Chelated AAZTA conjugates and their complexes

Abstract

Chelate compounds of formula (I) or pharmaceutically acceptable salts thereof, and complexes thereof with metals or radioisotopes thereof. The present invention further relates to the preparation of such ligands and complexes, and their use as diagnostic or therapeutic agents.

Description

[Technical Field]

[0001] The present invention generally relates to the field of nuclear medicine (NM). In particular, the present invention relates to a novel conjugate of the chelating agent 6-amino-6-methylperhydro-1,4-diazepinetetraacetic acid with [Tyr3]octreotate / D-phenylalanyl-L-cysteinyl-L-tyrosyl-D-tryptophyl-L-lysyl-L-threonyl-L-cysteinyl-L-threonine cyclic disulfide (SSTR agonist), and its complex with metal or radioisotope. The present invention further relates to the preparation of such ligands and complexes, and their use as diagnostic or therapeutic agents. [Background technology]

[0002] Background of the Invention Radiopharmaceuticals are unique pharmaceutical preparations containing radioisotopes used in major clinical fields for diagnosis and / or therapy. For biological applications, metal radionuclides are commonly coordinated to chelators (ligands) for safe administration. Furthermore, the ligands typically used to construct radiometal-based radiopharmaceuticals are bifunctional chelators (BFCs) equipped with reactive functional groups that can be covalently attached (conjugated) to targeting vectors (e.g., peptides, nucleotides, antibodies, nanoparticles) to achieve target-specific activity, exhibiting affinity and selectivity for selected targets. In this way, upon injection into a patient, the chelator tightly binds the radiometal ion, allowing the target molecule to deliver the isotope without losing the radiometal from the radiopharmaceutical, effectively providing a site-specific radiation source in vivo for imaging or therapy (Price, Chem. Soc. Rev., 2014, 43, 260). The use of radiometals is advantageous due to their relatively short half-lives. 18 F or 11Compared with the lengthy (multi-step) synthesis required to introduce C into the molecular backbone, it lends itself to rapid one-step complexation for the preparation of diagnostic probes. Chelating agents are classified into open-chain / acyclic ones (EDTA (ethylenediaminetetraacetic acid), DTPA (diethylenediaminetetraacetic acid)) and their derivatives) and macrocyclic ones (NOTA (2-[4,7-bis(carboxymethyl)-1,4,7-triazonan-1-yl]acetic acid), DOTA ((1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid)) and their derivatives). Among the currently used chelating agents, DOTA is one of the mainstay ligands in radiometal chemistry, 68 Ga, 111 In, 177 Lu, 86 / 90 Y, 225 Ac, and 44 / 47DOTA is currently one of the "gold standards" for many isotopes, including Sc. DOTA is widely used, and two different products for nuclear medicine, NETSPOT® (Ga-68 DOTATATE) and LUTATHERA® (Lu-177 DOTATATE), have already been approved by the FDA and are sold by Advanced Accelerator Applications USA INC. (now a Novartis Company). Despite interesting advantages, such as the commercial availability of various bifunctional intermediates, DOTA complex formation is particularly slow and the reaction conditions are extreme, limiting its use to temperature-tolerant targeting moieties. DOTA's significant limitations have stimulated the development of new chelators at many facilities worldwide, and it is currently one of the most highly researched areas in the field of radiopharmaceuticals. Another drawback revealed by an analysis of the state of the art in this field relates to current radiolabeling methods. Radiolabeling of temperature-sensitive biologically active moieties (e.g., peptides, miniantibodies, antibodies, etc.) can be carried out, for example, in a two-step process. The first step is the labeling of the bifunctional chelate with a radioactive metal at high temperature, and the second step is the conjugation of the radioactive complex with a biologically active moiety at a lower temperature (e.g., ambient / room temperature) followed by purification. This two-step process can cause further loss of radioactivity. To overcome these difficulties, mainly 68For Ga, other chelating agents have been proposed for producing "kit-type" labeling reagents (e.g., DATA-TOC (see, e.g., Nock et al. Dalton Trans. 2017, 46(42), 14584-14590); or THP-TATE (see, e.g., Ma et al. EJNMMI Research, 2015, 5:52); or NODAGA-TATE (see, e.g., Velikyan, Bioconjugate Chem. 2008, 19, 2, 569-573); or THP-PSMA (see, e.g., Young, J Nucl Med., 2017, 58(8): 1270-1277)), which can be used to prepare other radiometal-based radiodiagnostic / radiotherapeutic agents at room temperature and near-physiological conditions, preferably in a one-step process.

[0003] AAZTA (6-amino-6-methylperhydro-1,4-diazepinetetraacetic acid) has the formula: [ka]

[0004] AAZTA and AAZTA derivatives are disclosed in, for example, International Patent Applications WO2003 / 008390, WO2006 / 136564, WO2006 / 002873, WO2013 / 135750 and WO2008 / 071679 (BRACCO Imaging SpA).Bifunctional AAZTA derivatives have been disclosed in many papers (for example, Nagy et al., Angew. Chem. Int. Ed., 2017; Sinnes et al., J Nucl Med, 2017; Sinnes et al., J Nucl Med, 2016; Pfister et al., EJNMMI Res., 2015, 5(1):74; Wu Z. et al., Nuclear Medicine and Biology, 43, 6, 2016, 360-371).

[0005] Gd and Gd chelators with several chelators such as AAZTA, DTPA, DOTA, and HP-DOA3 (2-[4,7-bis(carboxylatomethyl)-10-(2-hydroxypropyl)-1,4,7,10-tetrazacyclododec-1-yl]acetate). 68 The synthesis of novel chelates of Ga was investigated by Manzoni et al. (Manzoni et al. ChemMedChem, 7, 2012, 1084-1093) as potential MRI and PET tumor imaging probes in conjugation with conformationally optimized RGD sequences.

[0006] [Tyr3]octreotate / D-phenylalanyl-L-cysteinyl-L-tyrosyl-D-tryptophyl-L-lysyl-L-threonyl-L-cysteinyl-L-threonine cyclic disulfide (TATE) is a peptide represented by the following formula and is known as a somatostatin receptor (SSTR) agonist. [ka]

[0007] SSTRs are found at high densities in many malignancies, including gastroenteropancreatic neuroendocrine tumors (GEP-NETs), CNS, breast, lung, and lymphatic cancers, and at lower densities in renal, medullary thyroid, prostate, and colon cancers. The role of SSTR agonists in neuroendocrine tumors (NETs) is well characterized, and massive SSTR overexpression is observed in some NETs. SSTRs are proven NET targets / biomarkers (Kwekkeboom et al, Endocrine-Related Cancer, 2010, 17 R53-R73). Summary of the Invention [Problem to be solved by the invention]

[0008] Applicants have discovered novel peptide-conjugated AAZTA derivatives, including the AAZTA ligand and the molecular vector TATE. [Means for solving the problem]

[0009] As observed by applicants, metal complexes of the novel probe AAZTA-TATE have unexpectedly improved in vivo properties relative to the corresponding metal complexes of DOTA-TATE.

[0010] Applicants further observed that metal complexes prepared with the bioconjugates of the present invention, including AAZTA and TATE, have unexpectedly high kinetic inertness. In particular, the bioconjugate ligands of the present invention are unexpectedly stable compared to the corresponding non-bioconjugate complexes of AAZTA.

[0011] This novel conjugate and its metal complexes are particularly suitable as new tools for diagnosing and treating pathologies associated with SSTR expression.

[0012] Summary of the Invention One aspect of the present invention is a compound of formula (I): [ka] or a pharmaceutically acceptable salt thereof.

[0013] The present invention further relates to a metal complex comprising a chelate compound of formula (I) or a pharmaceutically acceptable salt thereof and a metal ion.

