Pharmacokinetic enhancements of bifunctional chelates and uses thereof

Abstract

To provide conjugates including a chelating moiety of a metal complex and a therapeutic or targeting moiety, the conjugates having linkers that enhance excretion of the chelating moiety or the metal complex thereof.SOLUTION: The problem is solved by the invention described in the specification.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] Related Applications This application claims priority to and the benefit of U.S. Provisional Patent Application No. 62 / 502,260, entitled "Pharmacokinetic Enhancement of Bifunctional Chelates and Uses Thereof," filed May 5, 2017, the entire contents of which are incorporated herein by reference for all purposes. [Background technology]

[0002] Radiolabeled targeting moieties, or radioconjugates, are typically prepared by using bifunctional chelators to attach radiolabels to biomolecules while maintaining target affinity. Structurally, bifunctional chelates may contain a chelate, a linker, and a bridging group or targeting moiety. Modifications to the linker region of bifunctional chelates can provide pharmacokinetic advantages, such as increased elimination of radioactivity. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] WO92 / 22324 [Patent Document 2] WO98 / 46645 [Patent Document 3] U.S. Patent No. 6,180,370 [Non-patent literature]

[0004] [Non-Patent Document 1] Greene, Protective Groups in Organic Synthesis, 3rd edition (John Wiley & Sons, New York, 1999). [Non-patent document 2] Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977 [Non-patent document 3] Pharmaceutical Salts: Properties, Selection, and Use (PH Stahl and CG Wermuth (eds.)), Wiley-VCH, 2008 [Non-licensed Document 4] Bioconjugate Chem. 2000, 11, pp. 510-519 [Non-licensed Document 5] Bioconjugate Chem. 2012, 23, pp. 1029-1039 [Non-licensed Document 6] Mol Imaging Biol (2011) 13:215~221 pages [Non-licensed Document 7] Bioconjugate Chem. 2002, 13, pp. 110-115 [Non-licensed Document 8] Bioconjugate Chem. 2006, 17, pp. 1551-1560 [Non-licensed Document 9] Bioconjugate Chem. 2003, 14, pp. 927-933 [Non-licensed Document 10] Mol Cancer Ther; 12(11) November 2013, Methods in Molecular Biology, 2009, 539, pages 191~211 [Non-licensed Document 11] Bioconjugate chemistry, Volume 14, Issue 5, pages 927~33 (2003) [Non-licensed Document 12] Quadri and Vriesendorp [QJ Nucl. Med. 1998, 42, pp. 250-261] [Non-licensed Document 13] Wardら(1989)Nature 341:544~546 pages [Non-licensed Document 14] Harlow et al., Antibodies: A Laboratory Manual (Cold Spring Harbor Laboratory Press, 2nd ed., 1988) [Non-Patent Document 15] Brinkman et al., 1995, J. Immunol. Methods 182:41-50 [Non-Patent Document 16] Morrison, 1985, Science 229:1202 p. [Non-Patent Document 17] Segal et al., J. Immunol. Methods 248:1-6 (2001) [Non-Patent Document 18] Tutt et al., J. Immunol. 147: 60 (1991) [Non-Patent Document 19] Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, PA, 17th edition, 1985 [Non-Patent Document 20] Langer (Science 249:1527~1533, 1990) Summary of the Invention [Problem to be solved by the invention]

[0005] The present invention relates to linkers that enhance the excretion of a chelating moiety, or a metal complex thereof, when conjugated to a therapeutic moiety, a targeting moiety, or a bridging group. [Means for solving the problem]

[0006] Thus, in a first aspect, the present invention provides a compound having the structure: AL 1 -(L 2 ) n -B Formula I wherein A is a chelating moiety or a metal complex thereof; L 1is an optionally substituted C1-C6 alkyl, an optionally substituted C1-C6 heteroalkyl, an optionally substituted aryl or heteroaryl; B is a therapeutic moiety, targeting moiety, or bridging group, or a pharmaceutically acceptable salt thereof; n is 1 to 5; L 2 each independently represents the structure: (-X 1 -L 3 -Z 1 -) Formula II (In the formula, X 1 is C=O(NR 1 ), C=S(NR 1 ), O-C=O(NR 1 ), NR 1 C=O(O), NR 1 C=O(NR 1 ), -CH2PhC=O(NR 1 ), -CH2Ph(NH)C=S(NR 1 ), O, NR 1 and R 1 is H or optionally substituted C1-C6 alkyl or optionally substituted C1-C6 heteroalkyl, optionally substituted aryl or heteroaryl; L 3 is optionally substituted C1 to C 50 Alkyl or optionally substituted C1-C 50 Heteroalkyl or C5-C 20 Polyethylene glycol; Z 1 are CH2, C=O, C=S, OC=O, NR 1 C=O, NR 1 and R 1 is hydrogen, optionally substituted C1-C6 alkyl, pyrrolidine-2,5-dione) have] The present invention is characterized by a compound having the formula:

[0007] In some embodiments, the chelating moiety is selected from the group consisting of DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), DOTMA ((1R,4R,7R,10R)-α,α',α'',α'''-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), DOTAM (1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane), DOTPA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrapropionic acid), DOAM-acetic acid (2-(4,7,10-tris(2-amino-2-oxoethyl)-1,4,7,10-tetraazacyclododecane)), and DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid). -1-yl)acetic acid), DOTA-GA anhydride (2,2',2''-(10-(2,6-dioxotetrahydro-2H-pyran-3-yl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid), DOTP (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylenephosphonic acid)), DOTMP (1,4,6,10-tetraazacyclodecane-1,4,7,10-tetramethylenephosphonic acid), DOTA-4AMP (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(acetamido-methylenephosphonic acid)), CB-TE2A (1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-diacetic acid), NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid), NOTP (1,4,7-triazacyclononane-1,4,7-tri(methylenephosphonic acid)), TETPA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetrapropionic acid), TETA (1,4,8,11-tetraazacyclotetradecane- 1,4,8,11-tetraacetic acid), HEHA (1,4,7,10,13,16-hexaazacyclohexadecane-1,4,7,10,13,16-hexaacetic acid), PEPA (1,4,7,10,13-pentaazacyclopentadecane-N,N',N'',N''',N''''-pentaacetic acid), H4Octapa (N,N'-bis(6-carboxy-2-pyridylmethyl)-ethylenediamine-N,N '-diacetic acid), H2Dedpa (1,2-[[6-(carboxy)-pyridin-2-yl]-methylamino]ethane), H6phospa (N,N'-(methylenephosphonate)-N,N'-[6-(methoxycarbonyl)pyridin-2-yl]-methyl-1,2-diaminoethane), TTHA (triethylenetetramine-N,N,N',N'',N''',N'''-hexaacetic acid), DO2P (tetraazacyclododecanedimethanephosphonic acid), HP-DO3A (hydroxypropyltetraazacyclododecanetriacetic acid), EDTA (ethylenediaminetetraacetic acid), deferoxamine, DTPA (diethylenetriaminepentaacetic acid), DTPA-BMA (diethylenetriaminepentaacetic acid-bismethylamide), HOPO (octadentate hydroxypyridinone), or porphyrin.

[0008] Those skilled in the art will appreciate that the use of chelating moieties in the practice of the present invention is not limited to the specific constructs disclosed herein, but rather may include other known chelating moieties.

[0009] In some embodiments, the chelating moiety has the structure:

[0010] [ka]

[0011] (In the formula, Y 1 is -CH2OCH2(L 2 ) n -B, C=O(L 2 ) n -B, or C=S(L 2 ) n -B and Y 2 is -CH2CO2H; In the formula, Y 1 is H and Y 2 L 1 -(L 2 ) n -B) It has.

[0012] In some embodiments, L 1 The structure:

[0013] [ka]

[0014] (In the formula, R 2 is optionally substituted hydrogen or -COH) It has.

[0015] In some embodiments, the metal can be selected from Bi, Pb, Y, Mn, Cr, Fe, Co, Zn, Ni, Tc, In, Ga, Cu, Re, Sm, a lanthanide, or an actinide for use as an imaging or therapeutic agent. Particular examples of radionuclides suitable for complexation with compounds of formula (I) include: 47 Sc, 55 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 66 Ga, 67 Ga, 68 Ga, 82 Rb, 86 Y, 87 Y, 90 Y, 97 Ru, 105 Rh,109 Pd, 111 In, 117m Sn, 149 Pm, 149 Tb, 153 Sm, 177 Lu, 186 Re, 188 Re, 199 Au, 201 Tl, 203 Pb, 212 Pb, 212 Bi, 213 Bi, 225 Ac, and 227 Th is an example.

[0016] In some embodiments, B is a therapeutic moiety or a targeting moiety.

[0017] In some embodiments, the therapeutic or targeting moiety is an antibody, an antigen-binding fragment thereof, or other targeting proteins such as nanobodies, affibodies, and consensus sequences derived from fibronectin type III domains.

[0018] In some embodiments, the antibody, or antigen-binding fragment thereof, specifically binds to insulin-like growth factor-1 receptor (IGF-1R).

[0019] In some embodiments, the crosslinking group is an amino-reactive crosslinking group, a methionine-reactive crosslinking group, a thiol-reactive crosslinking group, or a sortase-mediated coupling sequence.

[0020] In some embodiments, the amino-reactive, methionine-reactive, or thiol-reactive crosslinking group comprises an activated ester such as a hydroxysuccinimide ester, N-hydroxysulfosuccinimide, 2,3,5,6-tetrafluorophenol ester, 4-nitrophenol ester, or imidate, an anhydride, a thiol, a disulfide, a maleimide, an azide, an alkyne, a strained alkyne, a strained alkene, a halogen, a sulfonate, a haloacetyl, an amine, a hydrazide, a diazirine, a phosphine, a tetrazine, an isothiocyanate, or an oxaziridine.

[0021] In some embodiments, the sortase recognition sequence may include a terminal glycine-glycine-glycine (GGG) and / or LPTXG amino acid sequence, where X is any amino acid.

[0022] Those skilled in the art will appreciate that the use of cross-linking groups in the practice of the present invention is not limited to the specific constructs disclosed herein, but rather may include other known cross-linking groups.

[0023] In some embodiments, the bridging group is

[0024] [ka]

[0025] is selected from the group consisting of:

[0026] In some embodiments, Y 1 is H.

[0027] In some embodiments, X 1 is C=O(NR 1 ) and R 1 is H.

[0028] In some embodiments, Z 1 is -CH2.

[0029] In some embodiments, L 2 has an n value of 1.

[0030] In some embodiments, the compound is

[0031] [ka]

[0032] is selected from the group consisting of:

[0033] In some embodiments, the metal is a radionuclide.

[0034] In some embodiments, the radionuclide is 111 In.

[0035] In some embodiments, the radionuclide is 68 It's Ga.

[0036] In some embodiments, the radionuclide is 86 It's Y.

[0037] In some embodiments, the metal is a beta-emitting radionuclide.

[0038] In some embodiments, the radionuclide is 67 Cu, 177 Lu' or 90 It's Y.

[0039] In some embodiments, the metal is an alpha-emitting radionuclide.

[0040] In some embodiments, the radionuclide is 225 Ac, 212 Pb, 227 Th or its progeny (daughter isotope).

[0041] In another aspect, the invention features a pharmaceutical composition including any of the above compounds and a pharmaceutically acceptable excipient.

[0042] In another aspect, the invention features a method of radiation treatment planning and / or treatment comprising administering to a subject in need thereof any of the above compounds or pharmaceutical compositions.

[0043] In another aspect, the invention features a method of detecting and / or treating cancer, comprising administering to a subject in need thereof a first dose of any of the compounds or pharmaceutical compositions in an amount effective for a radiation treatment regimen, followed by subsequent doses of therapeutically effective amounts of any of the compounds or pharmaceutical compositions.

[0044] In some embodiments, the compound or composition administered in the first dose and the compound or composition administered in the second dose are the same.

[0045] In some embodiments, the compound or composition administered in the first dose and the compound or composition administered in the second dose are different.

[0046] In some embodiments, the cancer is a solid tumor or a blood (liquid) cancer.

[0047] In some embodiments, the solid tumor cancer is breast cancer, non-small cell lung cancer, small cell lung cancer, pancreatic cancer, head and neck cancer, prostate cancer, colorectal cancer, sarcoma, adrenocortical carcinoma, neuroendocrine carcinoma, Ewing's sarcoma, multiple myeloma, or acute myeloid leukemia.

[0048] In some embodiments, the method further comprises administering an antiproliferative agent, a radiosensitizing agent, or an immunomodulatory or immunomodulatory agent.

[0049] In some embodiments, the compound or any of its compositions and the antiproliferative or radiosensitizing agent are administered within 28 days (eg, within 14, 7, 6, 5, 4, 3, 2, or 1 day) of each other.

[0050] In some embodiments, the compound or any of its compositions and the immunomodulatory agent or immunomodulatory agent are administered within 90 days (e.g., within 80, 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2, or 1 day) of each other.

[0051] In another aspect, the invention features a method for making a radioconjugate (e.g., any of the radioconjugates described herein). The method includes: (a) conjugating a bifunctional chelate to a biomolecule; (b) purifying the conjugate produced by step (a); and (c) chelating one or more radionuclides (e.g., one or more Ac-225 radionuclides) with the purified conjugate of step (b) at a temperature below 35°C (e.g., 20-25°C) to produce the radioconjugate (e.g., an actinium radioconjugate).

[0052] In some embodiments, the radioconjugate is a radioimmunoconjugate (eg, any of the radioimmunoconjugates described herein).

[0053] In some embodiments, the pH of the reaction mixture of the conjugation step (a) is less than 6.4 (e.g., less than or equal to 6.3, 6.2, 6.1, 6.0, 5.9, or 5.8).

[0054] In some embodiments, the pH of the reaction mixture of the conjugation step (c) is less than 5.5 (e.g., 5.4, 5.3, 5.2, 5.1, or 5.0 or less) or greater than 7.0 (e.g., 7.1, 7.2, 7.3, 7.4, 7.5 or more).

[0055] In some embodiments, the temperature of the reaction mixture for conjugation step (c) is between 20 and 34°C (e.g., 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C, 30°C, 31°C, 32°C, 33°C, or 34°C).

[0056] Chemical terminology: The term "acyl," as used herein, refers to a hydrogen or an alkyl group, as defined herein (e.g., a haloalkyl group), attached to the parent molecular group through a carbonyl group, as defined herein, and is exemplified by formyl (i.e., a carboxaldehyde group), acetyl, trifluoroacetyl, propionyl, butanoyl, and the like. Exemplary unsubstituted acyl groups contain 1 to 7, 1 to 11, or 1 to 21 carbons. In some embodiments, the alkyl group is further substituted with 1, 2, 3, or 4 substituents described herein.

[0057] The term "alkyl," as used herein, unless otherwise specified, includes both straight-chain and branched-chain saturated groups of 1 to 20 carbons (e.g., 1 to 10 or 1 to 6). Alkyl groups are exemplified by methyl, ethyl, n- and isopropyl, n-, sec-, iso- and tert-butyl, neopentyl, and the like; (1) C 1~6 Alkoxy; (2) C 1~6 (3) amino, as defined herein (e.g., unsubstituted amino (i.e., —NH) or substituted amino (i.e., —N(R N1 )2(wherein, R N1 is as defined for amino); (4) C 6~10 Aryl-C 1~6 Alkoxy; (5) Azido; (6) Halo; (7) (C 2~9 (8) hydroxy optionally substituted with an O-protecting group; (9) nitro; (10) oxo (e.g., carboxaldehyde or acyl); (11) C 1~7 Spirocyclyl; (12) Thioalkoxy; (13) Thiol; (14) -CO2R optionally substituted with an O-protecting group A' (In the formula, R A' is (a)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c) C 6~10 Aryl, (d) hydrogen, (e) C 1~6 ALC-C 6~10 Aryl, (f) Amino-C1~20 Alkyl, (g) (CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20 (h)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (15) —C(O)NR B' R C' (In the formula, R B' and R C' are each independently (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 aryl); (16) -SO2R D' (In the formula, R D' is (a)C 1~6 Alkyl, (b) C 6~10 Aryl, (c) C 1~6 ALC-C 6~10 (17) -SO2NR E' R F' (In the formula, R E' and R F' are each independently (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 Aryl and (d) C 1~6 ALC-C 6~10aryl); (18) —C(O)R G' (In the formula, R G' is (a)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c) C 6~10 Aryl, (d) hydrogen, (e) C 1~6 ALC-C 6~10 Aryl, (f) Amino-C 1~20 Alkyl, (g)-(CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20 (h)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (19)-NR H' C(O)R I' (In the formula, R H' is (a1) hydrogen and (b1) C 1~6 alkyl; R I' is (a2)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b2) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c2) C 6~10 Aryl, (d2) hydrogen, (e2) C 1~6 ALC-C 6~10Aryl, (f2) Amino-C 1~20 Alkyl, (g2)-(CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20 (h2)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (20)-NR J' C(O)OR K' (In the formula, R J' is (a1) hydrogen and (b1) C 1~6 alkyl; R K' is (a2)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b2) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c2) C 6~10 Aryl, (d2) hydrogen, (e2) C 1~6 ALC-C 6~10 Aryl, (f2) Amino-C 1~20 Alkyl, (g2)-(CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20(h2)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (21) amino-polyethylene glycols (wherein the alkyl group is an alkyl group); and (22) amidines. In some embodiments, each of these groups can be further substituted as described herein. For example, the alkylene group of a C1-alkaryl can be further substituted with an oxo group to provide the corresponding aryloyl substituent.

