and vehicle (saline; n=3) or 25 μg scavenger (formulated in 250 μL saline; n=12) was administered at t=24h. For vehicle, blood was sampled at t=1, 2, 3, 4, 24, 25, 26, 27 and 28h (30-40 μL; n=3 / time point). For scavenger, blood was sampled at t=1, 2, 3, 4, 24, 24.1 (5 min after CA injection), 24.3 (15 min after CA injection), 24.5, 25, 26, 27 and 28h (30-40 μL; n=3 / time point). The radioactivity concentration of each sample was determined by counting in a gamma counter (PerkinElmer Life Sciences Wallac Wizard 3). The count rate was background-corrected and decay-corrected, converted to activity using the system calibration factor specific to the isotope, normalized to the administered activity, and expressed as percent injected dose / g (ID / g). The percent change in blood activity over the time interval from baseline (ie, just before scavenger injection; y1) to after scavenger injection (y2) was calculated using the formula ((y2-y1) / y1)*100.

[0160] Dendrimer scavenger CCA-16-DOTA-Y during DOTA-PRIT 3+ Dose optimization. Tumor-bearing mice (n = 4 / group) were injected with BsAb (250 μg, 1.19 nmol; t = -28 h), followed by injection of different amounts of CCA-16-DOTA-Y 3+ Another group of tumor-bearing animals was given a dextran scavenger (62.5 μg; 0.125 l; 7.625 nmol (Y)DOTA) instead of the dendrimer scavenger CCA-16-DOTA-Y 3+ Four hours after administration of the scavenger, mice were injected with 177 Lu-DOTA-Bn (150-167 μCi; 30-33 pmol). Injection 177The mice were killed 48h after Lu activity and the organs were removed to determine the radioactivity concentration as described above. These organs included blood, tumor, heart, liver, spleen, stomach, small intestine, large intestine, kidney, muscle, bone and tail (injection site). The ratio of the injected dose percentage / g of tumor to non-tumor was also calculated.

[0161] [ 111 In] Preparation of Proteus-DOTA. Figure 8 The structure of Proteus-DOTA is shown (chemical formula: C 50 H 80 LqCy 11 O 19 S 3- ; Exact mass: 1345.48; Molecular weight: 1346.28). See also International Application No. PCT / US18 / 40911 filed on July 5, 2018, incorporated herein by reference. For the DOTA-PRIT study, from [ 111 In] indium chloride (153 MBq [4.14 mCi]) and 1 μL of 10 mg / mL Proteus-DOTA (10 μg; 7.42 nmole) were prepared [ 111 In]Proteus-DOTA. 111 In] Proteus-DOTA yield was 44%, and the final specific activity (SA) was 7178 GBq / g [194 Ci / g] or 9.67 E6 GBq / mol [2.61 E5 Ci / mol]. Prior to administration to mice, the Strata TM -X box (33μm polymer reverse phase C-18 30mg / 1mL #8B-S100-TAK, Inc., Torrance, California, USA) to purify [ 111 In] Proteus-DOTA, and the radiochemical purity was verified to be >98% using an in vitro binding assay with excess BsAb or by analytical reverse-phase HPLC with added radiodetection.

[0162] Statistical Analysis. Differences between means were determined using the unpaired Student's t test.

[0163] SPECT / CT imaging. For anti-GPA33-DOTA-PRIT+[ 111 SPECT / CT imaging and biodistribution studies after in vivo targeting of Proteus-DOTA were performed in patients who had received huA33-C825 BsAb and CCA-16-DOTA-Y 3+(25 μg; 2.76 nmol) of SW1222 tumor-bearing mice injected with 172 pmol / 1.67 MBq [45 μCi] of 111 In] 1 (n = 4) or 790pmol / 7.66MBq [207μCi] 111 In]1 (n = 1). The next day, 20 hours after injection, SPECT / CT was performed to give a larger administration activity [ 111 Single mice were imaged for In]Proteus-DOTA, and all animals were sacrificed 24 h after injection for biodistribution studies.