[0014] The compounds of formula (I) can form complexes with ions of metal atoms selected from: 43 Sc, 44 Sc, 44m Sc, 47 Sc, 51 Cr, 52 Fe, 52 Mn, 52m Mn, 55 Co, 58 Co, 60 Cu, 61 Cu, 62 Cu,64 Cu, 67 Cu, 67 Ga, 68 Ga, 72 As, 86 Y, 88 Y, 90 Y, 97 Ru, 99m Tc, 103 Ru, 105 Rh, 109 Pd, 111 In, 111 Ag, 112 Ag, 117m Sn, 140 La, 141 Ce, 142 Pr, 149 Pm, 149 Tb, 152 Tb, 155 Tb, 153 Sm, 159 Gd, 161 Tb, 165 Dy, 166 Dy, 166 Ho, 167 Tm, 168 Yb, 175 Yb, 177 Lu, 186 Re, 188 Re, 199 Au, 203 Pb, 212 Pb, 205 Bi, 206 Bi, 210 Bi, 211 Bi, 212 Bi, 213 Bi and 214 Bi.

[0015] The metal ion is preferably 44 Sc, 47 Sc, 68 Ga, 177 Lu, 212 Pb or 213 The ions are selected from Bi.

[0016] A further aspect of the invention relates to the use of the compounds as diagnostic imaging agents (eg, PET / SPECT imaging) or therapeutic agents (eg, alpha / beta / auger), particularly for the diagnosis or treatment of neuroendocrine tumors.

[0017] The present invention further relates to a method for preparing compounds of formula (I), comprising the synthesis of the compounds of formula (I) and their subsequent complexation with a selected metal ion. In general, when the metal ion is a radionuclide, this complexation can also be referred to as "radiolabeling."

[0018] According to another aspect, the present invention comprises, in particular 68 Ga or 44 A radiopharmaceutical composition suitable for PET / SPECT imaging comprising a compound of formula (I) when radiolabeled with Sc, or preferably a compound of formula (I) 213 Bi, 47 Sc, 177 Lu, 212 When radiolabeled with Pb, it relates to a radiopharmaceutical composition suitable for therapeutic purposes. DETAILED DESCRIPTION OF THE INVENTION

[0019] Detailed Description of the Invention In a first aspect, the present invention provides a compound of formula (I): [ka] or a pharmaceutically acceptable salt thereof.

[0020] Hereinafter, the compound of formula (I) above will be referred to as "AAZTA-TATE."

[0021] The preparation of AAZTA-TATE can be carried out using amide chemistry (for example, as detailed in the Examples), including the following steps: A. Solid-phase peptide synthesis of octapeptide (TATE) (Petersen, J. Control. Release, 160, 2012, 254-263); B. N-terminal acylation with 6-[bis[2-(1,1-dimethylethoxy)-2-oxoethyl]amino]-6-(5-carboxypentyl)tetrahydro-1H-1,4-diazepine-1,4(5H)-diacetic acid α,α'-bis(1,1-dimethylethyl) ester ((tBu)4-AAZTA-C4-COOH); C. Cleavage from the solid support and deprotection; D. Purification of the final compound by preparative HPLC.

[0022] Alternatively, AAZTA-TATE can be prepared by using other chemical routes readily available to one skilled in the art by using functional groups such as thioamides, esters, thioesters, ethers, thioethers, ureas, thioureas, triazoles, etc.

[0023] The compounds of formula (I) are suitable for forming metal complexes, for example, with metal elements having atomic numbers 20 to 31, 39, 42, 43, 44, 49, 57 to 83, or 43 Sc, 44 Sc, 44m Sc, 47 Sc, 51 Cr, 52 Fe, 52 Mn, 52m Mn, 55 Co, 58 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 67 Ga, 68 Ga, 72 As, 86 Y, 88 Y, 90 Y, 97 Ru, 99m Tc, 103 Ru, 105 Rh, 109 Pd, 111 In, 111 Ag, 112 Ag, 117m Sn, 140 La, 141 Ce, 142 Pr,149 Pm, 149 Tb, 152 Tb, 155 Tb, 153 Sm, 159 Gd, 161 Tb, 165 Dy, 166 Dy, 166 Ho, 167 Tm, 168 Yb, 175 Yb, 177 Lu, 186 Re, 188 Re, 199 Au, 203 Pb, 212 Pb, 205 Bi, 206 Bi, 210 Bi, 211 Bi, 212 Bi, 213 Bi and 214 It is possible to form a metal complex with an ion of a metal atom (abbreviated as "metal ion") selected from the group consisting of the respective radioisotope selected from Bi.

[0024] Preferably, the metal ions are ions of radioactive metal atoms. In this specification and claims, the expression radioactive metal atoms denotes radioactive forms of metal elements and is used interchangeably with the expressions radioactive metal, radionuclide, radioactive metal nuclide and radioisotope.

[0025] A further aspect of the present invention relates to the use of metal complexes of compounds of formula I as diagnostic imaging agents (e.g., PET / SPECT imaging) or therapeutic agents (e.g., alpha / beta / auger) particularly for the diagnosis or treatment of neuroendocrine tumors.

[0026] In particular, certain metal complexes of the present invention can be used as diagnostic imaging agents, preferably for in vivo diagnostic (eg, PET / SPECT) applications.

[0027] Examples of metal atoms suitable for in vivo diagnostic applications are: 43 Sc, 44m Sc,44 Sc, 52 Fe, 52 Mn, 52m Mn, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Ga, 68 Ga, 72 As, 86 Y, 99m Tc, 111 In, 152 Tb, 155 Tb, 199 Au and 203 Pb.

[0028] In a preferred embodiment, said metal complexes for in vivo diagnostic applications are used for the detection of neuroendocrine tumors.

[0029] Certain metal complexes of the present invention can be used as therapeutic agents, preferably in theranostic or radiotherapeutic (eg, beta / alpha) applications.

[0030] Examples of metal atoms suitable for therapeutic use are: 47 Sc, 67 Cu, 88 Y, 90 Y, 97 Ru, 103 Ru, 105 Rh, 109 Pd, 111 Ag, 112 Ag, 117m Sn, 140 La, 149 Pm, 149 Tb, 153 Sm, 161 Tb, 165 Dy, 166 Dy, 166 Ho, 167 Tm, 168 Yb, 175 Yb, 177 Lu, 186 Re, 188 Re, 199 Au, 205 Bi, 206 Bi,210 Bi, 211 Bi, 212 Bi, 212 Pb, 213 Bi and 214 Bi is selected from the group consisting of

[0031] In a preferred embodiment, said metal complexes for therapeutic use are used in the treatment of neuroendocrine tumors.

[0032] In certain embodiments of the present invention, the compounds of formula (I) are selected from transition metal and post-transition metal ( 68 The metal ions form metal complexes with metal ions selected from the group consisting of: 51 Cr, 52 Fe, 52 Mn, 52m Mn, 55 Co, 58 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 67 Ga, 103 Ru, 105 Rh, 109 Pd, 111 In, 186 Re, 188 Re, 203 Pb, 212 Pb, 205 Bi, 206 Bi, 210 Bi, 211 Bi, 212 Bi, 213 Bi and 214 The metal ion of the complex of the compound of formula (I) is preferably an ion of a metal atom selected from the group consisting of Ga, Co and Cu.

[0033] In other embodiments, the compound of formula (I) is 44 It forms a metal complex with an ion of a metal atom selected from the group of rare earth metals such as Sc. 43 Sc, 44m Sc, 47 Sc,86 Y, 88 Y, 90 Y, 140 La, 141 Ce, 142 Pr, 149 Pm, 149 Tb, 152 Tb, 155 Tb, 153 Sm, 159 Gd, 161 Tb, 165 Dy, 166 Dy, 166 Ho, 167 Tm, 168 Yb, 175 Yb, 177 Preferably, the metal ion of the complex of the compound of formula (I) is an ion of a metal atom selected from the group consisting of Sc, Y, La, Ce, Tb, Sm, Gd, Dy, Ho, Tm, Yb and Lu.

[0034] In a further embodiment, the compound of formula (I) is 213 It forms a metal complex with a metal ion selected from the group of soft metals such as Bi. 103 Ru, 105 Rh, 109 Pd, 186 Re, 188 Re, 203 Pb, 212 Pb, 205 Bi, 206 Bi, 210 Bi, 211 Bi, 212 Bi and 214 The metal ion of the complex of the compound of formula (I) is preferably a Bi or Pb ion.

[0035] In a particularly preferred embodiment, the compound of formula (I) is 44 Sc, 47 Sc, 68 Ga, 177 Lu, 212 Pb or 213 It forms a complex with a metal ion selected from Bi.