[0058] The term "alkylene" and the prefix "alk-" as used herein refer to a saturated divalent hydrocarbon group derived from a straight or branched chain saturated hydrocarbon by the removal of two hydrogen atoms and are exemplified by methylene, ethylene, isopropylene, and the like. x~y Alkylene" and the prefix "C x~y "Alk-" represents an alkylene group having x to y carbons. Exemplary values ​​of x are 1, 2, 3, 4, 5, and 6, and exemplary values ​​of y are 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, or 20 (e.g., C 1~6 , C 1~10 , C 2~20 , C 2~6 , C 2~10 , or C 2~20 In some embodiments, the alkylene can be further substituted with 1, 2, 3, or 4 substituents, as defined herein, to form an alkyl group.

[0059] The term "alkenyl," as used herein, unless otherwise specified, represents a monovalent straight or branched chain group of 2 to 20 carbons (e.g., 2 to 6 or 2 to 10 carbons) containing one or more carbon-carbon double bonds, and is exemplified by ethenyl, 1-propenyl, 2-propenyl, 2-methyl-1-propenyl, 1-butenyl, 2-butenyl, and the like. Alkenyl includes both cis and trans isomers. Alkenyl groups are optionally substituted with 1, 2, 3, or 4 substituents independently selected from amino, aryl, cycloalkyl, or heterocyclyl (e.g., heteroaryl), as defined herein, or any of the exemplary alkyl substituents described herein.

[0060] The term "alkynyl," as used herein, refers to a monovalent straight or branched chain group of 2 to 20 carbon atoms (e.g., 2 to 4, 2 to 6, or 2 to 10 carbons) containing a carbon-carbon triple bond, and is exemplified by ethynyl, 1-propynyl, etc. Alkynyl groups are optionally substituted with 1, 2, 3, or 4 substituents independently selected from aryl, cycloalkyl, or heterocyclyl (e.g., heteroaryl), as defined herein, or any of the exemplary alkyl substituents described herein.

[0061] As used herein, the term "amino" refers to -N(R N1 )2(wherein, R N1 are independently H, OH, NO2, N(R N2 )2, SO2OR N2 , SO2R N2 , SOR N2, an N-protecting group, alkyl, alkenyl, alkynyl, alkoxy, aryl, alkaryl, cycloalkyl, alkcycloalkyl, carboxyalkyl (optionally substituted with an O-protecting group such as, for example, an optionally substituted arylalkoxycarbonyl group or any of those described herein), sulfoalkyl, acyl (e.g., acetyl, trifluoroacetyl, or others described herein), alkoxycarbonylalkyl (optionally substituted with an O-protecting group such as, for example, an optionally substituted arylalkoxycarbonyl group or any of those described herein), heterocyclyl (e.g., heteroaryl), or alkheterocyclyl (e.g., alkheteroaryl), wherein these described R N1 Each group may be optionally substituted as defined herein for each group; or two R N1 combine to form a heterocyclyl or N-protecting group, R N2 are each independently H, alkyl, or aryl. The amino group of the present invention can be an unsubstituted amino (i.e., —NH) or a substituted amino (i.e., —N(R N1 )2). In a preferred embodiment, amino may be -NH2 or NHR N1 (In the formula, R N1 are independently OH, NO2, NH2, and NR N2 2. SO2OR N2 , SO2R N2 , SOR N2 , alkyl, carboxyalkyl, sulfoalkyl, acyl (e.g., acetyl, trifluoroacetyl, or others described herein), alkoxycarbonylalkyl (e.g., t-butoxycarbonylalkyl), or aryl; R N2 are H and C, respectively. 1~20 Alkyl (e.g., C 1~6 alkyl), or C 6~10 It may be aryl).

[0062] The term "amino acid," as used herein, refers to a molecule having a side chain, an amino group, and an acid group (e.g., a carboxy group of -COH or a sulfo group of -SOH), where the amino acid is attached to the parent molecule by the side chain, amino group, or acid group (e.g., the side chain). In some embodiments, the amino acid is attached to the parent molecular group by a carbonyl group, where the side chain or amino group is attached to the carbonyl group. Exemplary side chains include optionally substituted alkyl, aryl, heterocyclyl, alkaryl, alkheterocyclyl, aminoalkyl, carbamoylalkyl, and carboxyalkyl. Exemplary amino acids include alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, hydroxynorvaline, isoleucine, leucine, lysine, methionine, norvaline, ornithine, phenylalanine, proline, pyrrolysine, selenocysteine, serine, taurine, threonine, tryptophan, tyrosine, and valine. 1~6 Alkoxy; (2) C 1~6 (3) amino, as defined herein (e.g., unsubstituted amino (i.e., —NH) or substituted amino (i.e., —N(R N1 )2(wherein, R N1 is as defined for amino); (4) C 6~10 Aryl-C 1~6 Alkoxy; (5) Azido; (6) Halo; (7) (C 2~9 (heterocyclyl)oxy; (8) hydroxy; (9) nitro; (10) oxo (e.g., carboxaldehyde or acyl); (11) C 1~7 Spirocyclyl; (12) Thioalkoxy; (13) Thiol; (14) -CO2R A' (In the formula, R A' is (a)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c) C 6~10 Aryl, (d) hydrogen, (e) C 1~6 ALC-C 6~10Aryl, (f) Amino-C 1~20 Alkyl, (g) (CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20 (h)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (15) —C(O)NR B' R C' (In the formula, R B' and R C' are each independently (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 aryl); (16) -SO2R D' (In the formula, R D' is (a)C 1~6 Alkyl, (b) C 6~10 Aryl, (c) C 1~6 ALC-C 6~10 (17) -SO2NR E' R F' (In the formula, R E' and R F' are each independently (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 Aryl and (d) C 1~6 ALC-C 6~10aryl); (18) —C(O)R G' (In the formula, R G' is (a)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c) C 6~10 Aryl, (d) hydrogen, (e) C 1~6 ALC-C 6~10 Aryl, (f) Amino-C 1~20 Alkyl, (g)-(CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20 (h)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (19)-NR H' C(O)R I' (In the formula, R H' is (a1) hydrogen and (b1) C 1~6 alkyl; R I' is (a2)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b2) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c2) C 6~10 Aryl, (d2) hydrogen, (e2) C 1~6 ALC-C 6~10Aryl, (f2) Amino-C 1~20 Alkyl, (g2)-(CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20 (h2)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (20)-NR J' C(O)OR K' (In the formula, R J' is (a1) hydrogen and (b1) C 1~6 alkyl; R K' is (a2)C 1~20 Alkyl (e.g., C 1~6 alkyl), (b2) C 2~20 Alkenyl (e.g., C 2~6 alkenyl), (c2) C 6~10 Aryl, (d2) hydrogen, (e2) C 1~6 ALC-C 6~10 Aryl, (f2) Amino-C 1~20 Alkyl, (g2)-(CH2) s2 (OCH2CH2) s1 (CH2) s3 OR' (wherein s1 is an integer of 1 to 10 (for example, 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (for example, 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R' is H or C 1~20(h2)-NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 are each independently hydrogen or optionally substituted C 1~6 (21) amino-polyethylene glycols (which are alkyl); and (22) amidines, which may be substituted with up to four substituents. In some embodiments, each of these groups may be further substituted as described herein.

[0063] The term "aryl" as used herein refers to a monocyclic, bicyclic, or polycyclic carbocyclic ring system having one or two aromatic rings and is exemplified by phenyl, naphthyl, 1,2-dihydronaphthyl, 1,2,3,4-tetrahydronaphthyl, anthracenyl, phenanthrenyl, fluorenyl, indanyl, indenyl, and the like; (1) C 1~7 Acyl (e.g., carboxaldehyde); (2) C 1~20 Alkyl (e.g., C 1~6 Alkyl, C 1~6 Alkoxy-C 1~6 Alkyl, C 1~6 Alkylsulfinyl-C 1~6 Alkyl, Amino-C 1~6 Alkyl, azido-C 1~6 Alkyl, (carboxaldehyde)-C 1~6 Alkyl, Halo-C 1~6 Alkyl (e.g., perfluoroalkyl), hydroxy-C 1~6 Alkyl, nitro-C 1~6 Alkyl or C 1~6 Thioalkoxy-C 1~6 alkyl); (3) C 1~20Alkoxy (e.g., C such as perfluoroalkoxy) 1~6 Alkoxy); (4) C 1~6 Alkylsulfinyl; (5)C 6~10 Aryl;(6)Amino;(7)C 1~6 ALC-C 6~10 Aryl; (8) Azide; (9) C 3~8 Cycloalkyl; (10)C 1~6 ALC-C 3~8 Cycloalkyl; (11) halo; (12) C 1~12 Heterocyclyl (e.g., C 1~12 Heteroaryl); (13) (C 1~12 (14) Hydroxy; (15) Nitro; (16) C 1~20 Thioalkoxy (e.g., C 1~6 Thioalkoxy);(17)-(CH2) q CO2R A' (wherein q is an integer of 0 to 4, and R A' is (a)C 1~6 Alkyl, (b) C 6~10 aryl, (c) hydrogen, and (d) C 1~6 ALC-C 6~10 aryl); (18)-(CH2) q CONR B' R C' (wherein q is an integer of 0 to 4, and R B' and R C' is (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 (19)-(CH2) q SO2R D' (wherein q is an integer of 0 to 4, and R D' is (a) alkyl, (b) C 6~10 aryl, and (c) alk-C 6~10 aryl); (20)-(CH2) q SO2NR E' R F' (wherein q is an integer of 0 to 4, and R E' and RF' are each independently (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 (21) thiol; (22) C 6~10 Aryloxy; (23)C 3~8 Cycloalkoxy; (24)C 6~10 Aryl-C 1~6 Alkoxy; (25)C 1~6 ALC-C 1~12 Heterocyclyl (e.g., C 1~6 ALC-C 1~12 Heteroaryl);(26)C 2~20 Alkenyl; and (27) C 2~20 and optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from the group consisting of alkynyl. In some embodiments, each of these groups can be further substituted as described herein. For example, the alkylene group of a C1-alkaryl or C1-alkheterocyclyl can be further substituted with an oxo group to provide the corresponding aryloyl and (heterocyclyl)oyl substituent.

[0064] The term "arylalkyl," as used herein, refers to an aryl group, as defined herein, attached to the parent molecular group through an alkylene group, as defined herein. Exemplary unsubstituted arylalkyl groups include those having 7 to 30 carbons (e.g., 7 to 16 or 7 to 20 carbons, e.g., C 1~6 ALC-C 6~10 Aryl, C 1~10 ALC-C 6~10 Aryl, or C 1~20 ALC-C 6~10 In some embodiments, alkylene and aryl can each be further substituted with 1, 2, 3, or 4 substituents as defined herein for the corresponding group. Other groups following the prefix "alk-" are defined in the same manner, with "alk" being a C 1~6 The chemical structures to which alkylene refers and is attached are as defined herein.

[0065] The term "carbonyl" as used herein refers to a C(O) group, which can also be represented as C=O.

[0066] The term "carboxy" as used herein means -CO2H.

[0067] As used herein, the term "cyano" refers to a -CN group.

[0068] The term "cycloalkyl," as used herein, unless otherwise specified, refers to a monovalent saturated or unsaturated non-aromatic cyclic hydrocarbon group of 3 to 8 carbons, exemplified by cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, bicyclic heptyl, and the like. When a cycloalkyl group contains one carbon-carbon double bond or one carbon-carbon triple bond, the cycloalkyl group can be referred to as a "cycloalkenyl" or "cycloalkynyl" group, respectively. Exemplary cycloalkenyl and cycloalkynyl groups include cyclopentenyl, cyclohexenyl, cyclohexynyl, and the like. Cycloalkyl groups of the present invention include (1) C 1~7 Acyl (e.g., carboxaldehyde); (2) C 1~20 Alkyl (e.g., C 1~6 Alkyl, C 1~6 Alkoxy-C 1~6 Alkyl, C 1~6 Alkylsulfinyl-C 1~6 Alkyl, Amino-C 1~6 Alkyl, azido-C 1~6 Alkyl, (carboxaldehyde)-C 1~6 Alkyl, Halo-C 1~6 Alkyl (e.g., perfluoroalkyl), hydroxy-C 1~6 Alkyl, nitro-C 1~6 Alkyl or C 1~6 Thioalkoxy-C 1~6 alkyl); (3) C 1~20 Alkoxy (e.g., C such as perfluoroalkoxy) 1~6 Alkoxy); (4) C 1~6 Alkylsulfinyl; (5)C6~10 Aryl;(6)Amino;(7)C 1~6 ALC-C 6~10 Aryl; (8) Azide; (9) C 3~8 Cycloalkyl; (10)C 1~6 ALC-C 3~8 Cycloalkyl; (11) halo; (12) C 1~12 Heterocyclyl (e.g., C 1~12 Heteroaryl); (13) (C 1~12 (14) Hydroxy; (15) Nitro; (16) C 1~20 Thioalkoxy (e.g., C 1~6 Thioalkoxy);(17)-(CH2) q CO2R A' (wherein q is an integer of 0 to 4, and R A' is (a)C 1~6 Alkyl, (b) C 6~10 aryl, (c) hydrogen, and (d) C 1~6 ALC-C 6~10 aryl); (18)-(CH2) q CONR B' R C' (wherein q is an integer of 0 to 4, and R B' and R C' is (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 (19)-(CH2) q SO2R D' (wherein q is an integer of 0 to 4, and R D' is (a)C 6~10 Alkyl, (b) C 6~10 aryl, and (c) C 1~6 ALC-C 6~10 aryl); (20)-(CH2) q SO2NR E' R F' (wherein q is an integer of 0 to 4, and R E' and R F' are each independently (a) hydrogen, (b) C 6~10 Alkyl, (c) C6~10 aryl, and (d) C 1~6 ALC-C 6~10 (21) thiol; (22) C 6~10 Aryloxy; (23)C 3~8 Cycloalkoxy; (24)C 6~10 Aryl-C 1~6 Alkoxy; (25)C 1~6 ALC-C 1~12 Heterocyclyl (e.g., C 1~6 ALC-C 1~12 Heteroaryl; (26) oxo; (27) C 2~20 Alkenyl; and (28)C 2~20 In some embodiments, each of these groups can be further substituted as described herein. For example, the alkylene group of a C1-alkaryl or C1-alkheterocyclyl can be further substituted with an oxo group to provide the corresponding aryloyl and (heterocyclyl)oyl substituents.

[0069] As used herein, the term "diastereomers" means stereoisomers that are not mirror images of each other and are not superimposable with respect to one another.

[0070] As used herein, the term "enantiomer" refers to each individual optically active form of a compound of the invention having an optical purity or enantiomeric excess (as determined by standard methods in the art) of at least 80% (i.e., at least 90% of one enantiomer and at most 10% of the other enantiomer), preferably at least 90% and more preferably at least 98%.

[0071] The term "halogen" as used herein refers to a halogen selected from bromine, chlorine, iodine, or fluorine.

[0072] As used herein, the term "heteroalkyl" refers to an alkyl group, as defined herein, in which one or two member carbon atoms are replaced with nitrogen, oxygen, or sulfur, respectively. In some embodiments, heteroalkyl groups can be further substituted with one, two, three, or four substituents as described herein for alkyl groups. As used herein, the terms "heteroalkenyl" and "heteroalkynyl" refer to alkenyl and alkynyl groups, as defined herein, respectively, in which one or two member carbon atoms are replaced with nitrogen, oxygen, or sulfur, respectively. In some embodiments, heteroalkenyl and heteroalkynyl groups can be further substituted with one, two, three, or four substituents as described herein for alkyl groups.