[0164] Example 3: The scavengers of the present invention enhance the blood flow of radiolabeled bispecific antibodies (BsAb) in vivo Liquid Clear

[0165] right 131 I-BsAb( 131 In vivo experiments were performed using normal (tumor-free) nude mice to evaluate the effect of a single dose of an excess of dendrimer CA of the present invention on the blood clearance of 131 Effect of blood clearance of I-BsAb.

[0166] Animals were initially injected with 250 μg of 131 I-BsAb, which is the BsAb dose previously optimized for DOTA-PRIT. Twenty-four hours later, mice were injected with dendrimers of the present technology, CA (25 μg, 2.76 nmol) or vehicle control. 131 Serial blood samples were collected at different times (28 hours after I-BsAb) and the concentration of 1g / mL in each sample was determined by measuring in a gamma counter. 131 IActive concentration.

[0167] as follows Figure 1 As shown, the blood levels of CA group and vehicle group at baseline (24 h after injection) 131 There was no significant difference in I activity initially. CA effectively reduced the circulating 131 I-BsAb, as the mean blood activity concentration at 5 minutes after injection decreased significantly by 64%, from 6.7% ID / g to 2.4% ID / g. At 1 hour after injection of CA or vehicle, the mean blood activity was 5.8 (-13%) or 1.3% ID / g (-81%), respectively.

[0168] These results demonstrate that the depleting agents of the present technology may be used in methods of increasing the sensitivity of a tumor to radiation therapy and / or treating cancer in a subject in need thereof.

[0169] Example 4: The scavenger of the present invention can be used for DOTA PRIT Method

[0170] Figure 2 The results show that different doses of CCA-16-DOTA-Y 3+ The results of tumor pretargeting were achieved. Dendrimer CA doses of 5-25 μg resulted in 177 The average tumor size at 24 h after Lu activity 177 Lu-DOTA-Bn uptake was 27-41% ID / g, which was consistent with the tumor uptake observed in the case of dextran CA and vehicle groups. 177 Lu-DOTA-Bn uptake (about 35% ID / g) was comparable. A 25 μg dose of dendrimer CA (CCA-16-DOTA-Y 3+ ) resulted in a tumor: blood uptake ratio comparable to that observed with the 62.5 μg dose of dextran CA. N-acetylgalactosamine dendrimer scavenger CCA-16-DOTA-Y 3+ The effect on tumor uptake is dose-dependent. Figure 3 .also, Figure 4 A comparison of tumor pretargeting results achieved with the clearing agents of the present technology compared to no clearing agent (vehicle) and with a 500 kD dextran-DOTA hapten conjugate clearing agent (i.e., dextran CA) is shown. 3+ The tumor:normal tissue uptake ratios achieved in the case of were comparable to those observed in the case of dextran CA.

[0171] To prove [ 225 Ac]Proteus-DOTA can be used in combination with DOTA-PRIT for efficient in vivo tumor targeting, and a group of nude mice bearing SW1222 xenografts expressing GPA33 were administered [ 225 BsAb huA33-C825 (250 μg; 1.19 nmol) was injected intravenously 28 h before administration of [ 225 The mice were injected with scavengers (62.5 μg; 0.125 nmol dextran; 7.625 nmol (Y)DOTA) 4 h before [Ac]Proteus-DOTA (182 pmol, 1.85 kBq [50 nCi]). 225 Ac]Proteus-DOTA was used to determine the biodistribution of the cells. 111 In]Proteus-DOTA(172pmol / 1.67MBq[45μCi][111 In] Proteus-DOTA (n = 4) or 790 pmol / 7.66 MBq [207 μCi] [ 111 In]Proteus-DOTA(n=1), use CCA-16-DOTA-Y 3+ The study was repeated at 20 h after injection. 111 Single mice were imaged for In]Proteus-DOTA, and all animals were sacrificed 24 h after injection for biodistribution studies.