[0036] Compounds of formula (I) can be successfully labeled with "cold" or "hot" metals at room temperature with yields of over 90% in 5 minutes. As used herein, unless otherwise specified, the terms "cold" and "hot" metals refer to non-radioactive and radioactive metal ions, respectively.

[0037] Advantageously, the presence of the peptide TATE bound to AAZTA does not adversely affect the labeling process. Labeling can be performed in various buffer media, including NaOAc, NHOAc, or HEPES buffers of various molarities and pHs. The reaction temperature can range from room temperature (20°C) to 95°C, preferably from 20°C to 50°C. The reaction time can range from 1 to 60 minutes, preferably from 5 to 15 minutes.

[0038] The labeled compounds are stable in human plasma and DTPA solutions for several hours. 68 GaAAZTA-TATE is stable in DTPA solution at pH 4 and 7.4 for 2 hours (radiochemical purity (RCP) ≥ 98%). 44 ScAAZTA-TATE is stable (RCP ≥ 95%) for up to 24 hours at both pH levels.

[0039] Advantageously, high quantitative radiochemical yields (RCY) can be achieved with compounds of formula (I) by using relatively small amounts of ligand. Lower amounts of ligand for complete radiochelation result in tracers with high specific radioactivity. This increases tumor uptake, leading to superior imaging (more accurate quantification of receptor density at the tumor site, as well as the ability to identify tumors expressing very low receptor density) and treatment.

[0040] Complexes of compounds of formula (I) further exhibit significantly enhanced cellular internalization compared to their respective complexes with DOTA-TATE, as detailed in the in vitro examples.

[0041] Surprisingly, as shown in detail in the Examples, Applicants have found that metal complexes of AAZTA-TATE exhibit unexpectedly improved in vivo properties compared to complexes of DOTA-TATE with the same metals, e.g., metal complexes of AAZTA-TATE exhibited superior in vivo accumulation and superior in vivo imaging compared to metal complexes of DOTA-TATE.

[0042] Furthermore, as detailed in the Examples, Applicants have discovered that complexes of AAZTA-TATE exhibit unexpectedly greater kinetic inertness than the parent complex.

[0043] As used herein, given a metal complex of a compound of formula (I) in which the ligand is the bioconjugate AAZTA-TATE and the metal ion is M, the term "parent complex" refers to the corresponding non-targeted metal complex, which is a complex in which the ligand is AAZTA not conjugated to the target molecule TATE and the metal ion is M.

[0044] Comparison of dissociation half-life values ​​(t 1 / 2 =ln2 / k d , k d is the pseudo-first-order rate constant characterizing the dissociation of the complex), indicating that the kinetic inactivity of the AAZTA-TATE complex is higher than that of the corresponding non-targeted AAZTA-metal complex.

[0045] Thus, an embodiment of the present invention relates to metal complexes of the compound of formula (I) characterized by at least two-fold greater kinetic inertness relative to the corresponding non-target metal complexes of AAZTA.

[0046] A preferred embodiment of the present invention relates to metal complexes of the compound of formula (I) characterized by at least four-fold, preferably five-fold or more, greater kinetic inertness relative to the corresponding non-target metal complexes of AAZTA.

[0047] Generally, high kinetic inactivity indicates a low rate of dissociation in vivo, which reduces the risk associated with products of the dissociation reaction, such as free metal ions and free ligands.

[0048] In some embodiments, the metal complexes of AAZTA-TATE of the present invention surprisingly have higher conditional stability constants (pH=7.4, [Na + ]=0.15 M, 25 °C), which are generally at least two-fold higher than the corresponding complexes of AAZTA.

[0049] For example, the complexes Ga(AAZTA-TATE) and Bi(AAZTA-TATE) show associated increases in thermodynamic properties compared to their respective parent complexes of AAZTA.

[0050] Radiopharmaceutical metal complexes with AAZTA-TATE can be prepared using a radiolabeling kit that includes a vial containing the ligand (e.g., in lyophilized form, optionally in combination with suitable additives or excipients). The radionuclide can be eluted directly from a radionuclide generator (e.g., in the form of a chromatography column to which the radionuclide and each parent radiometal nuclide are adsorbed) using an elution solvent into a vial containing the ligand. The vial can be a single-dose vial or a multi-dose vial whose contents are divided into different syringes.

[0051] In a further embodiment, the present invention relates to a pharmaceutical composition comprising a chelating compound of formula I or a pharmaceutically acceptable salt thereof in combination with a pharmaceutically acceptable excipient.

[0052] In another aspect, the present invention relates to a radiopharmaceutical composition comprising an effective amount of a metal complex of a compound of formula (I) or a pharmaceutically acceptable salt thereof, and optionally one or more pharmaceutically acceptable excipients.

[0053] Details regarding dosages, dosage forms, modes of administration, and pharmaceutically acceptable excipients (e.g., carriers, diluents, adjuvants) are known in the art (see, e.g., Monograph from World Health Organization: Document QAS / 08.262 / FINAL, "RADIOPHARMACEUTICALS Final text for addition to The International Pharmacopoeia", November 2008).

[0054] In a preferred embodiment, the radiopharmaceutical composition of the metal complex of formula (I) can be administered by conventional parenteral modes, such as intravenous administration. For example, the pharmaceutical composition of the present invention is suitably formulated in an optionally buffered, isotonic, sterile aqueous solution for parenteral administration. Furthermore, radiochemical stabilizers, such as singlet oxygen / radical scavengers, such as ascorbic acid, gentisic acid, salicylic acid, etc., can be used to prevent radiolysis of the final product.

[0055] In this range, and unless otherwise specified, the term "effective amount" refers to any amount of the complex of Formula (I) of the present invention or a pharmaceutical composition thereof sufficient to fulfill the intended diagnostic or therapeutic purpose. The molar amount of the chelator or complex is generally on the order of nanomolar for diagnostic and / or therapeutic applications (e.g., 1-500 nmol, typically 5-50 nmol for diagnostics, and 50-250 nmol for beta therapy). Generally, for diagnostic applications, a radiation dose of 2-10 mCi / patient (Ci = Curie = 37 gigabecquerels (GBq)) is sufficient, for alpha therapy, 0.2-100 mCi / patient is sufficient, while for beta therapy, higher doses are required (e.g., 100-300 mCi / patient).

[0056] Both the ligand (i.e., AAZTA-TATE) and its metal complexes can exist in salt form, and in particular the carboxylic acid and amino groups (depending on the pH) can be salified with the preferred cations or anions of physiologically compatible bases or acids.

[0057] Thus, the present invention also relates to the chelate compounds of formula (I) and their metal complexes in the form of pharmaceutically acceptable salts.

[0058] As used herein, the term "pharmaceutically acceptable salt" refers to a compound of Formula I in which at least one of the carboxyl or amino groups is in ionic form (i.e., -COO - or -NH3 + , =NH2 + , ≡NH + ) and interacting with a corresponding counterion.

[0059] Preferred pharmaceutically acceptable salts are those in which the free carboxylic acid groups of the AAZTA ligand that are not involved in the chelation of a metal ion are salted with the corresponding cation, the actual number of said free carboxylic acid groups depending, for example, on the coordination number of the complexed metal ion and / or the pH of the solution containing the compound.

[0060] Alternatively, or in addition, the "pharmaceutically acceptable salts" may include derivatives of the compounds of the present invention in which the amine group of TATE is in the form of a salt with a corresponding anion.

[0061] Suitable cations that can be used to prepare salts of the complexes or ligands of the present invention can be inorganic or organic cations derived from either inorganic or organic bases. Examples of inorganic cations include, for example, alkali or alkaline earth metal ions such as potassium, sodium, calcium, or magnesium. Examples of organic cations include, for example, cations of primary, secondary, and tertiary amines such as ethanolamine, diethanolamine, morpholine, glucamine, N-methylglucamine, and N,N-dimethylglucamine.

[0062] Similarly, suitable anions that can be used to prepare salts of the complexes or ligands of the present invention can be either inorganic or organic anions derived from either inorganic or organic acids. Examples of anions of inorganic acids include ions of haloacids, such as chloride, bromide, or iodide, as well as other suitable ions such as sulfate or phosphate. Examples of anions of organic acids include those commonly used in the pharmaceutical field to prepare salts of basic substances, such as acetate, succinate, citrate, fumarate, maleate, or oxalate.