[0073] The term "heteroaryl," as used herein, refers to the subset of heterocyclyl, as defined herein, that is aromatic, i.e., that contains 4n+2 pi electrons in a monocyclic or polycyclic ring system. Exemplary unsubstituted heteroaryl groups are those of 1 to 12 (e.g., 1 to 11, 1 to 10, 1 to 9, 2 to 12, 2 to 11, 2 to 10, or 2 to 9) carbons. In some embodiments, heteroaryl is substituted with 1, 2, 3, or 4 substituents as defined for heterocyclyl groups.

[0074] The term "heteroarylalkyl" refers to a heteroaryl group, as defined herein, attached to the parent molecular group through an alkylene group, as defined herein. Exemplary unsubstituted heteroarylalkyl groups include those having 2 to 32 carbons (e.g., 2 to 22, 2 to 18, 2 to 17, 2 to 16, 3 to 15, 2 to 14, 2 to 13, or 2 to 12 carbons, e.g., C 1~6 ALC-C 1~12 Heteroaryl, C 1~10 ALC-C 1~12 Heteroaryl, or C 1~20 ALC-C 1~12In some embodiments, alkylene and heteroaryl can each be further substituted with 1, 2, 3, or 4 substituents as defined herein for the corresponding group. Heteroarylalkyl groups are a subset of heterocyclylalkyl groups.

[0075] The term "heterocyclyl," as used herein, unless otherwise specified, refers to a 5-, 6-, or 7-membered ring containing 1, 2, 3, or 4 heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur. Five-membered rings have 0 to 2 double bonds, and 6- and 7-membered rings have 0 to 3 double bonds. Exemplary unsubstituted heterocyclyl groups are those of 1 to 12 (e.g., 1 to 11, 1 to 10, 1 to 9, 2 to 12, 2 to 11, 2 to 10, or 2 to 9) carbons. The term "heterocyclyl" also refers to heterocyclic compounds having bridged polycyclic structures in which one or more carbons and / or heteroatoms bridge two non-adjacent members of a monocycle, e.g., a quinuclidinyl group. The term "heterocyclyl" includes bicyclic, tricyclic, and tetracyclic groups in which any of the above heterocycles is fused to one, two, or three carbon rings, such as an aryl ring, a cyclohexane ring, a cyclohexene ring, a cyclopentane ring, a cyclopentene ring, or another monocyclic heterocycle, such as indolyl, quinolyl, isoquinolyl, tetrahydroquinolyl, benzofuryl, benzothienyl, etc. Examples of fused heterocyclyls include tropane and 1,2,3,5,8,8a-hexahydroindolizine. Heterocycles include pyrrolyl, pyrrolinyl, pyrrolidinyl, pyrazolyl, pyrazolinyl, pyrazolidinyl, imidazolyl, imidazolinyl, imidazolidinyl, pyridyl, piperidinyl, homopiperidinyl, pyrazinyl, piperazinyl, pyrimidinyl, pyridazinyl, oxazolyl, oxazolidinyl, isoxazolyl, isoxazolidinyl, morpholinyl, thiomorpholinyl, and thiazolyl, including dihydro and tetrahydro forms in which one or more double bonds are reduced and replaced with hydrogen. , thiazolidinyl, isothiazolyl, isothiazolidinyl, indolyl, indazolyl, quinolyl, isoquinolyl, quinoxalinyl, dihydroquinoxalinyl, quinazolinyl, cinnolinyl, phthalazinyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, benzothiadiazolyl, furyl, thienyl, thiazolidinyl, isothiazolyl, triazolyl, tetrazolyl, oxadiazolyl (e.g., 1,2,3-oxadiazolyl), purinyl, thiadiazolyl (e.g., 1,2,3-thiadiazolyl), tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, dihydrothienyl, dihydroindolyl, dihydroquinolyl, tetrahydroquinolyl, tetrahydroisoquinolyl, dihydroisoquinolyl, pyranyl, dihydropyranyl, dithiazolyl, benzofuranyl, isobenzofuranyl, benzothienyl, and the like. Still other exemplary heterocyclyls include 2,3,4,5-tetrahydro-2-oxo-oxazolyl; 2,3-dihydro-2-oxo-1H-imidazolyl; 2,3,4,5-tetrahydro-5-oxo-1H-pyrazolyl (e.g., 2,3,4,5-tetrahydro-2-phenyl-5-oxo-1H-pyrazolyl); 2,3,4,5-tetrahydro-2,4-dioxo-1H-imidazolyl (e.g., 2,3,4,5-tetrahydro-2,4-dioxo-5-methyl-5-phenyl-1H-imidazolyl); 2,3-dihydro-2-thioxo-1,3,4-oxadiazolyl (e.g., 2,3-dihydro-2-thioxo-5-phenyl-1,3,4-oxadiazolyl); 4,5-Dihydro-5-oxo-1H-triazolyl (e.g., 4,5-dihydro-3-methyl-4-amino5-oxo-1H-triazolyl); 1,2,3,4-tetrahydro-2,4-dioxopyridinyl (e.g., 1,2,3,4-tetrahydro-2,4-dioxo-3,3-diethylpyridinyl); 2,6-dioxo-piperidinyl (e.g., 2,6-dioxo-3-ethyl-3-phenylpiperidinyl); 1,6-dihydro-6-oxopyridiminyl; 1,6-dihydro-4-oxopyrimidinyl (e.g., 2-(methylthio)-1,6-dihydro-4-oxo-5-methylpyrimidin-1-yl); 1,2,3,4-tetrahydro-2,4-dioxopyrimidinyl (e.g., 1,2,3,4-tetrahydro-2,4-dioxo-3-ethylpyrimidinyl); 1,6-dihydro-6-oxo-pyridazinyl (e.g., 1,6-dihydro-6-oxo-3-ethylpyridazinyl); 1,6-dihydro-6-oxo-1,2,4-triazinyl (e.g., 1,6-dihydro-5-isopropyl-6-oxo-1,2,4-triazinyl); 2,3-dihydro-2-oxo-1H-indolyl (e.g., 3,3-dimethyl-2,3-Dihydro-2-oxo-1H-indolyl and 2,3-dihydro-2-oxo-3,3'-spiropropane-1H-indol-1-yl; 1,3-dihydro-1-oxo-2H-iso-indolyl; 1,3-dihydro-1,3-dioxo-2H-iso-indolyl; 1H-benzopyrazolyl (e.g., 1-(ethoxycarbonyl)-1H-benzopyrazolyl); 2,3-dihydro-2-oxo-1H-benzimidazolyl (e.g., 3-ethyl-2,3-dihydro-2-oxo-1H-benzimidazolyl); 2,3-dihydro-2-oxo-benzoxazolyl (e.g., 5-chloro-2,3-dihydro-2-oxo-benzoxazolyl); 2,3-Dihydro-2-oxo-benzoxazolyl; 2-oxo-2H-benzopyranyl; 1,4-benzodioxanyl; 1,3-benzodioxanyl; 2,3-dihydro-3-oxo,4H-1,3-benzothiazinyl; 3,4-dihydro-4-oxo-3H-quinazolinyl (e.g., 2-methyl-3,4-dihydro-4-oxo-3H-quinazolinyl); 1,2,3,4-tetrahydro-2,4-dioxo-3H-quinazolyl (e.g., 1-ethyl-1,2,3,4-tetrahydro-2,4-dioxo-3H-quinazolyl); 1,2,3,6-tetrahydro-2,6-dioxo-7H-purinyl (e.g., 1,2,3,6-tetrahydro-1,3-dimethyl-2,6-dioxo-7H-purinyl); 1,2,3,6-tetrahydro-2,6-dioxo-1H-purinyl (e.g., 1,2,3,6-tetrahydro-3,7-dimethyl-2,6-dioxo-1H-purinyl); 2-oxobenzo[c,d]indolyl; 1,1-dioxo-2H-naphtha[1,8-c,d]isothiazolyl; and 1,8-naphthylenedicarboxamide. Further heterocycles include 3,3a,4,5,6,6a-hexahydro-pyrrolo[3,4-b]pyrrol-(2H)-yl and 2,5-diazabicyclo[2.2.1 ]heptan-2-yl, homopiperazinyl (or diazepanyl), tetrahydropyranyl, dithiazolyl, benzofuranyl, benzothienyl, oxepanyl, thiepanyl, azocanyl, oxecanyl, and thiocanyl. Heterocyclic groups also include those of the formula:

[0076] [ka]

[0077] wherein E' is selected from the group consisting of -N- and -CH-; F' is selected from the group consisting of -N=CH-, -NH-CH-, -NH-C(O)-, -NH-, -CH=N-, -CH-NH-, -C(O)-NH-, -CH=CH-, -CH-, -CHCH-, -CHO-, -OCH-, -O-, and -S-; and G' is selected from the group consisting of -CH- and -N-. Any of the heterocyclyl groups described herein can be substituted with: (1) C 1~7 Acyl (e.g., carboxaldehyde); (2) C 1~20 Alkyl (e.g., C 1~6 Alkyl, C 1~6 Alkoxy-C 1~6 Alkyl, C 1~6 Alkylsulfinyl-C 1~6 Alkyl, Amino-C 1~6 Alkyl, azido-C 1~6 Alkyl, (carboxaldehyde)-C 1~6 Alkyl, Halo-C 1~6 Alkyl (e.g., perfluoroalkyl), hydroxy-C 1~6 Alkyl, nitro-C 1~6 Alkyl or C 1~6 Thioalkoxy-C 1~6 alkyl); (3) C 1~20 Alkoxy (e.g., C such as perfluoroalkoxy) 1~6 Alkoxy); (4) C 1~6 Alkylsulfinyl; (5)C 6~10 Aryl;(6)Amino;(7)C 1~6 ALC-C 6~10 Aryl; (8) Azide; (9) C 3~8 Cycloalkyl; (10)C 1~6 ALC-C 3~8 Cycloalkyl; (11) halo; (12) C 1~12 Heterocyclyl (e.g., C 2~12 Heteroaryl); (13) (C1~12 (14) Hydroxy; (15) Nitro; (16) C 1~20 Thioalkoxy (e.g., C 1~6 Thioalkoxy);(17)-(CH2) q CO2R A' (wherein q is an integer of 0 to 4, and R A' is (a)C 1~6 Alkyl, (b) C 6~10 aryl, (c) hydrogen, and (d) C 1~6 ALC-C 6~10 aryl); (18)-(CH2) q CONR B' R C' (wherein q is an integer of 0 to 4, and R B' and R C' is (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 (19)-(CH2) q SO2R D' (wherein q is an integer of 0 to 4, and R D' is (a)C 1~6 Alkyl, (b) C 6~10 aryl, and (c) C 1~6 ALC-C 6~10 aryl); (20)-(CH2) q SO2NR E' R F' (wherein q is an integer of 0 to 4, and R E' and R F' are each independently (a) hydrogen, (b) C 1~6 Alkyl, (c) C 6~10 aryl, and (d) C 1~6 ALC-C 6~10 (21) thiol; (22) C 6~10 Aryloxy; (23)C 3~8 Cycloalkoxy; (24) Arylalkoxy; (25) C 1~6 ALC-C 1~12 Heterocyclyl (e.g., C1~6 ALC-C 1~12 Heteroaryl; (26) oxo; (27) (C 1~12 Heterocyclyl)imino; (28)C 2~20 Alkenyl; and (29)C 2~20 and alkynyl. In some embodiments, each of these groups can be further substituted as described herein. For example, the alkylene group of a C1-alkaryl or C1-alkheterocyclyl can be further substituted with an oxo group to provide the corresponding aryloyl and (heterocyclyl)oyl substituent.

[0078] As used herein, the term "hydrocarbon" refers to a group consisting solely of carbon and hydrogen atoms.

[0079] As used herein, the term "hydroxyl" refers to an -OH group. In some embodiments, the hydroxyl group can be substituted with 1, 2, 3, or 4 substituents (e.g., O-protecting groups) as defined herein for alkyl.

[0080] As used herein, the term "isomer" refers to any tautomer, stereoisomer, enantiomer, or diastereomer of any of the compounds of the invention. The compounds of the invention may possess one or more chiral centers and / or double bonds and therefore exist as stereoisomers, such as double bond isomers (i.e., geometric E / Z isomers) or diastereomers (e.g., enantiomers (i.e., (+) or (-)) or cis / trans isomers). In accordance with the present invention, the chemical structures depicted herein, and thus the compounds of the invention, encompass all corresponding stereoisomers, i.e., both stereomerically pure forms (e.g., geometrically pure, enantiomerically pure, or diastereoisomerically pure), and mixtures of enantiomers and stereoisomers, e.g., racemates. Mixtures of enantiomers and stereoisomers of the compounds of the invention can typically be resolved into their component enantiomers or stereoisomers by known methods such as chiral-phase gas chromatography, chiral-phase high performance liquid chromatography, crystallization of the compound as a chiral salt complex, or crystallization of the compound in a chiral solvent. Enantiomers and stereoisomers can also be obtained from stereomerically or enantiomerically pure intermediates, reagents, and catalysts by known asymmetric synthetic methods.

[0081] The term "N-protected amino" as used herein refers to an amino group, as defined herein, that is bound to one or two N-protecting groups, as defined herein.

[0082] As used herein, the term "N-protecting group" refers to a group intended to protect an amino group against undesired reactions during synthetic procedures. Commonly used N-protecting groups are disclosed in Greene, "Protective Groups in Organic Synthesis," 3rd Edition (John Wiley & Sons, New York, 1999), which is incorporated herein by reference.N-protecting groups include acyl, aryloyl, or carbamyl groups such as formyl, acetyl, propionyl, pivaloyl, t-butylacetyl, 2-chloroacetyl, 2-bromoacetyl, trifluoroacetyl, trichloroacetyl, phthalyl, o-nitrophenoxyacetyl, α-chlorobutyryl, benzoyl, 4-chlorobenzoyl, 4-bromobenzoyl, and 4-nitrobenzoyl, and chiral auxiliary groups such as protected or unprotected D, L, or D,L-amino acids such as alanine, leucine, and phenylalanine; sulfonyl-containing groups such as benzenesulfonyl and p-toluenesulfonyl; benzyloxycarbonyl, p-chlorobenzyloxycarbonyl, p-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl, 2-nitrobenzyloxycarbonyl, p-bromobenzyloxycarbonyl, 3,4-dimethoxybenzyloxycarbonyl, 3,5-dimethoxybenzyloxycarbonyl, and 2,4-dimethoxybenzyloxycarbonyl. hydroxycarbonyl, 4-methoxybenzyloxycarbonyl, 2-nitro-4,5-dimethoxybenzyloxycarbonyl, 3,4,5-trimethoxybenzyloxycarbonyl, 1-(p-biphenylyl)-1-methylethoxycarbonyl, α,α-dimethyl-3,5-dimethoxybenzyloxycarbonyl, benzhydryloxycarbonyl, t-butyloxycarbonyl, diisopropylmethoxycarbonyl, isopropyloxycarbonyl, ethoxycarbonyl, methyl Examples of N-protecting groups include carbamate-forming groups such as ethoxycarbonyl, allyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl, phenoxycarbonyl, 4-nitrophenoxycarbonyl, fluorenyl-9-methoxycarbonyl, cyclopentyloxycarbonyl, adamantyloxycarbonyl, cyclohexyloxycarbonyl, and phenylthiocarbonyl, alkaryl groups such as benzyl, triphenylmethyl, and benzyloxymethyl, and silyl groups such as trimethylsilyl. Preferred N-protecting groups are formyl, acetyl, benzoyl, pivaloyl, t-butylacetyl, alanyl, phenylsulfonyl, benzyl, t-butyloxycarbonyl (Boc), and benzyloxycarbonyl (Cbz).