[0172] As shown in Figure 5(A), for the experience with [ 225 For those animals that received pretargeted radioimmunotherapy with Ac]Proteus-DOTA, blood, tumor, and kidney uptake (expressed as percentage of injected activity per gram of tissue; %IA / g) at 24 h post-injection were 0.94±0.26, 16.71±2.95, and 1.08±0.55, respectively, corresponding to tumor-to-organ activity ratios of approximately 18:1 and 16:1 for blood and kidney, respectively. 111 In]Proteus-DOTA, the blood, tumor, and kidney uptake at 24 h after injection were 0.76 ± 0.09, 13.18 ± 0.97, and 1.02 ± 0.06, respectively, corresponding to tumor-to-organ activity ratios of approximately 17:1 and 13:1 in the blood and kidney, respectively. 225 Ac]Proteus-DOTA and 111 No significant differences were observed between Proteus-DOTA and liver [ 225 The uptake of Ac]Proteus-DOTA was about 3 times higher (for [ 225 Ac]Proteus-DOTA or [ 111 In]Proteus-DOTA were 1.40±0.47 vs 0.46±0.05; P<0.05), and bone 111 In]Proteus-DOTA uptake was higher (for [ 225 Ac]Proteus-DOTA or [ 111 In] Proteus-DOTA, they were undetectable vs. 0.15 ± 0.01; P < 0.001). These tumor and organ viability ratios were similar to those using the tracer 177 Lu-DOTA-Bn or 86Previous biodistribution studies of Y-DOTA-Bn with anti-GPA33-DOTA-PRIT in the same animal model were similar, with average tumor uptake of both DOTA-haptens at 24 h post-injection of approximately 8% ID / g (1.85-8.8 MBq; 10-50 pmol for either M-DOTA-Bn hapten) (see Cheal, SM et al. Eur J Nucl Med Mol Imaging 43:925-937 (2016)), suggesting that C825 has a significant effect on [ 225 The affinity of Ac]Proteus-DOTA is similar. See Figure 5(B).

[0173] Figure 6(A) shows the results of the injection of 172pmol / 1.67MBq [45μCi] of 111 In] Proteus-DOTA (n = 4) or 790 pmol / 7.66 MBq [207 μCi] [ 111 Results of tumor pretargeting achieved in animals treated with Proteus-DOTA (n=1). 111 In animals treated with In]Proteus-DOTA, the tumor to normal tissue ratio was generally higher. See Figure 6 (A) and Figure 7 .

[0174] As shown in Figure 6(B), 225 Ac]Proteus-DOTA (used in combination with dextran CA) and [ 111 In]Proteus-DOTA(with CCA-16-DOTA-Y 3+ The anti-GPA33-DOTA-PRIT results achieved with both 1:1 and 2:1 of the PRIT assays (using both 1:1 and 2:1 of the PRIT assays) were comparable.

[0175] These results demonstrate that the depleting agents of the present technology may be used in methods of increasing the sensitivity of a tumor to radiation therapy and / or treating cancer in a subject in need thereof.

[0176] Example 5: DOTA-pretargeted radioimmunotherapy using N-acetylgalactosamine dendrimer scavengers

[0177] The main hypothesis was tested that the dendrimer CA, CCA-16-DOTA-Y 3+ Anti-GPA33 PRIT+ 177 Lu-aminobenzyl DOTA ([ 177 Lu]LuDOTA-Bn) can replace dextran CA to achieve a high therapeutic index (TI). The therapeutic index is defined herein as the estimated tumor / normal tissue absorbed dose ratio.

[0178] Including dendrimer CA, namely CCA-16-DOTA-Y 3+ PRIT+ 177 Estimation of absorbed dose for Lu]LuDOTA-Bn: To calculate the PRIT+[ 177 The absorbed dose of Lu]LuDOTA-Bn was administered to nude mice (n=5 / group) bearing SW1222 xenografts expressing GPA33. 177 BsAb huA33-C825 (250 μg; 1.19 nmol) was injected intravenously 28 h before administration of [Lu]LuDOTA-Bn (20 pmol, 3.7 MBq [100 μCi]) and 177 The dendrimer scavenger CCA-16-DOTA-Y was injected intravenously 4 h before Lu]LuDOTA-Bn (20 pmol, 3.7 MBq [100 μCi]). 3+ (25 μg; 2.76 nmol). 177 The samples were sacrificed 1, 4, 24 or 48 h after administration of Lu]LuDOTA-Bn for biodistribution determination. For each tissue, the time-activity concentration data without decay correction were fitted to a single component, two components or more complex exponential function using Excel as appropriate, and analytical integration was performed to obtain the cumulative activity concentration per unit of administered activity (MBq-h / g / MBq). 177 Lu equilibrium dose constant (8.49 g-cGy / MBq-h) estimates tumor-to-tumor and selected organ-to-organ self-absorption doses, assuming only 177 The local absorption of Luβ-rays is complete, and the contribution of γ-rays and non-self-dose is negligible. Fig. 9 The results obtained from these studies are shown.