[0063] Examples of cations and / or anions of amino acids include, for example, those of glycine, lysine, arginine, ornithine, or aspartic acid and glutamic acid.

[0064] Thus, the preparation of compounds of formula (I), including their chelate complexes, either per se or in the form of physiologically acceptable salts, is a further object of the present invention.

[0065] The metal complexes of the present invention can be advantageously used as diagnostic imaging agents in PET or SPECT imaging methods or as targeted therapeutic agents (alpha / beta / auger), especially for the identification or treatment of neuroendocrine tumors.

[0066] The following examples will serve to further illustrate the invention. [Example]

[0067] Example 1: Synthesis of AAZTA-TATE 9-Fluorenylmethoxycarbonyl (Fmoc) amino acids and H-Thr-(tBu)-OH preloaded Wang resin were purchased from IRIS Biotech (Marktredwiz, Germany) and Novabiochem (Darmstadt, Germany). O-(7-Azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HATU) and 2,4,6-collidine were purchased from Sigma-Aldrich (Darmstadt, Germany). All solvents were purchased from VWR International (Radnor, USA) and used without further purification. 6-[bis[2-(1,1-dimethylethoxy)-2-oxoethyl]amino]-6-(5-carboxypentyl)tetrahydro-1H-1,4-diazepine-1,4(5H)-diacetic acid α,α'-bis(1,1-dimethylethyl) ester (AAZTA-C4-COOH tetra tert-butyl ester) was synthesized as described in Manzoni et al., ChemMedChem, 7, 2012, 1084-1093 (Supporting Information).

[0068] The immobilized linear octapeptide HD-Phe-Cys(Acm)-Tyr(tBu)-D-Trp(Boc)-Lys(Boc)-Thr(tBu)-Cys(Acm)-Thr(tBu)-Wang resin was synthesized automatically using the standard Fmoc protocol on Wang resin preloaded with H-Thr(tBu)-OH (115 mg, 0.87 mmol / g). Cleavage of the Fmoc group was achieved with 20% piperidine in DMF. After synthesis of the linear peptide, the resin was transferred to an SPPS manual reactor, and on-resin disulfide formation was carried out by adding Tl(CF3COO)3 (109 mg, 0.20 mmol, 2 equiv.) in DMF (2 mL) at room temperature over 75 min. The resin was washed thoroughly with DMF, and the N-terminus was acylated with 3.1, 2, and 2 equivalents (0.31, 0.20, and 0.2 mmol; 211, 134, and 134 mg, respectively) of 6-[bis[2-(1,1-dimethylethoxy)-2-oxoethyl]amino]-6-(5-carboxypentyl)tetrahydro-1H-1,4-diazepine-1,4(5H)-diacetic acid α,α'-bis(1,1-dimethylethyl) ester ((tBu)4-AAZTA-C4-COOH) in the presence of HATU (0.31, 0.2, and 0.2 mmol) and collidine (6.8, 4, and 4 equivalents) as a base (triple coupling). The first coupling was carried out overnight at room temperature at 38 rpm. The second and third couplings were carried out for 3 hours at room temperature at 38 rpm. After each coupling, the resin was washed thoroughly with DMF. The peptide was cleaved from the solid support by adding TFA / HO / TIS (95:2.5:2.5) overnight at room temperature. Final purification was performed using Atlantis prepD® C. 18Preparative HPLC was performed using an OBD 5 μm (19 × 100 mm) column. Eluents: (A) 0.1% TFA in H2O, (B) 0.1% TFA in CH3CN. Gradient profile: 25% B isocratic over 8.65 min, linear gradient from 25% to 45% B over 2.84 min, linear gradient from 45% to 100% B over 1.01 min. Flow rate: 20 mL / min. AAZTA-TATE isolated as a homogenous peak. The solvent was removed under vacuum, and the product was lyophilized from water to give a white solid (21 mg, 14% yield). Product purity was monitored by analytical HPLC using an Atlantis DC18 5 μm (4.6 × 150 mm) column with the same gradient profile and solvent mixture as above. Flow rate: 1 mL / min with UV detection at 230 nm. Purity: 99%. ESI-MS(m / z): Calculated value: C 67 H 91 N 13 O 21 S2 (M+2H) + 740.33 Actual value: 740.39.

[0069] Example 2: Radiolabeling experiments General conditions for radiolabeling experiments A suitable radio-HPLC method was developed to determine the radiochemical yield (RCY) of the reaction. Experiments were performed at 95°C (unless a specific radiolabel was performed at a lower temperature) in closed 1.5 mL Eppendorf tubes in a heating block (MyBlock, Sigma Aldrich) with a few drops of water in each cavity to facilitate uniform heating. 68 Ga is ITG 68 Ge / 68 The Ga generator was eluted with 0.05 M hydrochloric acid. The eluate was collected in 1 mL fractions. The fractions with the highest activity (100–120 MBq each) were used for labeling without further purification. 44Sc was obtained by irradiating 120–170 mg of natural calcium target (Sigma-Aldrich 441872, 99.99%) with a 16 MeV Ge PETtrace 800 cyclotron at a beam current of 30 μA for 30–60 min. The irradiated target material was dissolved in 3 M ultrapure hydrochloric acid and purified with 70 mg of DGA resin (Triskem DN-B25-S) and then with DOWEX (Sigma-Aldrich) resin according to the following protocol: The resin was loaded into a 1 mL disposable SPE syringe and washed with 2 mL of 3 M HCl, 1 M HNO3, 0.1 M HCl, and 3 M HCl. Calcium was removed by forcing the target solution through the resin, followed by 4 mL of 3 M HCl. To remove traces of iron and nickel, the resin was washed with 3 mL of 3 M HCl and 3 mL of 1 M HNO3 and eluted with 3 mL of 0.1 M HCl. 10–30 mg of DOWEX resin was washed with 1 mL of 3 M HCl and 5 mL of HO. The first 250–300 μL of the eluate from the DGA was discarded, and the next 2.7 mL was concentrated on the DOWEX resin, washed with 1 mL of HO, and eluted with 8 × 30 μL of 1 M pH=4 NHOAc buffer. The most active fraction was used for labeling experiments. Activity concentrations ranged from 1–3 GBq / mL. The resulting solution was tested for labeling efficiency with AAZTA-TATE.

[0070] Radio HPLC conditions Instrumentation: Waters Acquity I Class UPLC liquid chromatography (Berthold LB513 radioactivity detector with BSM, FTN, CM, PDA module, 20 μL MX flow cell). Peak areas are decay corrected using the first injection time of each measurement series. RCP% values ​​are calculated from the corrected area values. HPLC conditions: Column: Kinetex XBC18 3.6 μm, 4.6 × 50 mm UV wavelength:210;254nm Column temperature: 30℃ 68 Eluent for Ga labeling: A: 0.1% formic acid B:ACN:H2O (9:1) 44 Eluent for Sc labeling: A: 0.01M oxalic acid, pH 3 B:ACN:H2O (9:1) [Table 1]

[0071] 68 Ga and 44 Labeling experiments were performed to determine the optimal labeling conditions for AAZTA-TATE with Sc. All samples were run with and without the column to check for column activity loss. When formic acid was used as the eluent, the injected activity was collected and measured in a gamma counter to determine recovery. Peak area values ​​were decay-corrected to the time of the initial injection. Radiochemical purity values ​​were calculated from the corrected values.

[0072] sign In preliminary experiments, AAZTA-TATE was synthesized in ammonium acetate and HEPES buffers at pH 4 and pH 7 at 95°C. 44 Labeling with Sc was performed for 5 min. HEPES buffer at pH 4 provided the best results for quantitative labeling with 0.3 μM peptide. At room temperature, AAZTA-TATE could be labeled with over 90% yield in 5 min. 68 For Ga labeling, a higher peptide concentration (1 μM) and a longer reaction time (15–30 min) were required at room temperature; however, quantitative yields could be achieved after 5 min at 95 °C using the same precursor concentration. Based on these results, HEPES buffer at pH 4 at 95 °C was selected for preparation purposes in in vitro and in vivo experiments.