[0083] As used herein, the term "O-protecting group" refers to a group intended to protect an oxygen-containing (e.g., phenol, hydroxyl, or carbonyl) group against undesired reactions during synthetic procedures. Commonly used O-protecting groups are disclosed in Greene, "Protective Groups in Organic Synthesis," 3rd Edition (John Wiley & Sons, New York, 1999), which is incorporated herein by reference. Exemplary O-protecting groups include acyl, aryloyl, or carbamyl groups such as formyl, acetyl, propionyl, pivaloyl, t-butylacetyl, 2-chloroacetyl, 2-bromoacetyl, trifluoroacetyl, trichloroacetyl, phthalyl, o-nitrophenoxyacetyl, α-chlorobutyryl, benzoyl, 4-chlorobenzoyl, 4-bromobenzoyl, t-butyldimethylsilyl, tri-isopropylsilyloxymethyl, 4,4′-dimethoxytrityl, isobutyryl, phenoxyacetyl, 4-isopropylphenoxyacetyl, dimethylformamidino, and 4-nitrobenzoyl; alkylcarbonyl groups such as acyl, acetyl, propionyl, and pivaloyl; and optionally substituted O-protecting groups such as benzoyl. arylcarbonyl groups; silyl groups such as trimethylsilyl (TMS), tert-butyldimethylsilyl (TBDMS), triisopropylsilyloxymethyl (TOM), and triisopropylsilyl (TIPS); groups that form ethers with hydroxyl groups such as methyl, methoxymethyl, tetrahydropyranyl, benzyl, p-methoxybenzyl, and trityl; alkoxycarbonyl groups such as methoxycarbonyl, ethoxycarbonyl, isopropoxycarbonyl, n-isopropoxycarbonyl, n-butyloxycarbonyl, isobutyloxycarbonyl, sec-butyloxycarbonyl, t-butyloxycarbonyl, 2-ethylhexyloxycarbonyl, cyclohexyloxycarbonyl, and methyloxycarbonyl;Alkoxyalkoxycarbonyl groups such as methoxymethoxycarbonyl, ethoxymethoxycarbonyl, 2-methoxyethoxycarbonyl, 2-ethoxyethoxycarbonyl, 2-butoxyethoxycarbonyl, 2-methoxyethoxymethoxycarbonyl, allyloxycarbonyl, propargyloxycarbonyl, 2-butenoxycarbonyl, and 3-methyl-2-butenoxycarbonyl; haloalkoxycarbonyl groups such as 2-chloroethoxycarbonyl, 2-chloroethoxycarbonyl, and 2,2,2-trichloroethoxycarbonyl; benzyl oxycarbonyl, p-methylbenzyloxycarbonyl, p-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl, 2,4-dinitrobenzyloxycarbonyl, 3,5-dimethylbenzyloxycarbonyl, p-chlorobenzyloxycarbonyl, p-bromobenzyloxycarbonyl, fluorenylmethyloxycarbonyl, and the like; and arylalkoxycarbonyl groups which may be substituted, such as phenoxycarbonyl, p-nitrophenoxycarbonyl, o-nitrophenoxycarbonyl, 2,4-dinitrophenoxycarbonyl, and the like. optionally substituted aryloxycarbonyl groups such as phenoxycarbonyl, p-methyl-phenoxycarbonyl, m-methylphenoxycarbonyl, o-bromophenoxycarbonyl, 3,5-dimethylphenoxycarbonyl, p-chlorophenoxycarbonyl, 2-chloro-4-nitrophenoxycarbonyl; substituted alkyl, aryl, and alkaryl ethers (e.g., trityl; methylthiomethyl; methoxymethyl; benzyloxymethyl; siloxymethyl; 2,2,2-trichloroethoxymethyl; tetrahydropyranyl; tetramethylphenoxycarbonyl); tetrahydrofuranyl; ethoxyethyl; 1-[2-(trimethylsilyl)ethoxy]ethyl; 2-trimethylsilylethyl; t-butyl ether; p-chlorophenyl, p-methoxyphenyl, p-nitrophenyl, benzyl, p-methoxybenzyl, and nitrobenzyl; silyl ethers (e.g., trimethylsilyl; triethylsilyl; triisopropylsilyl; dimethylisopropylsilyl; t-butyldimethylsilyl; t-butyldiphenylsilyl; tribenzylsilyl; triphenylsilyl; and diphenylmethylsilyl);Carbonate protecting groups (e.g., methyl, methoxymethyl, 9-fluorenylmethyl; ethyl; 2,2,2-trichloroethyl; 2-(trimethylsilyl)ethyl; vinyl, allyl, nitrophenyl; benzyl; methoxybenzyl; 3,4-dimethoxybenzyl; and nitrobenzyl); carbonyl protecting groups (e.g., acetal and ketal groups such as dimethyl acetal and 1,3-dioxolane; acylal groups; and dithiane groups such as 1,3-dithiane and 1,3-dithiolane); carboxylic acid protecting groups (e.g., ester groups such as methyl ester, benzyl ester, t-butyl ester, orthoester); and oxazoline groups.

[0084] The term "oxo" as used herein refers to =O.

[0085] As used herein, the term "polyethylene glycol" refers to an alkoxy chain comprising one or more monomeric units, each consisting of -OCHCH-. Polyethylene glycol (PEG) may also be referred to as polyethylene oxide (PEO) or polyoxyethylene (POE), and these terms may be considered interchangeable for purposes of the present invention. For example, polyethylene glycol has the structural formula -(CH) s2 (OCH2CH2) s1 (CH2) s3 O- (wherein s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), and s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10)). Polyethylene glycol may also have -NR N1 (CH2) s2 (CH2CH2O) s1 (CH2) s3 NR N1 (In the formula, s1 is an integer of 1 to 10 (e.g., 1 to 6 or 1 to 4), s2 and s3 are each independently an integer of 0 to 10 (e.g., 0 to 4, 0 to 6, 1 to 4, 1 to 6, or 1 to 10), and R N1 each independently represents hydrogen or optionally substituted C 1~6The amino-polyethylene glycols may also be considered to include amino-polyethylene glycols (wherein the alkyl is alkyl).

[0086] As used herein, the term "stereoisomer" refers to all possible different isomeric and conformational forms that a compound (e.g., a compound of any formula described herein) may possess, in particular all possible stereochemical and conformational isomeric forms of the basic molecular structure, all diastereomers, enantiomers and / or conformers. Some compounds of the present invention may exist in different tautomeric forms, all of the latter being included within the scope of the present invention.

[0087] The term "sulfonyl" as used herein refers to an -S(O2)- group.

[0088] The term "thiol" as used herein refers to an --SH group.

[0089] definition As used herein, the term "administered in combination" or "combined administration" means that two or more agents are administered to a subject simultaneously or within such an interval that there may be an overlap in the effects of each agent on the patient. In some embodiments, they are administered within 90 days (e.g., within 80, 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2, or 1 day), within 28 days (e.g., within 14, 7, 6, 5, 4, 3, 2, or 1 day), within 24 hours (e.g., within 12, 6, 5, 4, 3, 2, or 1 hour), or within about 60, 30, 15, 10, 5, or 1 minute of each other. In some embodiments, the administration of the agents is sufficiently close together so that a combined (e.g., synergistic) effect is achieved.

[0090] As used herein, "antibody" refers to a polypeptide whose amino acid sequence, including immunoglobulins and fragments thereof, specifically binds to a designated antigen, or fragment thereof. Antibodies according to the present invention may be of any type (e.g., IgA, IgD, IgE, IgG, or IgM) or subtype (e.g., IgA1, IgA2, IgG1, IgG2, IgG3, or IgG4). One skilled in the art will appreciate that a characteristic sequence or portion of an antibody may comprise amino acids found in one or more regions of an antibody (e.g., variable region, hypervariable region, constant region, heavy chain, light chain, and combinations thereof). Furthermore, one skilled in the art will appreciate that a characteristic sequence or portion of an antibody may comprise one or more polypeptide chains and may comprise sequence elements found in the same polypeptide chain or different polypeptide chains.

[0091] As used herein, "antigen-binding fragment" refers to a portion of an antibody that retains the binding properties of the parent antibody.

[0092] The terms "bifunctional chelate" or "bifunctional conjugate," as used interchangeably herein, refer to a compound containing a chelating group or a metal complex thereof, a linker group, and a therapeutic moiety, targeting moiety, or bridging group.

[0093] The term "cancer" refers to any cancer caused by the growth of malignant neoplastic cells, such as tumors, neoplasms, carcinomas, sarcomas, leukemias, and lymphomas. "Solid tumor cancers" are cancers that involve an abnormal mass of tissue, e.g., sarcomas, carcinomas, and lymphomas. "Hematologic cancers" or "liquid cancers," used interchangeably herein, are cancers that are present in bodily fluids, e.g., lymphomas and leukemias.

[0094] As used herein, the term "chelate" refers to an organic compound or portion thereof that can bind to a central metal or radioactive metal atom at two or more points.

[0095] As used herein, the term "conjugate" refers to a molecule containing a chelating group or a metal complex thereof, a linker group, and optionally a therapeutic moiety, a targeting moiety, or a bridging group.

[0096] As used herein, the term "compound" is meant to include all stereoisomers, geometric isomers, and tautomers of the structures depicted.

[0097] The compounds described herein may be asymmetric (e.g., having one or more stereocenters). Unless otherwise specified, all stereoisomers, such as enantiomers and diastereomers, are intended. Compounds of the present disclosure that contain asymmetrically substituted carbon atoms can be isolated in optically active or racemic forms. Methods for preparing optically active forms from optically active starting materials, such as by resolution of racemic mixtures or by stereoselective synthesis, are known in the art. Many geometric isomers of olefins, C=N double bonds, and the like, may also be present in the compounds described herein, and all such stable isomers are contemplated in this disclosure. Cis and trans geometric isomers of the compounds of the present disclosure are described and can be isolated as a mixture of isomers or as separated isomeric forms.

[0098] The compounds of the present disclosure also include tautomeric forms. Tautomeric forms result from the swapping of a single bond with an adjacent double bond and the concomitant migration of a proton. Tautomeric forms include proton tautomers, which are isomeric protonation states with the same empirical formula and total charge. Examples of proton tautomers include ketone-enol pairs, amide-imidic acid pairs, lactam-lactim pairs, amide-imidic acid pairs, enamine-imine pairs, and cyclic forms in which protons can occupy more than one position in a heterocyclic ring system, such as 1H- and 3H-imidazole, 1H-, 2H- and 4H-1,2,4-triazole, 1H- and 2H-isoindole, and 1H- and 2H-pyrazole. Tautomeric forms may be in equilibrium or sterically locked into one form by appropriate substitution.

[0099] At various places in the present specification, substituents of compounds of the present disclosure are disclosed in groups or in ranges. It is specifically intended that the present disclosure include each and every individual subcombination of the members of such groups and ranges. For example, the term "C 1~6 "Alkyl" is specifically intended to individually disclose methyl, ethyl, C alkyl, C alkyl, C alkyl, and C alkyl. The form of the phrase "optionally substituted X" (e.g., optionally substituted alkyl) herein is intended to be equivalent to "X, where X is optionally substituted" (e.g., "alkyl, where said alkyl is optionally substituted"). It is not intended to imply that the feature "X" (e.g., alkyl) itself is optional.

[0100] As used herein, a "detection agent" refers to a molecule or atom that is useful in diagnosing disease by locating cells containing an antigen. Various methods of labeling polypeptides with detection agents are known in the art. Examples of detection agents include, but are not limited to, radioisotopes and radionuclides, dyes (such as those containing biotin-streptavidin complexes), contrast agents, luminescent agents (e.g., FITC, rhodamine, lanthanide phosphors, cyanines, and near-IR dyes), and magnetic agents such as gadolinium chelates.

[0101] As used herein, the term "radionuclide" refers to an atom that can undergo radioactive decay (e.g., 3 H, 14 C. 15 N, 18 F, 35 S, 47 Sc, 55 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 75 Br, 76 Br, 77 Br, 89 Zr, 86 Y, 87 Y, 90 Y,97 Ru, 99 Tc, 99m Tc, 105 Rh, 109 Pd, 111 In, 123 I, 124 I, 125 I, 131 I, 149 Pm, 149 Tb, 153 Sm, 166 Ho, 177 Lu, 186 Re, 188 Re, 198 Au, 199 Au, 203 Pb, 211 At, 212 Pb, 212 Bi, 213 Bi, 223 Ra, 225 Ac, 227 Th, 229 Th, 66 Ga, 67 Ga, 68 Ga, 82 Rb, 117m Sn, 201 TI). The terms radioactive nuclide, radioisotope, or radioactive isotope can also be used to describe a radionuclide. Radionuclides can be used as detection agents, as described above. In some embodiments, the radionuclide can be an alpha-emitting radionuclide.

[0102] As used herein, the term "effective amount" of an agent (e.g., any of the conjugates described above) is an amount sufficient to effect beneficial or desired results, such as clinical results, and thus "effective amount" will depend on the context in which it is applied.

[0103] As used herein, the term "immunoconjugate" refers to a conjugate that includes a targeting moiety, such as an antibody, nanobody, affibody, or consensus sequence derived from a fibronectin type III domain. In some embodiments, the immunoconjugate includes an average of at least 0.10 conjugates per targeting moiety (e.g., an average of at least 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, or 5 conjugates per targeting moiety).

[0104] As used herein, the term "radioconjugate" refers to any conjugate that includes a radioisotope or radionuclide, such as any of the radioisotopes or radionuclides described herein. In some embodiments, the radioisotope or radionuclide is a metal chelate.

[0105] As used herein, the term "radioimmunoconjugate" refers to any immunoconjugate that includes a radioisotope or radionuclide, such as any of the radioisotopes or radionuclides described herein. In some embodiments, the radioisotope or radionuclide is a metal chelate.

[0106] As used herein, the term "radioimmunotherapy" refers to a method of using a radioimmunoconjugate to produce a therapeutic effect. In some embodiments, radioimmunotherapy may involve administration of a radioimmunoconjugate to a subject in need thereof, where administration of the radioimmunoconjugate produces a therapeutic effect in the subject. In some embodiments, radioimmunotherapy may involve administration of a radioimmunoconjugate to cells, where administration of the radioimmunoconjugate kills the cells. When radioimmunotherapy involves selective killing of cells, in some embodiments, the cells are cancer cells in a subject with cancer.

[0107] As used herein, the term "pharmaceutical composition" refers to a composition containing a compound described herein formulated with a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition is manufactured or sold under the approval of a government regulatory agency as part of a therapeutic regimen for treating a disease in a mammal. The pharmaceutical composition can be formulated, for example, for oral administration in a unit dosage form (e.g., a tablet, capsule, caplet, gel cap, or syrup); for topical administration (e.g., as a cream, gel, lotion, or ointment); for intravenous administration (e.g., as a sterile solution free of particulate matter and in a solvent system suitable for intravenous use); or for any other formulation described herein.

[0108] As used herein, "pharmaceutically acceptable excipient" refers to any ingredient other than the compounds described herein (e.g., a vehicle capable of suspending or dissolving an active compound) that has non-toxic and non-inflammatory properties in patients. Excipients may include, for example, antiadhesives, antioxidants, binders, coatings, compression aids, disintegrants, dyes (colorants), emollients, emulsifiers, fillers (diluents), film formers or coatings, flavors, fragrances, glidants (flow enhancers), lubricants, preservatives, printing inks, radioprotectants, adsorbents, suspending or dispersing agents, sweeteners, or water for hydration. Exemplary excipients include, but are not limited to, ascorbic acid, histidine, phosphate buffer, butylated hydroxytoluene (BHT), calcium carbonate, calcium phosphate (dibasic), calcium stearate, croscarmellose, cross-linked polyvinylpyrrolidone, citric acid, crospovidone, cysteine, ethylcellulose, gelatin, hydroxypropyl cellulose, hydroxypropylmethylcellulose, lactose, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, methylparaben, microcrystalline cellulose, polyethylene glycol, polyvinylpyrrolidone, povidone, pregelatinized starch, propylparaben, retinol palmitate, shellac, silicon dioxide, sodium carboxymethylcellulose, sodium citrate, sodium starch glycolate, sorbitol, starch (corn), stearic acid, stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C, and xylitol.

[0109] As used herein, the term "pharmaceutically acceptable salt" refers to a salt of a compound described herein that is within the scope of sound medical judgment and suitable for use in contact with the tissues of humans and animals without undue toxicity, irritation, or allergic response. Pharmaceutically acceptable salts are well known in the art. For example, pharmaceutically acceptable salts are described in Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977 and Pharmaceutical Salts: Properties, Selection, and Use (P.H. Stahl and C.G. Wermuth (eds.), Wiley-VCH, 2008). Salts can be prepared in situ during the final isolation and purification of a compound described herein, or separately by reacting the free base with a suitable organic acid.

[0110] The compounds of the present invention may have ionizable groups so that they can be prepared as pharmaceutically acceptable salts. These salts may be acid addition salts with inorganic or organic acids, or salts can be prepared from inorganic or organic bases in the case of the acidic form of the compounds of the present invention. Frequently, compounds are prepared or used as pharmaceutically acceptable salts prepared as addition products of pharmaceutically acceptable acids or bases. Suitable pharmaceutically acceptable acids and bases, such as hydrochloric acid, sulfuric acid, hydrobromic acid, acetic acid, lactic acid, citric acid, or tartaric acid for forming acid addition salts, and potassium hydroxide, sodium hydroxide, ammonium hydroxide, caffeine, various amines, etc. for forming base salts, are known in the art. Methods for preparing suitable salts are well established in the art.