[0179] Fig.10 It was shown that the injection 177 Decay-corrected time-activity curves for tumor, blood, liver, spleen, kidney, and muscle up to 48 h after Lu]LuDOTA-Bn. Fig.11 The absorbed dose (cGy / MBq) and therapeutic index for various tissues are shown. Fig.11 As shown, the estimated [ 177 The absorbed doses for the Lu]LuDOTA-Bn were 11.7, 468.4, 9.97, 5.49, and 13.3 cGy / MBq, respectively. The ratio of the estimated absorbed dose of the tumor to the estimated absorbed dose of selected normal tissues (i.e., TI) ranged from about 40 (e.g., for blood and kidney) to 550 (e.g., for muscle). Fig.11 ).

[0180] For the optimized PRIT+[ 177 Lu]LuDOTA-Bn, the estimated absorbed doses (cGy / MBq) of tumor, blood, liver, spleen and kidney under single cycle PRIT were 65.8, 0.9 (TI 73), 6.3 (TI 10), 6.6 (TI10) and 5.3 (TI 12), respectively. See Cheal et al., Eur J Nucl Med Mol Imaging 43:925 (2016). Effective treatment was demonstrated, with no major treatment-related toxicity observed (e.g., 111.0 MBq delivered in a fractionated dosing strategy produced a CR frequency of 100%, including two of nine mice surviving for more than 140 days; absorbed doses of tumor, blood and kidney were 7304, 100 and 588 cGy). Cheal et al., Eur J Nucl Med Mol Imaging 43:925 (2016).

[0181] Based on estimates of human normal tissue radiation dose tolerance derived from clinical observations (Marks et al., Int J Radiat Oncol Biol Phys. 76: S10-9 (2010)), the maximum tolerated dose (MTD) is 250 cGy for bone marrow, 1,500 cGy for lung, 3,000 cGy for liver, and 2,000 cGy for kidney. 177 Lu]LuDOTA-Bn, the most tolerated pretargeted [ 177 Lu]LuDOTA-Bn activity is 278MBq, with the bone marrow being the dose limiting organ. At this activity, the estimated absorbed dose delivered to the tumor would be 18,292cGy (183Gy), with 250cGy to the blood (bone marrow) and 1,473cGy to the kidney.

[0182] For the dendrimer CA, namely CCA-16-DOTA-Y 3+ PRIT+ 177 Lu]LuDOTA-Bn, the most tolerated pretargeted [ 177Lu]LuDOTA-Bn activity is 21MBq, with the bone marrow being the dose-limiting organ. At this activity, the estimated absorbed dose delivered to the tumor will be 9836cGy (98Gy), and 246cGy to the blood (bone marrow) and 280cGy to the kidney. Therefore, due to the tumor absorbed dose of about 73Gy, an effective and safe CRC therapy is predicted in xenografts in mice, which can be achieved with an administered activity of 15.6MBq (and 183cGy to the blood (bone marrow) and 207cGy to the kidney).

[0183] Ultimately, these data demonstrate that dendrimer CAs can be used to treat inflammatory conditions with less administration compared to dextran CAs. 177 Cure was achieved in a mouse model of human CRC without Lu activity (e.g., 70 Gy to tumor) with differences in dosimetry to critical tissues (e.g., reduced dose to kidney).

[0184] These results demonstrate that the depleting agents of the present technology may be used in methods of increasing the sensitivity of a tumor to radiation therapy and / or treating cancer in a subject in need thereof.