[0073] In order to obtain the highest specific activity and quantitative RCP% for both AAZTA-TATE and DOTA-TATE, 44 Labeling with Sc was performed.44 The specific radioactivity obtained with Sc-AAZTA-TATE labeling typically ranged from 85 to 110 GBq / µmol, whereas with DOTA-TATE, the specific radioactivity ranged from 8 to 20 GBq / µmol. The results demonstrate the superior labeling performance that can be achieved by using AAZTA-TATE instead of DOTA-TATE.

[0074] quality control Scandium labeling was monitored by HPLC on a Kinetex column using an oxalate-acetonitrile gradient without any signs of loss of activity. Preliminary experiments observed the formation of a mixed complex between gallium and the oxalate salt of AAZTA, which eluted as a broad peak with poor retention. Ga 3+ is adsorbed on the solid surface, and therefore the free 68 Ga 3+ The content cannot be quantified without the presence of a chelating agent. The addition of DTPA solution (0.01 M) was found to reduce the activity loss. The method was checked by collecting the HPLC eluate and measuring it in a gamma counter.

[0075] Stability of labeled compounds The stability of the labeled compound was determined in human plasma and DTPA solution. 68 GaAAZTA-TATE was stable in DTPA solution at pH 4 and 7.4 for 2 hours (RCP 98%). 44 ScAAZTA-TATE reached approximately 95% radiochemical purity in 24 h at both pH values.

[0076] Further plasma stability experiments were performed to determine the amount of radiometal released from the chelator at each time point. 68 2 hours for Ga; 44 In the case of Sc, the cells were incubated for 24 hours at 37°C in a sealed syringe containing human plasma. 3+ is adsorbed to plasma proteins and must be removed from the sample prior to HPLC analysis. 68 Ga3+ Washing the plasma precipitate was ineffective, resulting in the loss of Ga in solution. 3+ DTPA was added to plasma samples prior to protein precipitation to stabilize the DTPA. At each time point, 50 μL of sample was taken, mixed with 50 μL of 10 mM pH=4 DTPA solution, and incubated at 37°C for 10 minutes. The sample was mixed with 25 μL of cold 50% ethanol. 75 μL of cold acetonitrile was added, and the sample was centrifuged at 9000 RPM for 10 minutes. A 50 μL sample was taken from the supernatant and injected into the HPLC.

[0077] The plasma stability of both radiolabeled compounds is shown in Table 2. [Table 2] The results are: 68 It was shown that GaAAZTA-TATE showed a high RCP% (approximately 95% after 2 hours of plasma incubation). 44 In the case of ScAAZTA-TATE, the results show that the RCP% after 24 hours of incubation is approximately 96%. In conclusion, the results show that the compounds of the invention are characterized by high quantitative RCP after radiolabeling and are stable in plasma for a long period of time.

[0078] Example 3: In vitro cell experiments Cell Culture AR42J (ATCC® CRL-1492 TMThe ) somatostatin receptor-positive rat pancreatic exocrine tumor and human A2780 (receptor-negative human ovarian carcinoma) cell lines were purchased from the American Type Culture Collection (ATCC). LNCaP cells were cultured in RPMI-1640 medium (GIBCO Life Technologies) supplemented with 10% fetal bovine serum (FBS, GIBCO Life Technologies) and 1% antibiotic and antimycotic solution (Sigma-Aldrich). A2780 cells were cultured in Dulbecco's modified Eagle's medium (DMEM, GIBCO Life Technologies) supplemented with 10% FBS and 1% antibiotic and antimycotic solution. All cell lines were cultured at 37°C with 5% CO2. For in vitro experiments, cells were used at 85% confluence, and cell viability, as assessed by trypan blue exclusion, was always greater than 95%.

[0079] Cellular uptake experiments AR42J and A2780 cells were trypsinized, centrifuged, resuspended in DMEM, and diluted to 1 × 10 6 mL -1 The test tube was filled with 0.37MBq of 68 Ga or 44 The samples were incubated in the presence of Sc-labeled DOTA or AAZTA-TATE for 15, 30, 60, and 90 minutes at 37°C. After incubation, the samples were washed three times with ice-cold PBS and then washed with PBS. 68 Radioactivity was measured using a calibrated gamma counter (Perkin Elmer Wizard gamma counter) for 1 min within the Ga sensitive energy window. Decay-corrected radiotracer incorporation was calculated as counts / (min). ★ (10 6 The uptake was expressed as a percentage of the total radioactivity of the radiotracer added to the cells (%ID / 10 6 Each experiment was performed in triplicate, and the data shown represent the mean (±SD) of at least three independent experiments.

[0080] In vitro saturation binding experiments For in vitro saturation binding experiments, melanotic AR42J cells were used. Cells were plated in 24-well plates (5 × 10 per well). 4 ) for 24 hours. 44 Sc- or 68 Ga-AAZTA-TATE was added to each well in a volume of 200 μL. After 30 and 90 min of incubation (in a CO2 incubator at 37°C), the medium was removed, and the cells were washed twice with PBS, then twice with 0.2 M glycine, and lysed with 1 M NaOH at 37°C for 10 min.

[0081] To measure the ligand-receptor interaction, two different in vitro cell experiments (internalization and saturation binding) were performed.

[0082] 3a. 44 Sc-AAZTA-TATE and 44 Cellular uptake experiment of Sc-DOTA-TATE Internalization experiments measure receptor-mediated cellular uptake of radioligand. For this purpose, AR42J and A2780 cells were trypsinized, centrifuged, resuspended in DMEM, and diluted to 1 × 10. 6 mL -1 The tube was filled with 0.37MBq of 44 The cells were incubated in the presence of Sc-labeled DOTA- or AAZTA-TATE for 15, 30, 60, and 90 minutes at 37°C. After incubation, the samples were washed three times with ice-cold PBS, and radioactivity was measured in a calibrated gamma counter (Perkin Elmer Wizard gamma counter). Decay-corrected radiotracer incorporation was calculated as counts / (min). ★ (10 6 The uptake was expressed as a percentage of the total radioactivity of the radiotracer added to the cells (%ID / 10 6Each experiment was performed in triplicate, and the data shown represent the mean (±SD) of at least three independent experiments. Results are expressed as a percentage of the total radioactivity of the analyzed tracer (radioactive tracer added to cells) in AR42J and the control cell line A2780. 44 Sc-AAZTA-TATE or 44 Sc-DOTA-TATE, %ID / 10 6 The uptake is reported as a percentage of the total uptake in the target cells.

[0083] The results shown in Table 3 are 44 Compared with Sc-DOTA-TATE, the complex 44 showed significantly enhanced internalization of Sc-AAZTA-TATE. [Table 3]

[0084] 3b. In vitro saturation binding experiments In vitro saturation binding experiments measure the specific receptor-mediated uptake of radiolabeled ligand at equilibrium with increasing concentrations of radioligand. Melanotic AR42J cells were used for in vitro saturation binding experiments. Cells were plated in 24-well plates (5 × 10 cells per well). 4 ) for 24 hours. 44 Sc-AAZTA-TATE and 44 Sc-DOTA-TATE was added to each well in a volume of 200 μL. After 30 and 90 min of incubation (in a CO2 incubator at 37°C), the medium was removed, and the cells were washed twice with PBS, then twice with 0.2 M glycine, and lysed with 1 M NaOH at 37°C for 10 min. Saturation curves were generated plotting specific binding against the concentration of radioligand. Both ligands showed good internalization, especially the complexes. 44 The saturation curve for Sc-AAZTA-TATE reaches a plateau at higher amounts of bound species (Table 4a and Table 4b). [Table 4]