[0111] Representative acid addition salts include, inter alia, acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxy-ethanesulfonate, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, and magnesium salts, as well as non-toxic ammonium, quaternary ammonium, and amine cations, such as, but not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, and ethylamine.

[0112] As used herein, the term "therapeutic moiety" refers to any molecule or any portion of a molecule that provides a therapeutic benefit. In some embodiments, a therapeutic moiety is a protein or polypeptide, e.g., an antibody, an antigen-binding fragment thereof. In some embodiments, a therapeutic moiety is a small molecule.

[0113] As used herein, the term "targeting moiety" refers to any molecule or any portion of a molecule that binds to a given target. In some embodiments, a targeting moiety is a protein or polypeptide, such as an antibody or antigen-binding fragment thereof, a nanobody, an affibody, or a consensus sequence derived from a fibronectin type III domain.

[0114] As used herein, the term "crosslinking group" refers to any reactive group capable of covalently linking two or more molecules. In some embodiments, the crosslinking group is an amino-reactive or thiol-reactive crosslinking group. In some embodiments, the amino-reactive or thiol-reactive crosslinking group includes activated esters or imidates, such as hydroxysuccinimide esters, 2,3,5,6-tetrafluorophenol esters, 4-nitrophenol esters, anhydrides, thiols, disulfides, maleimides, azides, alkynes, strained alkynes, strained alkenes, halogens, sulfonates, haloacetyls, amines, hydrazides, diazirines, phosphines, tetrazines, and isothiocyanates. In some embodiments, the crosslinking group may be glycine-glycine-glycine and / or leucine-proline-(any amino acid)-threonine-glycine, which are recognition sequences for coupling a targeting agent to a linker using a sortase-mediated coupling reaction. One of skill in the art will understand that the use of crosslinking groups in the practice of the present invention is not limited to the specific constructs disclosed herein, but rather may include other known crosslinking groups.

[0115] As used herein, the term "polypeptide" refers to a string of at least two amino acids linked to each other by peptide bonds. In some embodiments, a polypeptide may contain at least 3-5 amino acids, each linked to another amino acid by at least one peptide bond. One of skill in the art will appreciate that a polypeptide may contain one or more "unnatural" amino acids or other entities that can nevertheless be incorporated into a polypeptide chain. In some embodiments, a polypeptide may be glycosylated, e.g., a polypeptide may contain one or more covalently linked sugar moieties. In some embodiments, a single "polypeptide" (e.g., an antibody polypeptide) may comprise two or more individual polypeptide chains, which in some cases may be linked to each other, e.g., by one or more disulfide bonds or other means.

[0116] By "subject" is meant a human or non-human animal (eg, a mammal).

[0117] "Substantial identity" or "substantially identical" refers to a polypeptide sequence that has the same polypeptide sequence as a reference sequence, or a specified percentage of amino acid residues that are the same at corresponding positions in the reference sequence when the two sequences are optimally aligned. For example, an amino acid sequence that is "substantially identical" to a reference sequence has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to the reference amino acid sequence. For polypeptides, the length of the comparison sequence is generally at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 50, 75, 90, 100, 150, 200, 250, 300, or 350 contiguous amino acids (e.g., full-length sequence). Sequence identity can be measured using sequence analysis software with default settings (e.g., Sequence Analysis Software Package of the Genetics Computer Group, University of Wisconsin Biotechnology Center, 1710 University Avenue, Madison, WI 53705). Such software can match similar sequences by assigning degrees of homology to various substitutions, deletions, and other modifications.

[0118] As used herein, and as well understood in the art, "treating" a condition or "treatment" of a condition (e.g., a condition described herein, such as cancer) is an approach for obtaining a beneficial or desired result, such as a clinical result. Beneficial or desired results include, but are not limited to, alleviation or amelioration of one or more symptoms or conditions, whether detectable or undetectable; reduction in the extent of the disease, disorder, or condition; stabilization (i.e., not worsening) of the disease, disorder, or condition; prevention of the spread of the disease, disorder, or condition; delay or slowing of the progression of the disease, disorder, or condition; improvement or palliation of the disease, disorder, or condition; and remission (partial or complete remission). "Amelioration" of a disease, disorder, or condition means a decrease in the extent and / or undesirable clinical findings of the disease, disorder, or condition and / or a slowing or prolongation of the time course of progression compared to the extent or time course in the absence of treatment. [Brief explanation of the drawings]

[0119] [Figure 1] FIG. 1 depicts the general structure of a conjugate comprising a chelate, a linker, and a bridging group (top) and a conjugate comprising a chelate, a linker, and a targeting moiety (bottom). [Figure 2] 1 illustrates the synthesis of the bifunctional chelate 4-{[11-oxo-11-(2,3,5,6-tetrafluorophenoxy)undecyl]carbamoyl}-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl]butanoic acid (Compound B). The synthesis of Compound B is described in Example 3. [Figure 3] FIG. 1 describes the synthesis of the bifunctional chelate, 4-{[2-(2-{2-[3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propoxy]ethoxy}ethoxy)ethyl]carbamoyl}-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl]butanoic acid (Compound C). The synthesis of Compound C is described in Example 4. [Figure 4]1 is a series of graphs depicting the metabolic excretion profiles of non-targeting human IgG antibody conjugates [177Lu]-Compound B-HuMIgG and [177Lu]-Compound C-HuMIgG compared to [177Lu]-Compound A-HuMIgG. The methods and results are detailed in Example 7. DETAILED DESCRIPTION OF THE INVENTION

[0120] Radiolabeled targeting moieties (also known as radioimmunoconjugates) are designed to target proteins or receptors that are upregulated in disease states and deliver a radioactive payload to kill the target cells (radioimmunotherapy). Methods that deliver such payloads by radioactive decay result in the emission of alpha, beta, or gamma particles or Auger electrons that can cause direct effects on DNA (such as single- or double-stranded DNA breaks) or indirect effects such as bystander or crossfire effects.

[0121] A radioimmunoconjugate typically contains a biological targeting moiety, a radioisotope, and a conjugate linking the two. The conjugate is formed when a bifunctional chelate is added to the biological targeting molecule in a manner that minimizes structural changes to the compound while maintaining target affinity. Once radiolabeled, the final radioimmunoconjugate is formed.

[0122] Structurally, bifunctional chelates contain a chelate, a linker, and a bridging group (Figure 1). When developing new bifunctional chelates, most efforts focus on the chelating portion of the molecule. Several examples of bifunctional chelates with various cyclic and acyclic structures when conjugated to targeting moieties have been described (Bioconjugate Chem. 2000, 11, pp. 510-519; Bioconjugate Chem. 2012, 23, pp. 1029-1039; Mol Imaging Biol (2011) 13:215-221; Bioconjugate Chem. 2002, 13, pp. 110-115).

[0123] One key factor in developing safe and effective radioimmunoconjugates is maximizing efficacy while minimizing off-target toxicity in normal tissues. While this is one of the core tenets of developing new drugs, their application to radioimmunotherapy presents new challenges. To be therapeutically effective, radioimmunoconjugates do not need to block receptors, as is required for therapeutic antibodies, nor do they need to release a cytotoxic payload intracellularly, as is required for antibody-drug conjugates. However, the release of toxic particles occurs as a result of primary (radioactive) decay and is a random event that can occur anywhere in the body after administration. Once released, damage to surrounding cells within the radiation range occurs, leading to the potential for off-target toxicity. Therefore, limiting exposure of these releases to normal tissues is key to developing new drugs (Bioconjugate Chem. 2006, 17, pp. 1551-1560; Bioconjugate Chem. 2003, 14, pp. 927-933).

[0124] One potential way to reduce off-target exposure is to more efficiently remove radioactivity from the body. The most obvious mechanism is to increase the clearance rate of biological targeting agents. This approach also requires identifying a method to shorten the half-life of biological targeting agents, which is a topic that has not been described much about biological targeting agents. Regardless of the mechanism, increasing drug clearance also negatively affects pharmacodynamics / efficacy, in that faster drug removal from the body reduces the effective concentration at the site of action, which then requires a higher total dose, and does not achieve the desired result of reducing the total radioactive dose to normal tissues.

[0125] Other efforts have focused on accelerating the metabolism of the portion of the molecule containing the radioactive moiety. To this end, some efforts have been made to increase the rate of cleavage of radioactivity from biological targeting agents using what have been called "cleavable linkers." However, the term "cleavable linker" has a different meaning as it relates to radioimmunoconjugates. Cornelissen et al. described a cleavable linker as a bifunctional conjugate attached to a biological targeting agent via a reduced cysteine, while others have described the use of an enzymatic cleavage system that requires simultaneous administration of a radioimmunoconjugate and a cleaving agent / enzyme for release [Mol Cancer Ther; 12(11) November 2013, Methods in Molecular Biology, 2009, 539, 191-211, Bioconjugate Chemistry, Volume 14, Issue 5, 927-33 (2003)]. These methods are not practical from a drug development perspective (enzymatic cleavage systems) because they alter the nature of the biological targeting moiety, in the case of cysteine ​​bonds, or, in the case of the provided citation, they require the administration of two drugs.

[0126] The focus of the embodiments described herein centers on more efficient removal of radioactivity from the body following catabolism and / or metabolism of the radioimmunoconjugate by modifying the linker region of the bifunctional chelate.

[0127] This is a new approach, especially since it applies to radioimmunoconjugates, since there is little information describing the in vivo effects of linkers. One possible reason is that after the catabolism / metabolism of the radioimmunoconjugate, those skilled in the art would expect the radiolabeled conjugate to undergo rapid systemic elimination. When a bifunctional chelate was administered alone, the speculation was advanced experimentally; it cleared the bloodstream faster than a radioimmunoconjugate with the same bifunctional chelate. Based on these data, those skilled in the art would expect that after the catabolism / metabolism of the radioimmunoconjugate, metabolites containing the bifunctional chelate would also be rapidly eliminated.

[0128] However, rapid clearance of metabolites containing radiolabeled conjugates does not necessarily occur in vivo. Based on the results described below, the linker region of the bifunctional chelate does not affect the overall in vitro properties or in vivo pharmacokinetics and pharmacodynamics of the radioimmunoconjugate, but can directly affect the removal of radioactivity from the body after catabolism of the radioconjugate. Data are presented below demonstrating that certain commercially available bifunctional chelates result in a slower rate and a lower degree of removal of total radioactivity from the body when compared to the embodiments described herein.

[0129] The excretion profiles of the embodiments described in the Examples demonstrate an unexpected finding. As previously reported by Quadri and Vriesendorp [QJ Nucl. Med. 1998, 42, 250-261], simple modifications to the linker region of bifunctional chelates did not affect the urinary excretion of radioactivity, despite their hydrophobicity. The results provided below clearly demonstrate that both hydrophobic and hydrophilic linkers can affect excretion patterns. Furthermore, the following Examples demonstrate that hepatic biliary clearance also plays a role in excretion.

[0130] Thus, embodiments described herein have identified bifunctional chelates that, when conjugated to biological targeting moieties, achieve reduced systemic radioactivity by increasing the extent of catabolic / metabolic excretion while maintaining the pharmacokinetics of the intact molecule compared to known similar bifunctional chelates. This reduction in systemic radioactivity has been determined to be due to clearance of catabolic / metabolic by-products, but does not affect other in vitro and in vivo properties, such as specificity (in vitro binding), cellular retention, and in vivo tumor uptake. Taken as a whole, these embodiments achieve the desired properties of radioimmunoconjugates by reducing the body burden of radioactivity while maintaining on-target activity.

[0131] Therapeutic and targeting moieties A therapeutic moiety includes any molecule or any portion of a molecule that provides a therapeutic benefit. In some embodiments, a therapeutic moiety is a protein or polypeptide, e.g., an antibody or antigen-binding fragment thereof. In some embodiments, a therapeutic moiety is a small molecule. A targeting moiety includes any molecule or any portion of a molecule that binds to a given target. In some embodiments, a targeting moiety is a protein or polypeptide such as an antibody or antigen-binding fragment thereof, a nanobody, an affibody, and a consensus sequence derived from a type III fibronectin domain (e.g., centyrin or adnectin).

[0132] Polypeptides Polypeptides include, for example, any of a variety of hematological agents (including, for example, erythropoietin, blood clotting factors, etc.), interferons, colony stimulating factors, antibodies, enzymes, and hormones. The identity of the particular polypeptide is not intended to limit the disclosure, and any polypeptide of interest may be a polypeptide in the methods of the present invention.

[0133] The reference polypeptides described herein may comprise a target-binding domain that binds to a target of interest (e.g., binds to an antigen). For example, a polypeptide such as an antibody may bind to a transmembrane polypeptide (e.g., a receptor) or a ligand (e.g., a growth factor). Exemplary molecular targets (e.g., antigens) for the polypeptides (e.g., antibodies) described herein include CD proteins such as CD2, CD3, CD4, CD8, CD11, CD19, CD20, CD22, CD25, CD33, CD34, CD40, and CD52; members of the ErbB receptors, such as EGF receptor (EGFR, HER1, ErbB1), HER2 (ErbB2), HER3 (ErbB3), or HER4 (ErbB4) receptors; macrophage receptors, such as CRIg; tumor necrosis factors, such as TNFα or TRAIL / Apo-2; cell adhesion molecules, such as LFA-1, Mac1, p150,95, VLA-4, ICAM-1, VCAM, and αvβ3 integrin containing either the α or β subunit (e.g., anti-CD11a, anti-CD18, or anti-CD11b antibodies); EGF, EGFR (e.g., FGFR3), and These include growth factors and receptors such as VEGF, IgE, cytokines such as IL1, cytokine receptors such as IL2 receptor, blood group antigens, flk2 / flt3 receptor, obesity (OB) receptor, mpl receptor, CTLA-4, protein C, neuropilin, ephrins and receptors, netrins and receptors, slit and receptors, chemokines and chemokine receptors such as CCL5, CCR4, and CCR5, amyloid beta, complement factors such as complement factor D, lipoproteins such as oxidized LDL (oxLDL), and lymphotoxins such as lymphotoxin alpha (LTa). Other molecular targets include Tweak, B7RP-1, proprotein convertase subtilisin / kexin type 9 (PCSK9), sclerostin, c-kit, Tie-2, c-fms, and anti-M1.

[0134] antibody IgG antibodies consist of two identical light chain polypeptides and two identical heavy chain polypeptides linked together by disulfide bonds. The first domain, located at the amino terminus of each chain, is variable in amino acid sequence and provides the antibody binding specificity found in each individual antibody. These are known as the variable heavy (VH) and variable light (VL) regions. The other domains of each chain are relatively invariant in amino acid sequence and are known as the constant heavy (CH) and constant light (CL) regions. For IgG antibodies, the light chain contains one variable region (VL) and one constant region (CL). IgG heavy chains contain a variable region (VH), a first constant region (CH1), a hinge region, a second constant region (CH2), and a third constant region (CH3). In IgE and IgM antibodies, the heavy chain contains an additional constant region (CH4).

[0135] Antibodies described herein may include, for example, monoclonal antibodies, polyclonal antibodies, multispecific antibodies, human antibodies, humanized antibodies, camelid antibodies, chimeric antibodies, single-chain Fvs (scFvs), disulfide-linked Fvs (sdFvs), and anti-idiotypic (anti-Id) antibodies, as well as antigen-binding fragments of any of the above. Antibodies may be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass.

[0136] As used herein, the term "antigen-binding fragment" of an antibody refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen. Examples of binding fragments encompassed by the term "antigen-binding fragment" of an antibody include Fab fragments, F(ab')2 fragments, Fd fragments, Fv fragments, scFv fragments, dAb fragments (Ward et al. (1989) Nature 341:544-546), and isolated complementarity-determining regions (CDRs). These antibody fragments can be obtained using conventional techniques known to those of skill in the art, and the fragments can be screened for utility in the same manner as intact antibodies.

[0137] The antibodies or fragments described herein can be produced by any method known in the art for the synthesis of antibodies (e.g., Harlow et al., Antibodies: A Laboratory Manual (Cold Spring Harbor Laboratory Press, 2nd ed., 1988); Brinkman et al., 1995, J. Immunol. Methods 182:41-50; WO92 / 22324; WO98 / 46645). Chimeric antibodies can be produced, for example, using the method described in Morrison, 1985, Science 229:1202, and humanized antibodies can be produced by the method described in U.S. Pat. No. 6,180,370.

[0138] Further antibodies described herein are bispecific and multispecific antibodies, e.g., as described in Segal et al., J. Immunol. Methods 248:1-6 (2001); and Tutt et al., J. Immunol. 147:60 (1991).