[0185] Equivalent

[0186] The technology of the present invention is not limited to the specific embodiments described in this application, and the embodiments are intended to be a single description of the individual aspects of the technology of the present invention. It will be clear to those skilled in the art that many modifications and changes can be made to the technology of the present invention without departing from the spirit and scope of the technology of the present invention. It will be clear to those skilled in the art from the foregoing description that, in addition to the methods and equipment listed herein, functionally equivalent methods and equipment within the scope of the technology of the present invention. Such modifications and changes are intended to fall within the scope of the technology of the present invention. It should be understood that the technology of the present invention is not limited to specific methods, reagents, compounds, compositions or biological systems, and of course they can change. It should also be understood that the terms used herein are only used for the purpose of describing specific embodiments and are not intended to be restrictive.

[0187] In addition, where features or aspects of the disclosure are described in terms of Markush groups, those skilled in the art will recognize that the disclosure is also thereby described in terms of any individual member or subgroup of members of the Markush group.

[0188] It will be understood by those skilled in the art that for any and all purposes, particularly with respect to providing a written description, all ranges disclosed herein also encompass any and all possible sub-ranges and combinations of sub-ranges thereof. Any listed range can be easily identified as fully describing the same range and enabling the same range to be decomposed into at least equal halves, thirds, quarters, fifths, tenths, etc. As a non-limiting example, each range discussed herein can be easily decomposed into a lower third, a middle third, and an upper third, etc. It will also be understood by those skilled in the art that all words such as "up to", "at least", "greater than", "less than", etc. include the numbers and relate to the ranges that can subsequently be decomposed into sub-ranges as discussed above. Finally, it will be understood by those skilled in the art that a range includes each individual member. Therefore, for example, a group having 1-3 cells refers to a group having 1, 2, or 3 cells. Similarly, a group having 1-5 cells refers to a group having 1, 2, 3, 4, or 5 cells, etc.

[0189] All patents, patent applications, provisional applications, and publications mentioned or cited herein are incorporated by reference in their entirety, including all figures and tables, to the extent they are consistent with the explicit teachings of this specification.

Claims

1. A compound, wherein the compound is or a pharmaceutically acceptable salt and / or solvate thereof, wherein X 1 , X 2 , X 3 , X 4 , X 5 , X 6 , X 7 , X 8 , X 9 , X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 , X 17 , X 18 , X 19 and X 20 Each is independently H or a lone pair of electrons; Y 1 , Y 2 , Y 3 , Y 4 and Y 5 each independently is O or S; x is 1, 2, or 3; and y is 1, 2, 3, or 4.

2. A scavenger compound, the scavenger compound being or a pharmaceutically acceptable salt and / or solvate thereof, wherein M 1 、M 2 、M 3 、M 4 and M 5 Each is independently Lu 3+ Sc 3+ , Ga 3+ , Y 3+ 、In 3+ ,La 3+ 、Ce 3+ 、Eu 3+ , Tb 3+ or Gd 3+ ; X 1 , X 2 , X 3 , X 4 , X 5 , X 6 , X 7 , X 8 , X 9 , X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 , X 17 , X 18 , X 19 and X 20 Each is independently H or a lone pair of electrons; Y 1 , Y 2 , Y 3 , Y 4 and Y 5 each independently is O or S; x is 1, 2, or 3; and y is 1, 2, 3, or 4.

3. The scavenger compound according to claim 2, wherein M 1 、M 2 、M 3 、M 4 and M 5 Each independently is not a radionuclide.

4. The scavenger compound according to any one of claims 1 to 3, wherein Y 1 , Y 2 , Y 3 , Y 4 and Y 5 Each independently is S.

5. The scavenger compound according to any one of claims 1 to 3, wherein x is 1 or 2.

6. The scavenger compound according to any one of claims 1 to 3, wherein y is 2 or 3.

7. A composition comprising the compound according to claim 1 or the scavenger compound according to any one of claims 2 to 6 and a pharmaceutically acceptable carrier.

8. Use of a scavenger compound according to any one of claims 2 to 7 in the preparation of a kit for use in a method of increasing the sensitivity of a tumor to radiotherapy in a subject diagnosed with cancer, wherein the method comprises (a) administering to the subject an effective amount of an anti-DOTA bispecific antibody comprising the single-chain variable fragment C825scFv, wherein the anti-DOTA bispecific antibody is configured to localize to a tumor expressing a tumor antigen target; (b) administering to the subject an effective amount of a clearing agent according to any one of claims 2-7; and (c) administering to the subject an effective amount of a radiolabeled DOTA hapten, wherein the DOTA hapten is configured to form a complex with the anti-DOTA bispecific antibody.