[0085] Example 4: In vivo imaging results In vivo uptake in AR42J (PET / MRI imaging) In vivo experiments were performed using tumor cells (tumor volume 125±10 mm 3 In vivo imaging experiments were performed 12±1 days after subcutaneous injection of AR42J pancreatic tumor-bearing male CB17 SCID mice (n=5). 44 Sc- or 68 Ga-AAZTA-TATE (injected radioactivity range: 14–22 MBq) or 44 Sc-DOTA-TATE (injected radioactivity range: 17-22 MBq) was injected intravenously via the lateral tail vein. 44 Blocking experiments were performed by injecting an excess amount of AAZTA-TATE before injecting sc-AAZTA-TATE (n = 3). An in vivo kinetic scan (0–90 min) was performed, followed by a 20-min static scan at 2.5 h. During imaging experiments, mice were anesthetized with 3–1.5% isoflurane (Forane) using a dedicated small animal anesthesia device. To determine the anatomical localization of organs and tissues, whole-body T1-weighted MRI scans were performed using a preclinical nanoScanPET / MRI system (Mediso Ltd., Hungary) (3D GRE EXT multi-FOV; TR / TE 15 / 2 ms; Phase: 100; FOV 55 mm; NEX: 2). PET volumes were reconstructed using the three-dimensional Ordered Subsets Expectation Maximization (3DOSEM) algorithm (Tera-Tomo, Mediso Ltd., Hungary). PET and MRI images were automatically co-registered by the nanoScan PET / MRI acquisition software (Nucline). Reconstructed images were then scanned using InterView TMImages were analyzed using FUSION (Mediso Ltd., Hungary) image analysis software. Ellipsoidal 3D regions of interest (VOIs) were manually drawn around the edges of the tissue or organ radioactivity by visual inspection. Radiotracer uptake was expressed as the standardized uptake value (SUV). SUV was calculated as follows: SUV = [VOI radioactivity (Bq / ml)] / [injected radioactivity (Bq) / animal body weight (g)] (assuming a density of 1 g / mL).

[0086] Table 5 shows the mean SUV values ​​for in vivo PET-MRI uptake experiments using cancer-bearing animals (n=5 / group, n=3 / blocking experiment) at 2.5 hours post-injection. [Table 5] 44 In vivo results for ScAAZTA-TATE demonstrate high uptake of this compound into tumor tissue. In fact, the mean SUV (2.74 ± 0.95) was 44 This is significantly higher than the mean value of ScDOTA-TATE (1.01 ± 0.47). Furthermore, blocking experiments of AAZTA-TATE demonstrated tumor uptake and sst receptor-expressing tumors. 44 The accumulation of Sc AAZTA-TATE was shown to be highly specific. In conclusion, the results showed that the compounds of the present invention are characterized by higher tumor tissue uptake than the corresponding DOTA compounds, and blocking experiments confirmed that the tumor uptake of the compounds of the present invention is highly specific.

[0087] Example 5: Evaluation of the stability and kinetic inactivity of the compound Ga-AAZTA-TATE 5a. Equilibrium experiment of Ga-AAZTA-TATE complex The thermodynamic properties of the Ga-AAZTA-TATE complex were evaluated using the conditional stability constant (logK cond =logK therm / (1+α H ), where α H =K1 H [H+ ]+K1 H K2 H [H + ] 2 +...+K1 H K2 H ...K n H [H + ] n and K1 H , K2 H ,...K n H is the protonation constant of the free ligand in 0.15 M NaCl solution at pH = 7.0). Based on the equilibrium properties of the Ga(AAZTA) complex, the Ga(AAZTA)OH species predominates under physiological conditions (Eq. (1), Baranyai, Eur. J. Inorg. Chem., 2013, 147-162). The conditional stability constant of the Ga(AAZTA)OH species is (logK cond =logK therm / ([OH - ](1+α H )) Following this analogy, it can be assumed that the Ga(AAZTA-TATE)OH complex also prevails under physiological conditions.

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[0088] 5b. Kinetics of the transchelation reaction of Ga(AAZTA-TATE)OH complex with human serum transferrin The ligand exchange reaction between Ga(AAZTA-TATE)OH and human serum apotransferrin (sTf, Sigma) was studied spectrophotometrically (Eq. (3)) using an Agilent 8453 UV-Vis spectrophotometer with a 1.0 cm cell at 246 nm and pH = 7.4, by the formation of Ga-transferrin complexes in the presence of excess Ga(AAZTA-TATE)OH to ensure pseudo-first-order kinetic conditions ([Ga(AAZTA-TATE)OH] = 50 μM, [Trf] = 8 and 16 μM) as described by Baranyai, Eur. J. Inorg. Chem., 2013, 147-162. The concentration of the human serum transferrin solution was determined by the molar extinction coefficient ε 280 =91200cm -1 M -1 (Takahashi, J. Biochem. 1989, 106, 858-863) was used to determine the absorbance at 280 nm. The temperature was maintained at 25°C, and the ionic strength and bicarbonate concentration of the sample (0.15 M for NaCl and 0.025 M for NaHCO₃) were kept constant.

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[0089] Example 6: Evaluation of the stability and kinetic inactivity of Sc(AAZTA-TATE) 6a. Equilibrium Experiment The thermodynamic properties of the Sc(AAZTA-TATE) complex are characterized by the conditional stability constant (logK cond =logK therm / (1+α H ), α H =K1 H [H + ]+K1 H K2 H [H + ] 2 +...+K1 H K2 H ...K n H [H + ] n and K1 H , K2 H ,...K n H is the protonation constant of the free ligand in 0.15 M NaCl solution at pH 7.0. The log K cond To determine the value, Sc 3+ The competitive reaction of AAZTA-TATE and NTA with Sc(NTA)2 complex (δ Sc =59.9 ppm) 45 It was investigated by following with Sc NMR spectroscopy (Bruker Avance III 400 spectrometer at 9.4 T) (Eq. (4), H3NTA = nitrilotriacetic acid, pH = 7.4, 25 °C, 0.15 M NaCl).

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[0090] To compare the equilibrium properties of Sc(AAZTA-TATE) with those of Sc(AAZTA), the conditional stability constant of Sc(AAZTA) is 3+ The competitive reaction of AAZTA and NTA with Sc(NTA)2 complex (δ Sc = 59.9 ppm) 45 The reaction mixture was determined by tracing with 1C NMR spectroscopy (Equation (5), pH = 7.4, 25°C, 0.15M NaCl).

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[0091] The amount of Sc(NTA)2 complex is 3+ The Sc concentration increases with increasing [NTA] due to the competitive reaction between AAZTA-TATE and AAZTA and NTA for Sc ions (Equations (4) and (5)). 3+ The competitive reactions of AAZTA-TATE and AAZTA with NTA for ions (Eqs. (4) and (5)) are ScTATE and K. ScAA It can be characterized by an equilibrium constant, which can be expressed as:

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[0092] NTA ([NTA] tot =[H x NTA]+2[Sc(NTA)2]), Sc 3+ ions ([Sc 3+ ] tot = [Sc(NTA)2] + [Sc(AAZTA-TATE)]) and AAZTA-TATE ([AAZTA-TATE] tot =[Sc(AAZTA-TATE)]+[H x AAZTA-TATE]) and K ScTATE The value of (Equation (6)) is Sc 3+ -AAZTA-TATE-NTA system 45 (K) calculated from the integrals of the Sc(NTA)2 complex in the Sc NMR spectrum.ScTATE =3.42(9)). The stability constant (logβ c(NTA)2 = 24.08, 0.15 M NaCl, 25 °C), the protonation constant of the NTA ligand (logK1 H =9.13(1), logK2 H =2.63(2), logK3 H = 1.64(3; 25°C, 0.15 M NaCl), the conditional stability constant of Sc(NTA)2 is log K cond Sc(NTA)2 = 20.63. ScTATE Equilibrium constant (K ScTATE =3.42) and the conditional stability constant of Sc(NTA)2 (logK cond Sc(NTA)2 = 20.63), the conditional stability constant (logK cond Sc( AAZTA-TATE ) =logK ScTATE +logK cond Sc(NTA)2 ) is the log K in 0.15 M NaCl solution at pH 7.4 and 25°C. cond Sc(AAZTA-TATE) =21.2.