[0139] Insulin-like growth factor 1 (IGF-1R) antibodies The insulin-like growth factor 1 receptor is a transmembrane protein found on the surface of human cells that is activated by insulin-like growth factors 1 (IGF-1) and 2 (IGF-2). IGF-1R is involved in several cancers, including breast cancer, non-small cell lung cancer, prostate cancer, colon cancer, sarcoma, and adrenocortical carcinoma, and high levels of IGF-1R are expressed on the surface of tumor cells in these cancers.

[0140] In some embodiments, the antibody, or antigen-binding fragment thereof, specifically binds to insulin-like growth factor-1 receptor (IGF-1R).

[0141] Nanobodies Nanobodies are antibody fragments consisting of a single monomeric variable antibody domain. Nanobodies may also be referred to as single-domain antibodies. Like antibodies, nanobodies selectively bind to a specific antigen. Nanobodies may be heavy chain variable domains or light chain domains. Nanobodies may be naturally occurring or the product of biological engineering. Nanobodies can be biologically engineered by site-directed mutagenesis or mutagenic screening (e.g., phage display, yeast display, bacterial display, mRNA display, ribosome display).

[0142] Affibody Affibodies are polypeptides or proteins engineered to bind to specific antigens. As such, affibodies may be thought of as mimicking certain functions of antibodies. Affibodies may be engineered variants of the B domain in the immunoglobulin-binding region of Staphylococcus aureus protein A. Affibodies may also be engineered variants of the Z domain, a B domain with lower affinity for the Fab region. Affibodies can be biologically engineered by site-directed mutagenesis or mutagenic screening (e.g., phage display, yeast display, bacterial display, mRNA display, ribosome display).

[0143] Affibody molecules have been generated that exhibit specific binding to a variety of different proteins (e.g., insulin, fibrinogen, transferrin, tumor necrosis factor-α, IL-8, gp120, CD28, human serum albumin, IgA, IgE, IgM, HER2, and EGFR) with affinities (K ) in the μM to pM range. d ) is shown.

[0144] Fibronectin type III domain The type III fibronectin domain is an evolutionarily conserved protein domain found in various extracellular proteins. Type III fibronectin domains have been used as molecular scaffolds to produce molecules that can selectively bind to specific antigens. Variants of type III fibronectin domains (FN3) engineered for selective binding can also be called monobodies. The FN3 domain can be biologically engineered by site-directed mutagenesis or mutagenic screening (e.g., CIS display, phage display, yeast display, bacterial display, mRNA display, ribosome display).

[0145] Modified Polypeptides Polypeptides of the present invention may have a modified amino acid sequence. A modified polypeptide may be substantially identical to a corresponding reference polypeptide (e.g., the amino acid sequence of a modified polypeptide may have at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of the reference polypeptide). In certain embodiments, the modification does not significantly destroy a desired biological activity. The modification may reduce (e.g., at least 5%, 10%, 20%, 25%, 35%, 50%, 60%, 70%, 75%, 80%, 90%, or 95%) the biological activity of the original polypeptide, have no effect thereon, or may increase (e.g., at least 5%, 10%, 25%, 50%, 100%, 200%, 500%, or 1000%) it. The modified polypeptides may have or optimize polypeptide characteristics such as in vivo stability, bioavailability, toxicity, immunological activity, immunological identity, and conjugation properties.

[0146] Modifications include those that result from natural processes, such as post-translational processing, or from chemical modification techniques known in the art. Modifications can occur anywhere in a polypeptide, including the polypeptide backbone, the amino acid side-chains, and the amino or carboxyl termini. The same type of modification can be present in the same or varying degrees at several sites in a given polypeptide, and a polypeptide can contain more than one type of modification. Polypeptides can be branched as a result of ubiquitination, and they can be cyclic, with or without branching. Cyclic, branched, and branched cyclic polypeptides can result from post-translational natural processes or can be produced synthetically. Other modifications include pegylation, acetylation, acylation, addition of an acetomidomethyl (Acm) group, ADP-ribosylation, alkylation, amidation, biotinylation, carbamoylation, carboxyethylation, esterification, covalent attachment to a flavin, covalent attachment to a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a drug, covalent attachment of a marker (e.g., a fluorescent or radioactive marker), covalent attachment of a lipid or lipid derivative, covalent attachment of phosphatidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cysteine, formation of pyroglutamate, formylation, gamma-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA-mediated addition of amino acids to proteins, such as arginylation and ubiquitination.

[0147] Modified polypeptides may also include amino acid insertions, deletions, or substitutions, either conservative or non-conservative (e.g., D-amino acids, desamino acids), in the polypeptide sequence (e.g., where such changes do not substantially alter the biological activity of the polypeptide). In particular, the addition of one or more cysteine ​​residues to the amino or carboxy terminus of any of the polypeptides of the invention can facilitate conjugation of these polypeptides, e.g., by disulfide bonds. For example, polypeptides can be modified to include a single cysteine ​​residue at the amino terminus or a single cysteine ​​residue at the carboxy terminus. Amino acid substitutions can be conservative (i.e., where a residue is replaced by another of the same general type or group) or non-conservative (i.e., where a residue is replaced by an amino acid of a different type). Furthermore, naturally occurring amino acids can be substituted with non-naturally occurring amino acids (i.e., non-naturally occurring conservative amino acid substitutions or non-naturally occurring non-conservative amino acid substitutions).

[0148] Synthetically produced polypeptides may include substitutions of amino acids that are not naturally encoded by DNA (e.g., non-naturally occurring or unnatural amino acids). Examples of non-naturally occurring amino acids include D-amino acids, N-protected amino acids, amino acids with an acetylaminomethyl group attached to the sulfur atom of cysteine, pegylated amino acids, amino acids of the formula NH2(CH2), n These include omega-amino acids with COOH (where n is 2-6), neutral nonpolar amino acids such as sarcosine, t-butylalanine, t-butylglycine, N-methylisoleucine, and norleucine. Phenylglycine may be substituted for Trp, Tyr, or Phe; citrulline and methionine sulfoxide are neutral nonpolar, cysteic acid is acidic, and ornithine is basic. Proline may be substituted with hydroxyproline, which retains the conformation that provides the properties.

[0149] Analogs can be generated by substitution mutagenesis, which retain the biological activity of the original polypeptide. Examples of substitutions identified as "conservative substitutions" are shown in Table 1. If such substitutions result in undesirable changes, other types of substitutions designated as "exemplary substitutions" in Table 1, or further described herein with reference to amino acid classes, are introduced and the products screened.

[0150] [Table 1]

[0151] Substantial alteration of function or immunological identity is achieved by selecting substitutions that differ significantly in their effect on (a) the structure of the polypeptide backbone in the region of the substitution, e.g., as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) maintaining side chain bulk.

[0152] crosslinking group A crosslinking group is a reactive group capable of covalently linking two or more molecules. Crosslinking groups can be used to attach linkers and chelating moieties to therapeutic or targeting moieties. Crosslinking groups can also be used to attach linkers and chelating moieties to targets in vivo. In some embodiments, the crosslinking group is an amino-reactive or thiol-reactive crosslinking group, or a sortase-mediated coupling. In some embodiments, the amino-reactive, methionine-reactive, or thiol-reactive crosslinking group comprises an activated ester or imidate, such as a hydroxysuccinimide ester, a 2,3,5,6-tetrafluorophenol ester, a 4-nitrophenol ester, an anhydride, a thiol, a disulfide, a maleimide, an azide, an alkyne, a strained alkyne, a strained alkene, a halogen, a sulfonate, a haloacetyl, an amine, a hydrazide, a diazirine, a phosphine, a tetrazine, an isothiocyanate, or an oxaziridine. In some embodiments, the sortase recognition sequence may comprise a terminal glycine-glycine-glycine (GGG) and / or LPTXG amino acid sequence, where X is any amino acid. One of skill in the art will appreciate that the use of cross-linking groups in the practice of the present invention is not limited to the particular constructs disclosed herein, but rather may include other known cross-linking groups.

[0153] detection agent A detection agent is a molecule or atom that is conjugated to a polypeptide, such as an antibody or an antigen-binding fragment thereof, and administered to the polypeptide to be useful in diagnosing, planning radiation treatment, or treating disease by locating cells containing the antigen. Useful detection agents include, but are not limited to, radioisotopes, dyes (such as those containing biotin-streptavidin complexes), contrast agents, fluorescent compounds or molecules, luminescent agents, and enhancing agents for magnetic resonance imaging (MRI) (e.g., paramagnetic ions). To load a detection agent onto a polypeptide component, it may be necessary to react it with a reagent having a linker that binds to the detection agent or agents.

[0154] Radioisotopes and Radionuclides Radioisotopes and radionuclides known in the art for their use as detection agents include, but are not limited to: 3 H, 14 C. 15 N, 18 F, 35 S, 47 Sc, 55 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 75 Br, 76 Br, 77 Br, 89 Zr, 86 Y, 87 Y, 90 Y, 97 Ru, 99 Tc, 99m Tc, 105 Rh, 109 Pd, 111 In, 123 I, 124 I, 125 I, 131 I, 149 Pm, 149 Tb, 153 Sm, 166 Ho, 177 Lu, 186 Re, 188 Re, 198 Au, 199 Au, 203 Pb, 211 At, 212 Pb, 212 Bi, 213 Bi, 223 Ra, 225 Ac, 227 Th, 229 Th, 66 Ga, 67 Ga, 68 Ga, 82 Rb, 117m Sn, 201 Examples include Tl.

[0155] Chelating Moiety Chelating moieties known in the art for their use as detection agents include, but are not limited to, DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), DOTMA ((1R,4R,7R,10R)-α,α',α'',α'''-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), DOTAM (1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane), DOTPA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrapropionic acid), DOAM-acetic acid (2-(4,7,10-tris(2-amino-2-oxoethyl)-1, ... 10-tetraazacyclododecan-1-yl)acetic acid), DOTA-GA anhydride (2,2',2''-(10-(2,6-dioxotetrahydro-2H-pyran-3-yl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid, DOTP (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylenephosphonic acid)), DOTMP (1,4,6,10-tetraazacyclodecane-1,4,7,10-tetramethylenephosphonic acid), DOTA-4AMP (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(acetamido-methylenephosphonic acid), CB-TE2A (1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-diacetic acid), NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid), NOTP (1,4,7-triazacyclononane-1,4,7-tri(methylenephosphonic acid), TETPA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetrapropionic acid), TETA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11- tetraacetic acid), HEHA (1,4,7,10,13,16-hexaazacyclohexadecane-1,4,7,10,13,16-hexaacetic acid), PEPA (1,4,7,10,13-pentaazacyclopentadecane-N,N',N'',N''',N''''-pentaacetic acid), H4Octapa (N,N'-bis(6-carboxy-2-pyridylmethyl)-ethylenediamine-N,N'-diacetic acid), H2Dedpa (1,2 -[[6-(carboxy)-pyridin-2-yl]-methylamino]ethane), H6phospa (N,N'-(methylenephosphonate)-N,N'-[6-(methoxycarbonyl)pyridin-2-yl]-methyl-1,2-diaminoethane), TTHA (triethylenetetramine-N,N,N',N'',N''',N'''-hexaacetic acid), DO2P (tetraazacyclododecanedimethanephosphonic acid), HP-DO3A (hydroxypropyltetraazacyclododecanetriacetic acid), EDTA (ethylenediaminetetraacetic acid), deferoxamine, DTPA (diethylenetriaminepentaacetic acid), DTPA-BMA (diethylenetriaminepentaacetic acid-bismethylamide), HOPO (octadentate hydroxypyridinone), or porphyrin. Chelating groups can be used to chelating metals such as manganese, iron, and gadolinium, as well as to chelating metals such as manganese, iron, and gadolinium. 47 Sc, 55 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 66 Ga, 67 Ga, 68 Ga, 82 Rb, 86 Y, 87 Y, 90 Y, 97 Ru, 99m Tc, 105 Rh,109 Pd, 111 In, 117m Sn, 149 Tb, 149 Pm, 153 Sm, 177 Lu, 186 Re, 188 Re, 199 Au, 201 Tl, 203 Pb, 212 Pb, 212 Bi, 213 Bi, 225 Ac, and 227 It can be used in metal chelate combinations with isotopes such as Th (eg, isotopes in the general energy range of 60 to 4,000 keV).

[0156] Linker The linkers of the present invention have Formula I: AL 1 -(L 2 ) n -B Formula I wherein A is a chelating moiety or a metal complex thereof; L 1 is an optionally substituted C1-C6 alkyl, an optionally substituted C1-C6 heteroalkyl, an optionally substituted aryl or heteroaryl; B is a therapeutic moiety, targeting moiety, or bridging group, or a pharmaceutically acceptable salt thereof; n is 1 to 5; L 2 each independently represents the structure: (-X 1 -L 3 -Z 1 -) Formula II (In the formula, X 1 is C=O(NR 1 ), C=S(NR 1 ), O-C=O(NR 1 ), NR 1 C=O(O), NR 1 C=O(NR 1 ), -CH2PhC=O(NR 1), -CH2Ph(NH)C=S(NR 1 ), O, NR 1 and R 1 is H or optionally substituted C1-C6 alkyl or optionally substituted C1-C6 heteroalkyl, optionally substituted aryl or heteroaryl; L 3 is optionally substituted C1 to C 50 Alkyl or optionally substituted C1-C 50 Heteroalkyl or C5-C 20 Polyethylene glycol; Z 1 are CH2, C=O, C=S, OC=O, NR 1 C=O, NR 1 and R 1 is hydrogen, optionally substituted C1-C6 alkyl, pyrrolidine-2,5-dione) have] The structure may be:

[0157] The conjugate of the present invention comprises three different modules which together provide it with increased efficacy compared to those known in the art.

[0158] 1. Chelating moiety or its metal complex: Module A is included for the incorporation of a detection agent (e.g., a chelating moiety or a metal complex thereof). The metal complex may comprise an imaging radionuclide.

[0159] 2. Linker: The linkers of the present invention have Formula I: AL 1 -(L 2 ) n -B Formula I wherein A is a chelating moiety or a metal complex thereof; L 1 is an optionally substituted C1-C6 alkyl, an optionally substituted C1-C6 heteroalkyl, an optionally substituted aryl or heteroaryl; B is a therapeutic moiety, targeting moiety, or bridging group, or a pharmaceutically acceptable salt thereof; n is 1 to 5; L 2 each independently represents the structure: (-X 1 -L 3 -Z 1 -) Formula II (In the formula, X 1 is C=O(NR 1 ), C=S(NR 1 ), O-C=O(NR 1 ), NR 1 C=O(O), NR 1 C=O(NR 1 ), -CH2PhC=O(NR 1 ), -CH2Ph(NH)C=S(NR 1 ), O, NR 1 and R 1 is H or optionally substituted C1-C6 alkyl or optionally substituted C1-C6 heteroalkyl, optionally substituted aryl or heteroaryl; L 3 is optionally substituted C1 to C 50 Alkyl or optionally substituted C1-C 50 Heteroalkyl or C5-C 20 Polyethylene glycol; Z 1 are CH2, C=O, C=S, OC=O, NR 1 C=O, NR 1 and R 1 is hydrogen, optionally substituted C1-C6 alkyl, pyrrolidine-2,5-dione) have] The structure may be:

[0160] 3. Therapeutic moieties, targeting moieties, or cross-linking groups: Module B is a therapeutic moiety (e.g., antibody, antigen-binding fragment), a targeting moiety (e.g., nanobody, affibody, consensus sequence derived from fibronectin type III domain), or a cross-linking group (e.g., amino-reactive, thiol-reactive cross-linking group, or sortase-mediated coupling).

[0161] Administration and Dosage The present invention also features pharmaceutical compositions containing a therapeutically effective amount of a compound of the present invention. The compositions can be formulated for use in various drug delivery systems. One or more physiologically acceptable excipients or carriers can also be included in the composition for appropriate formulation. Suitable formulations for use in the present invention can be found in Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, PA, 17th Edition, 1985. For a brief review of methods for drug delivery, see, for example, Langer (Science 249:1527-1533, 1990).