9. Use of a scavenger compound according to any one of claims 2 to 7 in the preparation of a kit for use in a method of treating cancer in a subject in need thereof, wherein the method comprises (a) administering to the subject an effective amount of an anti-DOTA bispecific antibody comprising the single-chain variable fragment C825scFv, wherein the anti-DOTA bispecific antibody is configured to localize to a tumor expressing a tumor antigen target; (b) administering to the subject an effective amount of a clearing agent according to any one of claims 2-7; and (c) administering to the subject an effective amount of a radiolabeled DOTA hapten, wherein the DOTA hapten is configured to form a complex with the anti-DOTA bispecific antibody.

10. The use according to claim 9, wherein the method further comprises administering to the subject sequentially, separately or simultaneously at least one chemotherapeutic agent selected from the group consisting of anthracyclines, antibiotics, enzyme inhibitors, platinum coordination complexes, adrenocortical inhibitors, hormone antagonists, antimetabolites, alkylating agents, antimitotic agents, antiangiogenic agents, mTOR inhibitors, proteasome inhibitors, and pro-apoptotic agents.

11. The use according to claim 9, wherein the method further comprises administering to the subject sequentially, separately or simultaneously at least one chemotherapeutic agent selected from the group consisting of a tyrosine kinase inhibitor, a heat shock protein (HSP90) inhibitor and an HDAC inhibitor.

12. The method of claim 9, wherein the method further comprises administering to the subject sequentially, separately or simultaneously at least one chemotherapeutic agent selected from the group consisting of nitrogen mustards, alkyl sulfonates, nitrosoureas, gemcitabine, triazenes, taxanes, COX-2 inhibitors, epipodophyllotoxins, vinca alkaloids, endostatin, paclitaxel, camptothecin, SN-38, doxorubicin, methotrexate and CPT-11.

13. The method of claim 8 , wherein the tumor antigen target is selected from the group consisting of GPA33, HER2 / neu, GD2, MAGE-1, MAGE-3, BAGE, GAGE-1, GAGE-2, MUM-1, CDK4, N-acetylglucosamine transferase, p15, gp75, β-catenin, ErbB2, cancer antigen 125 (CA-125), carcinoembryonic antigen (CEA), RAGE, MART (melanoma antigen), MUC-1, MUC-2, MUC-3, MUC-4, MUC-5ac, MUC-16, MUC-17, tyrosinase, Pmel 17 (gp100), GnT-V intron V sequence (N-acetylglucosamine transferase V intron V sequence), prostate cancer psm, PRAME (melanoma antigen), β-catenin, EBNA (Epstein-Barr virus nuclear antigen) 1-6, p53, lung resistance protein (LRP) Bcl-2, prostate specific antigen (PSA), Ki-67, CEACAM6, colon specific antigen-p (CSAp), HLA-DR, CD40, CD74, CD138, EGFR, EGP-1, EGP-2, VEGF, PlGF, insulin-like growth factor (ILGF), tenascin, platelet-derived growth factor, IL-6, CD20, CD19, PSMA, CD33, CD123, MET, DLL4, Ang-2, HER3, IGF-1R, CD30, TAG-72, SPEAP, CD45, L1-CAM, Lewis Y (Le y ) antigens, E-cadherin, V-cadherin and EpCAM.

14. The use according to any one of claims 8-12, wherein the anti-DOTA bispecific antibody is administered intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intradermally, intraperitoneally, transtracheally, subcutaneously, intracerebroventricularly, orally or intranasally.

15. The use according to any one of claims 8 to 12, wherein the radiolabeled DOTA hapten is administered intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intradermally, intraperitoneally, transtracheally, subcutaneously, intracerebroventricularly, orally or intranasally.

16. The use according to any one of claims 8 to 12, wherein the clearing agent is administered intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intradermally, intraperitoneally, transtracheally, subcutaneously, intracerebroventricularly, orally or intranasally.