[0093] NTA ([NTA] tot =[H x NTA]+2[Sc(NTA)2]), Sc 3+ ions ([Sc 3+ ] tot =[Sc(NTA)2]+[Sc(AAZTA)]) and AAZTA([AAZTA] tot =[Sc(AAZTA)]+[H x AAZTA]) and K ScAA The value of (Equation (7)) is Sc 3+ -AAZTA-NTA system 45 (K) calculated from the integrals of the Sc(NTA)2 complex in the Sc NMR spectrum. ScAA =70.3(5)). K ScAA Equilibrium constant (K ScAA=70.3) and the conditional stability constant of Sc(NTA)2 (logK cond Sc(NTA)2 = 20.63), the conditional stability constant (logK cond Sc( AAZTA ) =logK ScAA +logK cond Sc(NTA)2 ) is the log K in 0.15 M NaCl solution at pH 7.4 and 25°C. cond Sc( AAZTA ) = 22.5. Based on this evidence, the conditional stability constant of the Sc(AAZTA-TATE) complex is approximately 1 log K unit lower than that of the Sc(AAZTA) complex in 0.15 M NaCl solution at pH = 7.4 and 25 °C.

[0094] 6b. Kinetics of the transchelation reaction between Sc(AAZTA-TATE) and NTA ligand (NTA = nitrilotriacetic acid) The ligand exchange reaction between Sc(AAZTA-TATE) and NTA (Sigma) was carried out at pH 5.5 and 25°C. 45 The transchelation of Sc(AAZTA-TATE) was monitored by following the integral of the Sc(NTA)2 complex (Eq. (8)) formed during the reaction as described above.

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[0095] For kinetic experiments, two samples (2 × 0.8 mL samples) were prepared with 0.2 mM Sc(AAZTA-TATE) in the presence of 2500-fold and 5000-fold excess NTA ligand to ensure pseudo-first-order kinetic conditions. The pH was adjusted to 5.5 by stepwise addition of concentrated NaOH or HCl. Due to the high concentration of NTA present, no buffer was used to keep the pH constant. For the Sc(AAZTA-TATE)-NTA reaction system, 45In the Sc NMR spectrum, the transchelation reaction between Sc(AAZTA-TATE) and NTA results in an increase in the integral of the Sc(NTA)2 complex, as described in equation (8). The pseudo-first-order rate constant (k d ) was calculated by fitting the integral (Sc(NTA)2)-time data set to equation (9).

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[0096] Based on this similarity, it can be inferred that the mechanism of the transchelation reaction of Sc(AAZTA-TATE) is very similar to that of the parent Sc(AAZTA) complex (Nagy, Angew Chem Int Ed Engl 2017, 56, 2118-2122). The transchelation reaction of Sc(AAZTA-TATE) involves the rate-determining step, H + The dissociation of the Sc(AAZTA-TATE) complex assisted by 3+ This is followed by a rapid reaction between the exchanged NTA ligands. The pseudo-first-order rate constants (k) characterizing the transchelation reaction between Sc(AAZTA-TATE) and NTA were obtained at [Sc(AAZTA-TATE)] = 0.2 mM, [NTA] = 0.5 and 1.0 M, and pH = 5.50 and 5.35. d ) are (1.7±0.2)×10 -7 s -1 , (1.8±0.3)×10 -7 s -1 These kinetic data clearly demonstrate that [NTA] does not substantially affect the dissociation rate of Sc(AAZTA-TATE). The dissociation half-lives (t 1 / 2 =ln2 / k d )(Sc(AAZTA):t 1 / 2= 469 and 600 hours (Nagy, Angew Chem Int Ed Engl 2017, 56, 2118-2122); Sc(AAZTA-TATE):t 1 / 2 Comparison of the reaction times (=1100 and 1050 h, 25 °C) indicates that the kinetic inactivity of Sc(AAZTA-TATE) is approximately two-fold higher than that of Sc(AAZTA).

[0097] Example 7: Evaluation of the stability and kinetic inactivity of compound Bi-AAZTA-TATE 7a. Equilibrium experiment The stability constant of Bi(AAZTA) is 3+ The competitive reaction of AAZTA and NTA with Bi (Eq. (10)) was determined by UV spectrophotometry in the wavelength range of 210-350 nm. 3+ The concentrations of NTA and AAZTA were 30.2 μM and 10 mM, respectively, and the concentration of AAZTA was varied between 0 and 50 μM in 0.15 M NaClO4 (6 × 2 mL samples of previously prepared Bi(NTA)2 complexes to which AAZTA was added). The pH was adjusted to 7.4 by stepwise addition of concentrated NaOH or HClO4. To reach equilibrium, the samples were kept at 50 °C for 1 week and at 25 °C for 2 weeks (the time required to reach equilibrium was measured spectrophotometrically). Spectrophotometric measurements were performed using a PerkinElmer Lambda 365 UV-Vis spectrophotometer with a 1.0 cm cell at 25 °C. For equilibrium calculations, the protonation constants of the AAZTA and NTA ligands (AAZTA: log K1 H =10.29, logK2 H =6.51, logK3 H =3.86, logK4 H =1.94, logK5 H =1.00;NTA:logK1 H =9.22, logK2 H =2.98, logK3 H = 1.06; 25 °C, 0.15 M NaClO4) and the stability constants (log K Bi(NTA) =16.97, logβ Bi(NTA)2= 26.21, 25 °C, 0.15 M NaClO4) was determined as described in the literature (Baranyai, Eur. J. Inorg. Chem., 2013, 147-162; Karadakov, Talanta, 1970, 17, 883-887). Equilibrium constants were calculated using the program PSEQUAD (L. Zekany and I. Nagypal in PSEQUAD, Vol. (Ed. D. Leggett), Springer US, 1985, pp. 291-353).

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[0098] Furthermore, the thermodynamic properties of the Bi(AAZTA-TATE) complexes were evaluated using the conditional stability constants (logK cond =logK therm / (1+α H The log K of the Bi(AAZTA-TATE) complex was cond To determine the value of Bi 3+ The competitive reaction of AAZTA-TATE and NTA (Eq. (11)) with HCl was analyzed by capillary zone electrophoresis (Hewlett-Packard HP) in 0.15 M NaClO solution at pH 7.4 as disclosed by Chang, J. Chinese. Chem. Soc. 1999, 46, 519-528 (HNTA = nitrilotriacetic acid). 3D The equilibrium characterization of Bi(AAZTA-C4-TATE) was performed using a capillary electrophoresis system. 3+ The concentrations of NTA and AAZTA were 30.2 μM and 15.0 mM, respectively. The concentration of AAZTA-C4-TATE was varied between 0.0 and 50.4 μM in 0.15 M NaClO4 (5 × 1 mL samples of previously prepared Bi(NTA)2 complexes with AAZTA-C4-TATE). The pH was adjusted to 7.4 by the stepwise addition of concentrated NaOH or HClO4. To reach equilibrium, the samples were kept at 50 °C for 1 week and at 25 °C for 2 weeks (the time required to reach equilibrium was measured by capillary electrophoresis).

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[0099] Bi 3+ The amount of Bi(AAZTA-TATE) complex increases with increasing AAZTA-TATE amount according to the competitive reaction between AAZTA-TATE and NTA for NTA ions (Eq. (11)). tot=[H x NTA]+2[Bi(NTA)2]), Bi 3+ ion ([Bi 3+ ] tot =[Bi(NTA)2]+[Bi(AAZTA-TATE)]) and AAZTA-TATE([AAZTA-TATE] tot =[Bi(AAZTA-TATE)]+[H x AAZTA-TATE]) from the total concentration of K BiTATE The value was found to be 51 (9). The stability constant of Bi(NTA)2 and the protonation constant (logK1 H =9.22, logK2 H =2.98, logK3 H = 1.06; 25°C, 0.15M NaClO4), the conditional stability constant of Bi(NTA)2 is log K in 0.15M NaClO4 solution at pH 7.4 and 25°C. cond Bi(NTA)2 = 22.5. BiTATE Equilibrium constant (K Ga TATE = 51) and the conditional stability constant (log K cond Bi(NTA)2 = 22.5), the conditional stability constant (logK cond Bi(AAZTA-TATE) =logK BiTATE +logK cond Bi(NTA)2 ) is the log K in 0.15M NaClO4, pH = 7.4, and 25°C. cond Bi(AAZTA-TATE) =24.3. Based on this evidence, the conditional stability constant of the Bi(AAZTA-TATE) complex is approximately 0.8 log K units higher than that of the Bi(AAZTA) complex in 0.15 M NaCl4 solution at pH = 7.4 and 25 °C.