[0162] The pharmaceutical compositions are intended for parenteral, intranasal, topical, oral, or local administration, such as by transdermal means, for prophylactic and / or therapeutic treatment. The pharmaceutical compositions can be administered parenterally (e.g., by intravenous, intramuscular, or subcutaneous injection), or by oral ingestion, or by local application in a blood vessel or in an area affected by a cancerous condition, or by intraarticular injection. Additional routes of administration include intravascular, intraarterial, intratumoral, intraperitoneal, intraventricular, intradural, as well as intranasal, ophthalmic, intrascleral, intraorbital, rectal, topical, or aerosol inhalation administration. Sustained-release administration, such as by depot injection or erodible implant or component, is also specifically encompassed by the present invention. Thus, the present invention provides compositions for parenteral administration comprising the agent dissolved or suspended in an acceptable carrier, preferably an aqueous carrier, such as water, buffered water, saline, or PBS. The compositions may contain pharmaceutically acceptable auxiliary substances necessary to approximate physiological conditions, such as pH adjusting and buffering agents, tonicity adjusting agents, wetting agents, or surfactants, among others. The present invention also provides compositions for oral delivery, which may contain inactive ingredients such as binders or fillers for the formulation of unit dosage forms such as tablets or capsules. Furthermore, the present invention provides compositions for topical administration, which may contain inactive ingredients such as solvents or emulsifiers for the formulation of creams, ointments, gels, pastes, or eye drops.

[0163] These compositions can be sterilized by conventional sterilization techniques or sterile filtered. The resulting aqueous solutions can be packaged for immediate use or lyophilized, and the lyophilized preparations can be combined with a sterile aqueous carrier prior to administration. The pH of the preparations is typically 3 to 11, more preferably 5 to 9 or 6 to 8, and most preferably 6 to 7, e.g., 6 to 6.5. The resulting solid-form compositions can be packaged into multiple single-dose units, each containing a fixed amount of the agent or agents, such as in a sealed package of tablets or capsules. The solid-form compositions can also be packaged in flexible-volume containers, such as squeeze tubes designed for topically applicable creams or ointments.

[0164] A composition containing an effective amount can be administered for radiation treatment planning, diagnosis, or therapeutic treatment. When administered for radiation treatment planning or diagnostic purposes, the conjugate is administered to a subject in an amount effective to determine a diagnostically effective dose and / or a therapeutically effective dose. In therapeutic applications, the composition is administered to a subject (e.g., a human) already suffering from a condition (e.g., cancer) in an amount sufficient to cure or at least partially halt the symptoms of the disorder and its complications. An amount sufficient to achieve this purpose is defined as a "therapeutically effective amount," which is an amount of compound sufficient to substantially ameliorate at least one symptom associated with the disease or medical condition. For example, in the treatment of cancer, an agent or compound that reduces, prevents, delays, inhibits, or halts any symptoms of the disease or condition is therapeutically effective. A therapeutically effective amount of an agent or compound is not required to cure the disease or condition, but provides treatment for the disease or condition such that the onset of the disease or condition is delayed, hindered, or prevented, or the symptoms of the disease or condition are ameliorated, or the duration of the disease or condition is altered, or for example, made less severe, or recovery is promoted in the individual. The conjugates of the invention can be used to treat cancer by administering to a subject a first dose of either the conjugate or composition in an amount effective for a radiation treatment regimen, followed by a second dose of either the conjugate or composition in a therapeutically effective amount.

[0165] The effective amount for these uses may depend on the severity of the disease or condition and the weight and general condition of the subject. Those skilled in the art can determine the therapeutically effective amount of the compositions of the present invention and the methods of the present invention applied to mammals (e.g., humans), taking into account individual differences in the age, weight, and condition of the mammal. Because certain conjugates of the present invention exhibit enhanced ability to target and persist in cancer cells, the dose of the compound of the present invention may be less than the equivalent dose required for the therapeutic effect of the unconjugated drug (e.g., less than or equal to about 90%, 75%, 50%, 40%, 30%, 20%, 15%, 12%, 10%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1%). The agents of the present invention are administered to a subject (e.g., a mammal, such as a human) in an effective amount, which is an amount that produces the desired result in the treated subject.

[0166] Single or multiple administrations of the compositions of the invention comprising an effective amount can be carried out with the dose level and pattern selected by the treating physician. The dose and administration schedule can be determined and adjusted based on the severity of the disease or condition in the subject and monitored over the course of treatment according to methods commonly practiced by clinicians or others described herein.

[0167] The conjugates of the present invention can be used in combination with conventional treatment methods or therapies, or can be used separately from conventional treatment methods or therapies.

[0168] When compounds of the invention are administered in combination therapy with other agents, they may be administered sequentially or simultaneously to an individual. Alternatively, pharmaceutical compositions according to the invention may comprise a combination of a compound of the invention, as described herein, associated with a pharmaceutically acceptable excipient, and another therapeutic or prophylactic agent known in the art.

[0169] As used interchangeably herein, "antiproliferative" or "antiproliferative agent" refers to any anticancer agent, including those listed in Table 2, any of which may be used in combination with the conjugates of the present invention to treat the medical conditions described herein. Antiproliferative agents also include organoplatinum derivatives, naphthoquinone and benzoquinone derivatives, chrysophanic acid and its anthroquinone derivatives.

[0170] As used interchangeably herein, "immunomodulatory agent" or "immunomodulatory agent" refers to any immune modulator, including those listed in Table 2, any of which may be used in combination with the conjugates of the invention to treat the medical conditions described herein.

[0171] As used herein, "radiosensitizer" includes any agent that increases the sensitivity of cancer cells to radiation therapy. Radiation sensitizers include, but are not limited to, 5-fluorouracil, platinum analogs (e.g., cisplatin, carboplatin, oxaliplatin), gemcitabine, EGFR antagonists (e.g., cetuximab, gefitinib), farnesyltransferase inhibitors, COX-2 inhibitors, bFGF antagonists, and VEGF antagonists.

[0172] [Table 2A]

[0173] [Table 2B]

[0174] [Table 2C]

[0175] [Table 2D]

[0176] [Table 2E]

[0177] [Table 2F]

[0178] [Table 2G]

[0179] The following examples are intended to illustrate the synthesis of a representative number of conjugates and the use of these conjugates for the treatment of cancer. As such, the examples are intended to illustrate, rather than limit, the invention. Additional compounds not specifically exemplified can be synthesized using conventional methods in combination with the methods described herein. [Example]

[0180] Example 1 General Materials and Methods The antibody used was HuMIgG (Aldrich, I4506). Lutetium-177 was received from Perkin Elmer as lutetium chloride in 0.05N hydrochloric acid solution.

[0181] Analytical HPLC-MS was performed using a Waters Acquity HPLC-MS system containing a Waters Acquity Binary Solvent Manager, a Waters Acquity Sample Manager (samples cooled to 10°C), a Water Acquity Column Manager (column temperature 30°C), a Waters Acquity Photodiode Array Detector (monitoring at 254 nm and 214 nm), a Waters Acquity TQD with electrospray ionization, and a Waters Acquity BEH C18, 2.1 x 50 (1.7 μm) column. Preparative HPLC was performed using a Waters HPLC system containing a Waters 1525 Binary HPLC pump, a Waters 2489 UV / Visible Detector (monitoring at 254 nm and 214 nm), and a Waters XBridge Prep phenyl or C18, 19 x 100 mm (5 μm) column.

[0182] HPLC elution method 1: Waters Acquity BEH C18 2.1 x 50 mm (1.7 μm) column; mobile phase A: HO (0.1% v / v TFA); mobile phase B: acetonitrile (0.1% v / v TFA); flow rate = 0.3 mL / min; initial = 90% A, 3–3.5 min = 0% A, 4 min = 90% A, 5 min = 90% A.

[0183] HPLC elution method 2: Waters XBridge Prep phenyl 19 × 100 mm (5 μm) column; mobile phase A: HO (0.1% v / v TFA); mobile phase B: acetonitrile (0.1% v / v TFA); flow rate = 10 mL / min; initial = 80% A, 13 min = 0% A.

[0184] HPLC elution method 3: Waters Acquity BEH C18 2.1 x 50 mm (1.7 μm) column; Mobile phase A: H2O (0.1% v / v TFA); Mobile phase B: acetonitrile (0.1% v / v TFA); Flow rate = 0.3 mL / min; Initial = 90% A, 8 min = 0% A, 10 min = 0% A, 11 min = 90% A, 12 min = 90% A.

[0185] HPLC elution method 4: Waters XBridge Prep C18 OBD 19 x 100 mm (5 μm) column; Mobile phase A: H2O (0.1% v / v TFA); Mobile phase B: acetonitrile (0.1% v / v TFA); Flow rate: 10 mL / min; Initial = 80% A, 3 min = 80% A, 13 min = 20% A, 18 min = 0% A.

[0186] HPLC elution method 5: Waters XBridge Prep C18 OBD 19 x 100 mm (5 μm) column; Mobile phase A: H2O (0.1% v / v TFA); Mobile phase B: acetonitrile (0.1% v / v TFA); Flow rate: 10 mL / min; Initial = 90% A, 3 min = 90% A, 13 min = 0% A, 20 min = 0% A.

[0187] HPLC elution method 6: Waters XBridge Prep C18 OBD 19 x 100 mm (5 μm) column; Mobile phase A: H2O (0.1% v / v TFA); Mobile phase B: acetonitrile (0.1% v / v TFA); Flow rate: 10 mL / min; Initial = 75% A, 13 min = 0% A, 15 min = 0% A.

[0188] HPLC elution method 7: Waters XBridge Prep C18 OBD 19 x 100 mm (5 μm) column; Mobile phase A: H2O (0.1% v / v TFA); Mobile phase B: acetonitrile (0.1% v / v TFA); Flow rate: 10 mL / min; Initial = 80% A, 12 min = 0% A, 15 min = 0% A.

[0189] HPLC elution method 8: Waters XBridge Prep C18 OBD 19 x 100 mm (5 μm) column; Mobile phase A: H2O (0.1% v / v TFA); Mobile phase B: acetonitrile (0.1% v / v TFA); Flow rate: 10 mL / min; Initial = 90% A, 12 min = 0% A, 15 min = 0% A.

[0190] Analytical size-exclusion chromatography (SEC) was performed using a Waters system containing a Waters 1525 Binary HPLC pump, a Waters 2489 UV / Visible Detector (monitoring at 280 nm), a Bioscan Flow Count radiation detector (FC-3300), and a TOSOH TSKgel G3000SWxl, 7.8 x 300 mm column. The isocratic SEC method had a flow rate of 1 mL / min and used a mobile phase of 0.1 M phosphoric acid, 0.6 M NaCl, 0.025% sodium azide, pH 7.

[0191] MALDI-MS (positive ion) was performed using a MALDI Bruker Ultraflextreme Spectrometer.

[0192] Radioactive thin-layer chromatography (radio-TLC) was performed using a Bioscan AR-2000 Imaging Scanner and was carried out on iTLC-SG glass microfiber chromatography paper (Agilent Technologies, SGI0001) plates using citrate buffer (0.1 M, pH 5.5).

[0193] Example 2 Synthesis of 4-{[11-oxo-11-(2,3,5,6-tetrafluorophenoxy)undecyl]carbamoyl}-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl]butanoic acid (Compound B) The bifunctional chelate, 4-{[11-oxo-11-(2,3,5,6-tetrafluorophenoxy)undecyl]carbamoyl}-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl]butanoic acid (Compound B), was synthesized according to the scheme provided in Figure 2. To a solution of 5-(tert-butoxy)-5-oxo-4-(4,7,10-tris(2-(tert-butoxy)-2-oxoethyl)-1,4,7,10-tetraazacyclododecan-1-yl)pentanoic acid (DOTA-GA-(tBu)4, 50 mg, 0.07 mmol) in ACN (2.0 mL) was added DSC (50 mg, 0.21 mmol) followed by pyridine (0.20 mL, 2.48 mmol). The reaction was stirred at room temperature for 1 hour. To the reaction mixture was added 11-aminoundecanoic acid (70 mg, 0.36 mmol) followed by a PBS solution (1.0 mL) at room temperature. The reaction was stirred at room temperature for 72 hours. The reaction mixture was filtered using a syringe filter and directly purified by preparative HPLC using Method 6 to give intermediate 2-A (71 mg, 74.8%).

[0194] To a solution of intermediate 2-A (40 mg, 0.03 mmol), TFP (90 mg, 0.54 mmol), and EDC (40 mg, 0.27 mmol) in ACN (1.0 mL) was added pyridine (0.05 mL, 50 mg, 0.62 mmol) at room temperature. The solution was stirred at room temperature for 24 hours. The reaction was purified directly by preparative HPLC using Method 7 to provide intermediate 2-B (33 mg, 82.5%) as a wax after concentration using a Biotage V10 Rapid Evaporator.

[0195] Intermediate 2-B (33 mg, 0.022 mmol) was dissolved in DCM / TFA (1.0 mL / 2.0 mL) and allowed to stir at room temperature for 24 hours. The reaction was concentrated by airflow and directly purified by preparative HPLC using Method 8, which afforded Compound B (14 mg, 50.0%) as a clear wax after concentration. An aliquot was analyzed by HPLC-MS elution method 3; retention time: 4.15 min; MS (positive ESI); found m / z 808.1 [M+H]. + ; C 36 H 54 F4N5O 11 (Calculated value 808.8).

[0196]

number

[0197] Example 3 Synthesis of 4-{[2-(2-{2-[3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propoxy]ethoxy}ethoxy)ethyl]carbamoyl}-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl]butanoic acid (Compound C) The bifunctional chelate, 4-{[2-(2-{2-[3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propoxy]ethoxy}ethoxy)ethyl]carbamoyl}-2-[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl]butanoic acid (Compound C), was synthesized according to the scheme provided in Figure 3. To a solution of 5-(tert-butoxy)-5-oxo-4-(4,7,10-tris(2-(tert-butoxy)-2-oxoethyl)-1,4,7,10-tetraazacyclododecan-1-yl)pentanoic acid (DOTA-GA(tBu)4, 100 mg, 0.143 mmol) in ACN (8.0 mL) was added DSC (73 mg, 0.285 mmol) and pyridine (0.80 mL, 9.89 mmol). The reaction mixture was stirred at ambient temperature for 90 min. This solution was added to a half-solution of amino-PEG3-acid (63 mg, 0.285 mmol) in 1.2 mL of DMF in a 100 mL round-bottom flask. After 4 h at ambient temperature, the reaction was worked up by concentrating to dryness under a stream of air. The crude material was purified by HPLC elution method 2 (the crude material was dissolved in 6 mL of 20% ACN / HO). The product-containing fractions were pooled, concentrated under reduced pressure, and then co-evaporated with ACN (3 × 2 mL). Intermediate 1-A was obtained in 82% yield.

[0198] To a vial containing intermediate 1-A (82 mg, 60 μmol), ACN (2 mL), NEt (50 μL, 360 μmol, 6 equiv.), HBTU (23 mg, 60 μmol, 1 equiv.), and TFP solution (50 mg, 300 μmol, 5 equiv., dissolved in 250 μL of ACN) were added. The resulting clear solution was stirred at ambient temperature for 3 h. The reaction was worked up by concentrating to dryness under a stream of air, then diluted with ACN / HO (1:1, 3 mL total volume), and purified on preparative HPLC using elution method 4. Product-containing fractions were pooled, concentrated under reduced pressure, and coevaporated with ACN (3 × 2 mL). Intermediate 1-B was obtained as a clear residue (67 mg, 74% yield).

[0199] To a vial containing intermediate 1-B (67 mg, 64 μmol), DCM (2 mL) and TFA (2 mL) were added, and the resulting solution was stirred at ambient temperature for 16 h. Additional TFA (2 mL) was added, and the reaction was stirred at ambient temperature for 6 h. The reaction was concentrated to dryness under airflow, and the crude product was finally dissolved in ACN / HO (1 mL of 10% ACN / HO). The crude reaction solution was then purified by preparative HPLC using elution method 5. Fractions containing the product were pooled, frozen, and lyophilized. Compound C was obtained as a white solid (36 mg, 63% yield). An aliquot was analyzed by HPLC-MS elution method 3: retention time: 3.11 min; MS (positive ESI): found m / z 828.4 [M+H]. + ; C 34 H 50 F4N5O 14 (Calculated value 828.3).

[0200]

number

[0201] Example 4 [ 177 Synthesis of [Lu]-Compound A-Human IgG Compound A (1.34 μmol) was dissolved in sodium acetate buffer (20 μL, pH 6.5) and added to a solution containing antibody human IgG antibody (6.7 nmol) in bicarbonate buffer (pH 8.5). After 45 minutes at ambient temperature, the antibody conjugate product was purified by HPLC SEC column (1 mL / min, elution with acetate buffer (pH 6.5, 1 mM ascorbic acid)). MALDI-TOF-MS (positive ion): Compound A-human IgG: Found m / z 150360 [M+H] + Human IgG: Measured m / z 148339 [M+H] + .