17. The method of any one of claims 8 to 12, wherein the cancer is selected from breast cancer, colorectal cancer, cervical cancer, ovarian cancer, liver cancer, bladder cancer, brain cancer, lung cancer, stomach cancer, pancreatic cancer, thyroid cancer, kidney cancer, prostate cancer, melanoma, sarcoma, endometrial cancer, salivary gland cancer, vulvar cancer, penile cancer, and head and neck cancer.

18. The use according to any one of claims 8 to 12, wherein the cancer is selected from Wilms tumor, squamous cell carcinoma and astrocytoma.

19. The use according to claim 17, wherein the brain cancer is pituitary adenoma, meningioma, neuroblastoma, glioblastoma or craniopharyngioma.

20. The use according to any one of claims 8 to 12, wherein the complex emits a level of radioactivity expressed as a percentage of the injected dose per gram of tissue.

21. The use of any one of claims 8-12, wherein the subject exhibits a tumor to normal tissue uptake ratio of 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 55:1, 60:1, 65:1, 70:1, 75:1, 80:1, 85:1, 90:1, 95:1, or 100:

1.

22. The use according to any one of claims 8 to 12, wherein the radiolabeled DOTA hapten is labeled with a radionuclide selected from: 213 Bi, 211 At 225 Ac, 152 Dy, 212 Bi, 223 Ra, 219 Rn, 215 Po, 211 Bi, 221 Fr. 217 At 255 Fm, 86 Y. 90 Y. 89 Sr. 165 Dy, 186 Re, 188 Re, 177 Lu, 67 Cu, 111 In, 67 Ga, 51 Cr, 58 Co. 99m Tc, 103m Rh, 195m Pt, 119 Sb, 161 Ho, 189m Os, 192 Ir, 201 Tl, 203 Pb, 68 Ga, 227 Th and 64 Cu.

23. The use according to claim 22, wherein the radiolabeled DOTA hapten comprises one or more of the following: Proteus-DOTA, S-2-(R-aminobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (DOTA-Bn), DOTA-Bn-biotin, BAD (((S)-2-(4-(2-bromo)-acetamido)-benzyl)-DOTA), NBD ((S)-2-(4-nitrobenzyl)- DOTA), DOTA-RGD, DOTA-PEG-E(c(RGDyK))2, DOTA-8-AOC-BBN, p-NO2-Bn-DOTA, DOTA-PESIN, DOTA-biotin-sarcosine (DOTA-biotin), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid mono(N-hydroxysuccinimide ester) (DOTA-NHS), or DOTATyrLysDOTA.

24. A kit comprising the compound of claim 1 or the scavenger compound of any one of claims 2-7 and instructions for use.

25. The kit according to claim 24, further comprising at least one anti-DOTAB sAb comprising the single-chain variable fragment C825scFv.

26. The kit of claim 25, wherein the at least one anti-DOTA BsAb binds to a tumor antigen target selected from the group consisting of GPA33, HER2 / neu, GD2, MAGE-1, MAGE-3, BAGE, GAGE-1, GAGE-2, MUM-1, CDK4, N-acetylglucosamine transferase, p15, gp75, β-catenin, ErbB2, cancer antigen 125 (CA-125), carcinoembryonic antigen (CEA), RAGE, MART (melanoma antigen), MUC-1, MUC-2, MUC-3, MUC-4, MUC-5ac, MUC-16, MUC-17, tyrosinase, Pmel 17 (gp100), GnT-V intron V sequence (N-acetylglucosamine transferase V intron V sequence), prostate cancer psm, PRAME (melanoma antigen), β-catenin, EBNA (Epstein-Barr virus nuclear antigen) 1-6, p53, lung resistance protein (LRP) Bcl-2, prostate specific antigen (PSA), Ki-67, CEACAM6, colon specific antigen-p (CSAp), HLA-DR, CD40, CD74, CD138, EGFR, EGP-1, EGP-2, VEGF, PlGF, insulin-like growth factor (ILGF), tenascin, platelet-derived growth factor, IL-6, CD20, CD19, PSMA, CD33, CD123, MET, DLL4, Ang-2, HER3, IGF-1R, CD30, TAG-72, SPEAP, CD45, L1-CAM, Lewis Y (Le y ) antigens, E-cadherin, V-cadherin and EpCAM.

27. The kit according to any one of claims 24-26, further comprising a DOTA hapten.

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