[0100] 7b. Kinetics of the transchelation reactions between Bi(AAZTA-TATE) and AAZTA ligand, and between Bi(AAZTA) and DTPA ligand The kinetic properties of Bi(AAZTA) were determined by following the transchelation reaction between Bi(AAZTA) and DTPA using UV spectrophotometry at 278 nm in 0.15 M NaClO4 solution at 25 °C (the stability constant of Bi(DTPA) is log K in 0.6 M NaClO4 at 25 °C). Bi(DTPA) = 29.3 (V. Kornev, A. Troubachev, Russ. J. Inorg. Chem, 1987, 32, 1419). In these experiments, the concentration of Bi(AAZTA) was 0.1 mM, and DTPA was applied in 10- and 20-fold excess to ensure pseudo-first-order conditions. The pH was adjusted to 8.5, 9.0, 9.5, 10.0, 10.5, and 11.0 by stepwise addition of concentrated NaOH or HClO4. To keep the pH value constant, 0.01 M N-methyl-piperazine (pH > 10) buffer was used. At pH > 10, OH - No buffer was used to maintain a constant pH due to the high concentration of . The pseudo-first-order rate constant (k d ) was calculated by fitting the absorbance-time data set for the Bi(AAZTA)-DTPA system to equation (9). The dissociation rate of the Bi(AAZTA) complex (k d ) is independent of [DTPA] and increases with increasing pH. - ] with increasing d The increase in value is due to the rate-determining step of OH - Dissociation of the Bi(AAZTA) complex assisted by β-AzTa (followed by free Bi 3+ This can be interpreted in terms of the k characterizing the dissociation reaction of Bi(AAZTA). d Rate constant and half-life (t 1 / 2 =ln2 / k d ) is k d =1.67×10 -6 s -1 , t 1 / 2 = 115 hours (pH = 9.0, 25°C, in 0.15M NaClO4).

[0101] To investigate the kinetic inactivity of the Bi(AAZTA-TATE) complex, the transchelation reaction between the Bi(AAZTA-TATE) complex and AAZTA (Eq. (12)) was investigated by capillary zone electrophoresis (Hewlett-Packard HP 3D The reaction was carried out by capillary electrophoresis (CES) in 0.15 M NaClO4 at pH 9.0 at 25°C in the presence of 20- and 40-fold excesses of AAZTA to ensure pseudo-first-order kinetic conditions.

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[0102] For kinetic experiments, two samples (2 × 1 mL samples in 0.15 M NaClO4 solution) were prepared with 50.1 μM Bi(AAZTA-TATE) in the presence of 20-fold and 40-fold excess AAZTA ligand to ensure pseudo-first-order kinetic conditions. The pH was adjusted to 9.0 by stepwise addition of concentrated NaOH or HClO4. 0.01 M N-methyl-piperazine (NMP) buffer was used to maintain a constant pH. The samples were maintained at 25 °C. In the electropherogram of the Bi(AAZTA-TATE)-AAZTA reaction system, the area of ​​the Bi(AAZTA-TATE) complex decreased as a function of time, indicating a transchelation reaction between the Bi(AAZTA-TATE) complex (Eq. 12) and the AAZTA ligand. The pseudo-first-order rate constant (k d ) was calculated by fitting the area of ​​the Bi(AAZTA-TATE) complex-time data set to equation (9). The pseudo-first-order rate constants (k d ) are (1.7±0.1)×10 -7 s -1 and (2.0±0.2)×10 -7 s -1 These kinetic data clearly demonstrate that [AAZTA] does not substantially affect the dissociation rate of Bi(AAZTA-TATE). The rate-determining step is the OH- The dissociation of the Bi(AAZTA-TATE) complex assisted by 3+ This is thought to be followed by a rapid reaction between the hydroxyl group and the exchanged AAZTA ligand.

[0103] The dissociation half-lives (t 1 / 2 =ln2 / k d )(Bi(AAZTA):t 1 / 2 = 115 hours; Bi(AAZTA-TATE):t 1 / 2 Comparison of the kinetic inactivation times (=118 h in 0.15 M NaClO4, pH=9.0, 25 °C) indicates that the kinetic inactivation of Bi(AAZTA-TATE) is approximately 10-fold higher than that of Bi(AAZTA) at near physiological conditions.

Claims

1. Formula (I): 【Chemical 1】 or a pharmaceutically acceptable salt thereof.

2. A metal complex comprising the chelate compound or a pharmaceutically acceptable salt thereof according to claim 1 and a metal ion.

3. The metal ions are 43 Sc, 44 Sc, 44m Sc, 47 Sc, 51 Cr, 52 Fe, 52 Mn, 52m Mn, 55 Co, 58 Co, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Cu, 67 Ga, 68 Ga, 72 As, 86 Y, 88 Y, 90 Y, 97 Ru, 99m Tc, 103 Ru, 105 Rh, 109 Pd, 111 In, 111 Ag, 112 Ag, 117m Sn, 140 La, 141 Ce, 142 Pr, 149 Pm, 149 Tb, 152 Tb, 155 Tb, 153 Sm, 159 Gd, 161 Tb, 165 Dy, 166 Dy, 166 Ho, 167 Tm, 168 Yb, 175 Yb, 177 Lu, 186 Re, 188 Re, 199 Au, 203 Pb, 212 Pb, 205 Bi, 206 Bi, 210 Bi, 211 Bi, 212 Bi, 213 Bi and 214 3. The metal complex according to claim 2, which is an ion of a metal atom selected from Bi.

4. The metal ions are 43 Sc, 44m Sc, 44 Sc, 52 Fe, 52 Mn, 52m Mn, 60 Cu, 61 Cu, 62 Cu, 64 Cu, 67 Ga, 68 Ga, 72 As, 86 Y, 99m Tc, 111 In, 152 Tb, 155 Tb, 199 Au and 203 3. The metal complex according to claim 2, which is an ion of a metal atom selected from Pb.

5. The metal ions are 47 Sc, 67 Cu, 88 Y, 90 Y, 97 Ru, 103 Ru, 105 Rh, 109 Pd, 111 Ag, 112 Ag, 117m Sn, 140 La, 149 Pm, 149 Tb, 153 Sm, 161 Tb, 165 Dy, 166 Dy, 166 Ho, 167 Tm, 168 Yb, 175 Yb, 177 Lu, 186 Re, 188 Re, 199 Au, 205 Bi, 206 Bi, 210 Bi, 211 Bi, 212 Bi, 212 Pb, 213 Bi and 214 3. The metal complex according to claim 2, which is an ion of a metal atom selected from Bi.

6. The metal ions are 44 Sc, 47 Sc, 68 Ga, 177 Lu, 212 Pb or 213 The metal complex according to claim 3, which is a Bi ion.

7. The metal ions are 44 Sc, 68 Ga or 213 The metal complex according to claim 6, which is a Bi ion.

8. Metal ions 68 The metal complex according to claim 7, which is a Ga ion.

9. Metal ions 44 The metal complex according to claim 7, which is an ion of Sc.

10. Metal ions 213 The metal complex according to claim 7, which is a Bi ion.

11. 11. The metal complex of any one of claims 2 to 10, wherein the pharmaceutically acceptable salt comprises a compound of formula I having at least one carboxylic acid group in ionic form and an inorganic or organic cation.

12. 11. The metal complex of any one of claims 2 to 10, wherein the pharmaceutically acceptable salt comprises a compound of formula I having at least one amino group in ionic form and an inorganic or organic anion.

13. 5. The metal complex of claim 4 for use as an imaging diagnostic agent for in vivo diagnosis (PET / SPECT).

14. 5. The metal complex of claim 4 for use in detecting neuroendocrine tumors.

15. 6. The metal complex of claim 5 for use as a therapeutic agent for theranostic or radiotherapy.

16. 6. The metal complex of claim 5 for use in the treatment of neuroendocrine tumors.

17. A pharmaceutical composition comprising the chelate compound of claim 1 in combination with a pharmaceutically acceptable excipient.

18. A pharmaceutical composition comprising a metal complex according to any one of claims 2 to 16 in combination with a pharmaceutically acceptable excipient.

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