[0202] In a typical reaction, Lu-177 (1.1 mCi, 5 μL) was added to a solution of Compound A-human IgG (90 μg in acetate buffer (pH 6.5) and ascorbic acid (1 μL, 0.1 M in acetate buffer (pH 6.5)). The radiolabeling reaction was incubated at 37° C. for 90 minutes. The crude product, [ 177 Lu]-Compound A-human IgG was purified by Sephadex G-50 resin-packed column and eluted with acetate buffer (pH 6.5, 1 mM ascorbic acid). Radio-TLC radiochemical purity: 98%; radiochemical yield: 45%; non-activity: 15.1 mCi / mg.

[0203] Example 5 [ 177 Synthesis of [Lu]-Compound B-Human IgG Compound B (1.17 μmol) was dissolved in sodium acetate buffer (0.117 mL, pH 6.5). An aliquot of Compound B solution (2 μL, 10 nmol) was added to a solution containing human IgG antibody (6.7 nmol) in bicarbonate buffer (pH 8.5). After 1 hour at ambient temperature, the antibody conjugate product was purified using a Sephadex G-50 resin-packed column. The antibody conjugate Compound A-human IgG was eluted from the column using acetate buffer (pH 6.5). MALDI-TOF-MS (positive ion): Compound B-human IgG observed m / z 149949 [M+H] + ; Human IgG measured value m / z 148540 [M+H] + .

[0204] In a typical reaction, Lu-177 (1.1 mCi, 5 μL) was added to a solution of Compound B-human IgG (100 μg in acetate buffer (pH 6.5) and ascorbic acid (1 μL, 0.1 M in acetate buffer (pH 6.5)). The radiolabeling reaction was incubated at 37° C. for 30 minutes. The crude product, 177Lu-Compound B-human IgG was purified by HPLC SEC column (1 mL / min, elution with acetate buffer (pH 6.5, 1 mM ascorbic acid)) and concentrated by ultrafiltration (Vivaspin, 10 kDa). Radio-TLC radiochemical purity: 98%; radiochemical yield: 51%; non-activity: 9.68 mCi / mg.

[0205] Example 6 [ 177 Synthesis of [Lu]-Compound C-Human IgG Compound C (0.96 μmol) was dissolved in sodium acetate buffer (95 μL, pH 6.5). An aliquot of Compound C solution (2 μL, 20 nmol) was added to a solution containing human IgG antibody (6.7 nmol) in bicarbonate buffer (pH 8.5). After 1 hour at ambient temperature, the antibody conjugate product was purified using a Sephadex G-50 resin-packed column. The antibody conjugate Compound C-human IgG was eluted from the column using acetate buffer (pH 6.5). MALDI-TOF-MS (positive ion): Compound C-human IgG observed m / z 150095 [M+H] + ; Human IgG measured value m / z 148540 [M+H] + .

[0206] In a typical reaction, Lu-177 (1.1 mCi, 5 μL) was added to a solution of Compound C-human IgG (100 μg in acetate buffer (pH 6.5) and ascorbic acid (1 μL, 0.1 M in acetate buffer (pH 6.5)). The radiolabeling reaction was incubated at 37° C. for 30 minutes. The crude product, [ 177 [Lu]-Compound C-human IgG was purified by HPLC SEC column (1 mL / min, eluted with acetate buffer (pH 6.5, 1 mM ascorbic acid)) and concentrated by ultrafiltration (Vivaspin, 10 kDa). Radio-TLC radiochemical purity: 98%; radiochemical yield: 37%; non-activity: 9.99 mCi / mg.

[0207] Example 7 Pharmacokinetic and metabolic studies of human IgG-based compounds Groups of four or five mice (normal CD-1 mice or athymic CD-1 nude mice) were intravenously injected with approximately 15 microcuries of radiolabeled test compound. Test compounds with various linkers were synthesized and radiolabeled with lutetium-177. For pharmacokinetic studies, animals were sacrificed at specific time points, and blood and tumors (if applicable) were analyzed for total radioactivity. For metabolic studies, animals were placed in metabolic cages (4-5 per cage) for collection of urine and feces every 24 hours up to 7 days. The radioactivity content of urine and fecal samples was quantified and converted to total urine or fecal output based on body weight. Excretion profiles for urine, feces, or total excretion (urine + feces) were generated by plotting the cumulative % injected dose (%ID) against time.

[0208] A non-targeting human IgG antibody was used for metabolic excretion studies to demonstrate that the change in radioactivity excretion profile directed by conjugation with linker Compound B and Compound C is a general process. The human IgG preparation used consisted of a purified mixture of all IgG isotypes (IgG1-4).

[0209] Non-targeting human IgG antibody conjugate [ 177 Lu]-Compound B-HuMIgG, and [ 177 The metabolic excretion profile of [Lu]-compound C-HuMIgG was 177 Lu]-Compound A-HuMIgG. 177 Lu]-Compound A-HuMIgG was excreted slowly, with only 13% of the injected dose (ID) cleared over 7 days via low-level urinary excretion. Compounds B and C directed distinct elimination pathways and increased the total excretion of radioactivity over 7 days when compared to Compound A-HuMIgG. 177 Lu]-compound B-HuMIgG is eliminated via feces and [ 177The elimination of [Lu]-Compound C-HuMIgG was roughly equally divided between urine and feces. Thus, while the linker type affected the route, rate, and extent of compound excretion (Figure 4), it was found not to alter the overall blood pharmacokinetics of the total radioactivity associated with the radioimmunoconjugate. We also observed that the improved elimination profile of Compound B or Compound C when conjugated to an antibody was a general and reproducible effect.

[0210] Example 8 [ 225 Ac]-Compound A-Human IgG Synthesis Compound A (1.34 μmol) was dissolved in sodium acetate buffer (20 μL, pH 6.5) and added to a solution containing human IgG antibody (6.7 nmol) in bicarbonate buffer (pH 8.5). After 45 minutes at ambient temperature, the antibody conjugate product was purified by HPLC SEC column (1 mL / min, elution with acetate buffer (pH 6.5, 1 mM ascorbic acid)). MALDI-TOF-MS (positive ion): Compound A-human IgG observed m / z 150360 [M+H] + ; Human IgG measured value m / z 148339 [M+H] + .

[0211] In a typical reaction, Ac-225 (1.1 mCi, 5 μL) is added to a solution of Compound A-human IgG (90 μg in acetate buffer (pH 6.5) and ascorbic acid (1 μL, 0.1 M in acetate buffer (pH 6.5)). The radiolabeling reaction is incubated for 90 minutes at ambient temperature (e.g., 20-25°C). The crude product, [ 225 Ac]-Compound A-human IgG was purified by a Sephadex G-50 resin-packed column and eluted with acetate buffer (pH 6.5, 1 mM ascorbic acid).

[0212] Example 9 [ 225 Ac]-Compound B-Human IgG Synthesis Compound B (1.17 μmol) was dissolved in sodium acetate buffer (0.117 mL, pH 6.5). An aliquot of Compound B solution (2 μL, 10 nmol) was added to a solution containing human IgG antibody (6.7 nmol) in bicarbonate buffer (pH 8.5). After 1 hour at ambient temperature, the antibody conjugate product was purified using a Sephadex G-50 resin-packed column. The antibody conjugate, Compound A-human IgG, was eluted from the column using acetate buffer (pH 6.5). MALDI-TOF-MS (positive ion): Compound B-human IgG observed m / z 149949 [M+H] + ; Human IgG measured value m / z 148540 [M+H] + .

[0213] In a typical reaction, Ac-225 (1.1 mCi, 5 μL) is added to a solution of Compound B-human IgG (100 μg in acetate buffer (pH 6.5) and ascorbic acid (1 μL, 0.1 M in acetate buffer (pH 6.5)). The radiolabeling reaction is incubated for 30 minutes at ambient temperature (e.g., 20-25°C). The crude product, [ 225 Ac]-Compound B-human IgG was purified by HPLC SEC column (1 mL / min, eluted with acetate buffer (pH 6.5, 1 mM ascorbic acid)) and concentrated by ultrafiltration (Vivaspin, 10 kDa).

[0214] Example 10 [ 225 Ac]-Compound C-Human IgG Synthesis Compound C (0.96 μmol) was dissolved in sodium acetate buffer (95 μL, pH 6.5). An aliquot of Compound C solution (2 μL, 20 nmol) was added to a solution containing human IgG antibody (6.7 nmol) in bicarbonate buffer (pH 8.5). After 1 hour at ambient temperature, the antibody conjugate product was purified using a Sephadex G-50 resin-packed column. The antibody conjugate, Compound C-human IgG, was eluted from the column using acetate buffer (pH 6.5). MALDI-TOF-MS (positive ion): Compound C-human IgG observed m / z 150095 [M+H] +; Human IgG measured value m / z 148540 [M+H] + .

[0215] In a typical reaction, Ac-225 (1.1 mCi, 5 μL) is added to a solution of Compound C-human IgG (100 μg in acetate buffer (pH 6.5) and ascorbic acid (1 μL, 0.1 M in acetate buffer (pH 6.5)). The radiolabeling reaction is incubated for 30 minutes at ambient temperature (e.g., 20-25°C). The crude product, [ 225 Ac]-Compound C-human IgG was purified by HPLC SEC column (1 mL / min, eluted with acetate buffer (pH 6.5, 1 mM ascorbic acid)) and concentrated by ultrafiltration (Vivaspin, 10 kDa).

[0216] Other embodiments While the invention has been described in relation to particular embodiments thereof, it will be understood that it can be further modified, and this application is generally intended to cover any variation, use, or adaptation of the invention in accordance with the principles of the invention, including such departures from the present disclosure as are within known or customary practice within the art to which this invention pertains and which can be applied to the essential features described above.

Claims

Claim 1: A compound of formula I: AL 1 -(L 2 ) n -B Formula I wherein A is a chelating moiety or a metal complex thereof; L 1 is optionally substituted C 1 -C 6 alkyl, optionally substituted C 1 -C 6 heteroalkyl, optionally substituted aryl or heteroaryl; B is a bridging group capable of covalently conjugating the AL 1 -(L 2 ) n - moiety in Formula I to a therapeutic or targeting moiety; n is 1 to 5; Each L 2 independently represents the structure: (-X 1 -L 3 -Z 1 -) Formula II wherein X 1 is C═O(NR 1 ), C═S(NR 1 ), OC═O(NR 1 ), NR 1 C═O(O), NR 1 C═O(NR 1 ), —CH 2 PhC═O(NR 1 ), —CH 2 Ph(NH)C═S(NR 1 ), O, or NR 1 , where R 1 is H or optionally substituted C 1 -C 6 alkyl or optionally substituted C 1 -C 6 heteroalkyl, optionally substituted aryl or heteroaryl; L 3 is an optionally substituted C 1 -C 50 alkylene, an optionally substituted C 1 -C 50 heteroalkylene, or a C 4 -C 20 polyethyleneoxide-ethylene; Z 1 is CH 2 , C═O, C═S, OC═O, NR 1 C═O, or NR 1 , where R 1 is hydrogen or optionally substituted C 1 -C 6 alkyl, or pyrrolidine-2,5-dione; Z 1 and B are directly bonded by a covalent bond, and L 1 is bonded to one atom constituting a ring in the chelating agent of A. or a pharmaceutically acceptable salt thereof.

2. The method of claim 1, wherein the chelating moiety is selected from the group consisting of DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), DOTMA ((1R,4R,7R,10R)-α,α',α'',α'''-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), DOTAM (1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane), DOTPA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrapropionic acid), DOAM-acetic acid (2-(4,7,10-tris(2-amino-2-oxoethyl)-1,4,7,10-tetraazacyclododecane-1-yl)-acetate), and DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid). (2,2',2''-(10-(2,6-dioxotetrahydro-2H-pyran-3-yl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid), DOTP (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylenephosphonic acid)), DOTMP (1,4,6,10-tetraazacyclodecane-1,4,7,10-tetramethylenephosphonic acid), DOTA-4AMP (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(acetamido-methylenephosphonic acid)), CB-TE2A (1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4,11-diacetic acid), NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid), NOTP (1,4,7-triazacyclononane-1,4,7-tri(methylenephosphonic acid)), TETPA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetrapropionic acid), TETA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid), HEHA (1,4,7,10,13,16-hexaazacyclohexadecane-1,4,7,10,13,16-hexaacetic acid), PEPA (1,4,7,10,13-pentaazacyclopentadecane-N,N',N'',N''',N''''-pentaacetic acid), H 4 Octapa (N,N'-bis(6-carboxy-2-pyridylmethyl)-ethylenediamine-N,N'-diacetic acid), H 2 Dedpa (1,2-[[6-(carboxy)-pyridin-2-yl]-methylamino]ethane), H 6 The compound of claim 1, which is phospa (N,N'-(methylenephosphonate)-N,N'-[6-(methoxycarbonyl)pyridin-2-yl]-methyl-1,2-diaminoethane), TTHA (triethylenetetramine-N,N,N',N'',N''',N'''-hexaacetic acid), DO2P (tetraazacyclododecanedimethanephosphonic acid), HP-DO3A (hydroxypropyltetraazacyclododecanetriacetic acid), EDTA (ethylenediaminetetraacetic acid), deferoxamine, DTPA (diethylenetriaminepentaacetic acid), DTPA-BMA (diethylenetriaminepentaacetic acid-bismethylamide), or porphyrin.

3. The chelating moiety of claim 1, wherein the chelating moiety has the structure: 【Chemical 1】 (In the formula, Y 1 is —CH 2 OCH 2 (L 2 ) n —B, C═O(L 2 ) n —B, or C═S(L 2 ) n —B and Y 2 is —CH 2 CO 2 H; or Y 1 is H and Y 2 is L 1 -(L 2 ) n -B).

3. The compound of claim 2, having the formula:

4. The compound of claim 2 or 3, wherein Y 1 is H.

5. The compound of claim 1, wherein X 1 is C═O(NR 1 ) and R 1 is H.

6. L 1 is a compound having the structure: 【Chemistry 2】 (wherein R 2 is hydrogen or —CO 2 H).

2. The compound of claim 1, having the formula:

7. The compound of any one of claims 1 to 6, wherein X 1 is C═O(NR 1 ), R 1 is H, and L 3 is C 4 to C 50 alkylene or C 4 to C 20 polyethyleneoxide-ethylene.

8. The compound of any one of claims 1 to 7, wherein Z 1 is CH 2 .

9. The metal of the metal complex is selected from Bi, Pb, Y, Mn, Cr, Fe, Co, Zn, Ni, Tc, In, Ga, Cu, Re, Sm, lanthanides, and actinides; or 2. The compound of claim 1, wherein the metal of the metal complex is a radionuclide selected from the group consisting of 47Sc, 55Co, 60Cu, 61Cu, 62Cu, 64Cu, 67Cu, 66Ga, 67Ga, 68Ga, 82Rb, 86Y, 87Y, 90Y, 97Ru, 99mTc, 105Rh, 109Pd, 111In, 117mSn, 149Pm, 149Tb, 153Sm, 177Lu, 186Re, 188Re, 199Au, 201Tl, 203Pb, 212Pb, 212Bi, 213Bi, 225Ac, and 227Th.

10. A compound according to any one of claims 1 to 9, wherein the metal is a radionuclide.

11. The compound of claim 10, wherein the radionuclide is 225 Ac, 111 In, or 68 Ga.

12. The compound of claim 1, wherein the therapeutic or targeting moiety is an antibody, an antigen-binding fragment thereof, or other targeting protein such as a nanobody, an affibody, and a consensus sequence derived from a type III fibronectin domain.

13. The compound of claim 12, wherein the antibody or antigen-binding fragment thereof specifically binds to insulin-like growth factor-1 receptor (IGF-1R).

14. The compound of claim 1, wherein the bridging group is an amino-reactive, methionine-reactive, or thiol-reactive bridging group.

15. The compound of claim 14, wherein the amino-reactive, methionine-reactive, or thiol-reactive bridging group comprises an activated ester such as a hydroxysuccinimide ester, a 2,3,5,6-tetrafluorophenol ester, a 4-nitrophenol ester, or an imidate, an anhydride, a thiol, a disulfide, a maleimide, an azide, an alkyne, a strained alkyne, a strained alkene, a halogen, a sulfonate, a haloacetyl, an amine, a hydrazide, a diazirine, a phosphine, a tetrazine, an isothiocyanate, or an oxaziridine.

16. The bridging group is 【Chemistry 3】 15. The compound of claim 14, selected from the group consisting of:

17. 【Catalog 4】 2. The compound of claim 1, wherein:

18. 【Chemical 5】 2. The compound of claim 1, wherein:

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